bioRxiv Channel: Brown University

Stem cells in Nanomia bijuga (Siphonophora), a colonial animal with localized growth zones

Stefan Siebert — 2014-01-06

BackgroundSiphonophores (Hydrozoa) have unparalleled colony-level complexity, precision of colony organization, and functional specialization between zooids (i.e., the units that make up colonies) Previous work has shown that, unlike other colonial animals, most growth in siphonophores is restricted to one or two well-defined growth zones that are the sites of both elongation and zooid budding. It remained unknown, however, how this unique colony growth and development is realized at the cellular level.nnResultsTo understand the colony-level growth and development of siphonophores at the cellular level, we characterize the distribution of proliferating cells and interstitial stem cells (i-cells) in the siphonophore Nanomia bijuga. Within the colony we find that i-cells are present at the tip of the horn, the structure within the growth zone that gives rise to new zooids. They persist in the youngest zooid buds, but as each zooid matures i-cells become progressively restricted to specific regions within the zooids until they are mostly absent from the oldest zooids. I-cell marker-gene expression remained in gametogenic regions. I-cells are not found in the stem between maturing zooids. Domains of high cell proliferation include regions where i-cells can be found, but also include some areas without i-cells such as the stem within the growth zones. Cell proliferation in regions devoid of marker gene expression indicates the presence of mitotically active epithelial cell lineages and, potentially, progenitor cell populations.nnConclusionsRestriction of stem cells to particular regions in the colony may play a major role in facilitating the precision of siphonophore growth, and also lead to a reduced developmental plasticity in other, typically older, parts of the colony. This helps explain why siphonophore colonies have such precise colony-level organization.

Optimizing Real Time fMRI Neurofeedback for Therapeutic Discovery and Development

Luke Stoeckel — 2014-03-18

While reducing the burden of brain disorders remains a top priority of organizations like the World Health Organization and National Institutes of Health (BRAIN, 2013), the development of novel, safe and effective treatments for brain disorders has been slow. In this paper, we describe the state of the science for an emerging technology, real time functional magnetic resonance imaging (rtfMRI) neurofeedback, in clinical neurotherapeutics. We review the scientific potential of rtfMRI and outline research strategies to optimize the development and application of rtfMRI neurofeedback as a next generation therapeutic tool. We propose that rtfMRI can be used to address a broad range of clinical problems by improving our understanding of brain-behavior relationships in order to develop more specific and effective interventions for individuals with brain disorders. We focus on the use of rtfMRI neurofeedback as a clinical neurotherapeutic tool to drive plasticity in brain function, cognition, and behavior. Our overall goal is for rtfMRI to advance personalized assessment and intervention approaches to enhance resilience and reduce morbidity by correcting maladaptive patterns of brain function in those with brain disorders.

Automation and Evaluation of the SOWH Test with SOWHAT

Samuel H. Church — 2014-05-19

The Swofford-Olsen-Waddell-Hillis (SOWH) test is a method to evaluate incongruent phylogenetic topologies. It is used, for example, when an investigator wishes to know if the maximum likelihood tree recovered in their analysis is significantly different than an alternative phylogenetic hypothesis. The SOWH test compares the observed difference in likelihood between the topologies to a null distribution of differences in likelihood generated by parametric resampling. The SOWH test is a well-established and important phylogenetic method, but it can be difficult to implement and its sensitivity to various factors is not well understood. We wrote SOWHAT, a program that automates the SOWH test. In test analyses, we find that variation in parameter estimation as well as the use of a more complex model of parameter estimation have little impact on results, but that results can be inconsistent when an insufficient number of replicates are used to estimate the null distribution. We provide methods of analyzing the sampling as well as a simple stopping criteria for sufficient bootstrap replicates, which increase the overall reliability of the approach. Applications of the SOWH test should include explicit evaluations of sampling adequacy. SOWHAT is available for download from https://github.com/josephryan/SOWHAT.

Phylogenomic analyses of deep gastropod relationships reject Orthogastropoda

Felipe Zapata — 2014-07-10

Gastropods are a highly diverse clade of molluscs that includes many familiar animals, such as limpets, snails, slugs, and sea slugs. It is one of the most abundant groups of animals in the sea and the only molluscan lineage that has successfully colonised land. Yet the relationships among and within its constituent clades have remained in flux for over a century of morphological, anatomical and molecular study. Here we re-evaluate gastropod phylogenetic relationships by collecting new transcriptome data for 40 species and analysing them in combination with publicly available genomes and transcriptomes. Our datasets include all five main gastropod clades: Patellogastropoda, Vetigastropoda, Neritimorpha, Caenogastropoda and Heterobranchia. We use two different methods to assign orthology, subsample each of these matrices into three increasingly dense subsets, and analyse all six of these supermatrices with two different models of molecular evolution. All twelve analyses yield the same unrooted network connecting the five major gastropod lineages. This reduces deep gastropod phylogeny to three alternative rooting hypotheses. These results reject the prevalent hypothesis of gastropod phylogeny, Orthogastropoda. Our dated tree is congruent with a possible end-Permian recovery of some gastropod clades, namely Caenogastropoda and some Heterobranchia subclades.

Analysis of geographic patterns of molecular, morphological, and bioclimatic variation to evaluate hypotheses of species boundaries in the South American montane genus Escallonia (Escalloniaceae)

Felipe Zapata — 2014-09-30

Escallonia is a morphologically and ecologically diverse clade of shrubs and small trees widely distributed in three hotspots of plant diversity. Previous studies suggested that extant species may have radiated rapidly and/or recently resulting in complex patterns of molecular variation across this genus. This result is apparently mirrored in morphology because species also display complex and overlapping patterns of morphological variation. Taken together, these patterns call into question the identity of all species within Escallonia. To evaluate the currently proposed hypotheses of species boundaries, I used molecular, morphological, and bioclimatic a data for 35 species and assessed three species criteria: genealogical exclusivity, morphological gaps, and climatic niche differences. Interpreting these data in the context of species as segments of evolutionary lineages, I provide evidence that most species (ca. 70%) within Escallonia represent distinct species on independent evolutionary trajectories. Instead of rejecting the current hypotheses of species limit, I argue for taxonomic stability as it provides a useful framework for studies aiming to understand the mechanisms driving the origin and evolution of species in hotspots of biodiversity.

The Histology of Nanomia bijuga (Hydrozoa: Siphonophora)

Samuel H Church — 2014-10-29

The siphonophore Nanomia bijuga is a pelagic hydrozoan (Cnidaria) with complex morphological organization. Each siphonophore is made up of many asexually produced, genetically identical zooids that are functionally specialized and morphologically distinct. These zooids predominantly arise by budding in two growth zones, and are arranged in precise patterns. This study describes the cellular anatomy of several zooid types as well as of the stem and gas-filled float, called the pneumatophore. The distribution of cellular morphologies across zooid types enhances our understanding of zooid function. The unique absorptive cells in the palpon, for example, indicate specialized intracellular digestive processing in this zooid type. Furthermore, there are multiple areas of both endodermal and ectodermal epithelial complexity. Though cnidarians are usually thought of as mono-epithelial, we characterize at least two cellular populations in this species which are not connected to a basement membrane. This work provides a greater understanding of epithelial diversity within the cnidarians, and will be a foundation for future studies on Nanomia bijuga, including functional assays and gene expression analyses.

Phylogenomic analyses support traditional relationships within Cnidaria

Felipe Zapata — 2015-04-06

Cnidaria, the sister group to Bilateria, is a highly diverse group of animals in terms of morphology, lifecycles, ecology, and development. How this diversity originated and evolved is not well understood because phylogenetic relationships among major cnidarian lineages are unclear, and recent studies present contrasting phylogenetic hypotheses. Here, we use transcriptome data from 15 newly-sequenced species in combination with 26 publicly available genomes and transcriptomes to assess phylogenetic relationships among major cnidarian lineages. Phylogenetic analyses using different partition schemes and models of molecular evolution, as well as topology tests for alternative phylogenetic relationships, support the monophyly of Medusozoa, Anthozoa, Octocorallia, Hydrozoa, and a clade consisting of Staurozoa, Cubozoa, and Scyphozoa. Support for the monophyly of Hexacorallia is weak due to the equivocal position of Ceriantharia. Taken together, these results further resolve deep cnidarian relationships, largely support traditional phylogenetic views on relationships, and provide a historical framework for studying the evolutionary processes involved in one of the most ancient animal radiations.

Fast ventral stream neural activity enables rapid visual categorization

Maxime Cauchoix — 2015-04-13

AO_SCPLOWBSTRACTC_SCPLOWPrimates can recognize objects embedded in complex natural scenes in a glimpse. Rapid categorization paradigms have been extensively used to study our core perceptual abilities when the visual system is forced to operate under strong time constraints. However, the neural underpinning of rapid categorization remains to be understood, and the incredible speed of sight has yet to be reconciled with modern ventral stream cortical theories of shape processing.nnHere we recorded multichannel subdural electrocorticogram (ECoG) signals from intermediate areas (V4/PIT) of the ventral stream of the visual cortex while monkeys were actively engaged in detecting the presence or absence of animal targets in natural scenes. Using multivariate pattern analysis (MVPA) techniques, we quantified at millisecond precision task-relevant signals conveyed by ECoG data. Reliable neural decoding was possible shortly after stimulus onset from single trials with a degree of generalization to experimental manipulations closely mimicking monkeys accuracy and reaction time.nnTogether, the present study suggests that rapid ventral stream neural activity induces a selective task-relevant signal subsequently used to drive visual categorization.nnClassificationBiological sciences / Neuroscience

Bayesian Nonparametric Inference of Population Size Changes from Sequential Genealogies

Julia A Palacios — 2015-05-11

Sophisticated inferential tools coupled with the coalescent model have recently emerged for estimating past population sizes from genomic data. Accurate methods are available for data from a single locus or from independent loci. Recent methods that model recombination require small sample sizes, make constraining assumptions about population size changes, and do not report measures of uncertainty for estimates. Here, we develop a Gaussian process-based Bayesian nonparametric method coupled with a sequentially Markov coalescent model which allows accurate inference of population sizes over time from a set of genealogies. In contrast to current methods, our approach considers a broad class of recombination events, including those that do not change local genealogies. We show that our method outperforms recent likelihood-based methods that rely on discretization of the parameter space. We illustrate the application of our method to multiple demographic histories, including population bottlenecks and exponential growth. In simulation, our Bayesian approach produces point estimates four times more accurate than maximum likelihood estimation (based on the sum of absolute differences between the truth and the estimated values). Further, our methods credible intervals for population size as a function of time cover 90 percent of true values across multiple demographic scenarios, enabling formal hypothesis testing about population size differences over time. Using genealogies estimated with ARGweaver, we apply our method to European and Yoruban samples from the 1000 Genomes Project and confirm key known aspects of population size history over the past 150,000 years.

Sequencing ultra-long DNA molecules with the Oxford Nanopore MinION

John M Urban — 2015-05-13

Oxford Nanopore Technologies nanopore sequencing device, the MinION, holds the promise of sequencing ultra-long DNA fragments >100kb. An obstacle to realizing this promise is delivering ultra-long DNA molecules to the nanopores. We present our progress in developing cost-effective ways to overcome this obstacle and our resulting MinION data, including multiple reads >100kb.

CRISPR/Cas9-Assisted Transformation-Efficient Reaction (CRATER), a novel method for selective transformation

Lynn J. Rothschild — 2015-09-27

The CRISPR/Cas9 system has revolutionized genome editing by providing unprecedented DNA-targeting specificity. Here we demonstrate that this system can be applied to facilitate efficient plasmid selection for transformation as well as selective gene insertion into plasmid vectors by cleaving unwanted plasmid byproducts after restriction enzyme digestion and ligation. Using fluorescent and chromogenic proteins as reporters, we demonstrate that CRISPR/Cas9 cleavage excludes unwanted ligation byproducts and increases transformation efficiency of desired inserts from 20% up to 97% {+/-} 3%. This CRISPR/Cas9-Assisted Transformation-Efficient Reaction (CRATER) protocol is a novel, inexpensive, and convenient method for obtaining specific cloning products.

Quantifying the unobserved protein-coding variants in human populations provides a roadmap for large-scale sequencing projects

James Zou — 2015-11-06

Introduction Introduction Results Discussion 1. Preliminaries 2.1 Performance Guarantees 2. The UnseenEst Algorithm 3. Datasets 4. Validation experiments 5. Related works 6.1 Faithful samples 6.2 Chebyshev construction 6.3 Proof of Proposition... 6. Proofs of the... References References Recent efforts aggregating the genomes and exomes of tens of thousands of individuals have provided unprecedented insights into the landscape of rare human genetic variation1,2 and generated critical resources for clinical and population genetics. The recently announced U.S. Precision Med ...

pong: fast analysis and visualization of latent clusters in population genetic data

Aaron A Behr — 2015-11-14

1 MotivationA series of methods in population genetics use multilocus genotype data to assign individuals membership in latent clusters. These methods belong to a broad class of mixed-membership models, such as latent Dirichlet allocation used to analyze text corpora. Inference from mixed-membership models can produce different output matrices when repeatedly applied to the same inputs, and the number of latent clusters is a parameter that is often varied in the analysis pipeline. For these reasons, quantifying, visualizing, and annotating the output from mixed-membership models are bottlenecks for investigators across multiple disciplines from ecology to text data mining.nn2 ResultsWe introduce pong, a network-graphical approach for analyzing and visualizing membership in latent clusters with a native D3.js interactive visualization. pong leverages efficient algorithms for solving the Assignment Problem to dramatically reduce runtime while increasing accuracy compared to other methods that process output from mixed-membership models. We apply pong to 225,705 unlinked genome-wide single-nucleotide variants from 2,426 unrelated individuals in the 1000 Genomes Project, and identify previously overlooked aspects of global human population structure. We show that pong outpaces current solutions by more than an order of magnitude in runtime while providing a customizable and interactive visualization of population structure that is more accurate than those produced by current tools.nn3 Availabilitypong is freely available and can be installed using the Python package management system pip. pongs source code is available at https://github.com/abehr/pong.nn4 Contactaaron_behr@alumni.brown.edu,nnsramachandran@brown.edu

Reference-free deconvolution of DNA methylation data and mediation by cell composition effects

Eugene Andres Houseman — 2016-01-22

We propose a simple method for reference-free deconvolution that provides both proportions of putative cell types defined by their underlying methylomes, the number of these constituent cell types, as well as a method for evaluating the extent to which the underlying methylomes reflect specific types of cells. We have demonstrated these methods in an analysis of 23 Infinium data sets from 13 distinct data collection efforts; these empirical evaluations show that our algorithm can reasonably estimate the number of constituent types, return cell proportion estimates that demonstrate anticipated associations with underlying phenotypic data; and methylomes that reflect the underlying biology of constituent cell types. Thus the methodology permits an explicit quantitation of the mediation of phenotypic associations with DNA methylation by cell composition effects. Although more work is needed to investigate functional information related to estimated methylomes, our proposed method provides a novel and useful foundation for conducting DNA methylation studies on heterogeneous tissues lacking reference data.

Information-dependent Enrichment Analysis Reveals Time-dependent Transcriptional Regulation of the Estrogen Pathway of Toxicity

Salil N. Pendse — 2016-02-04

The twenty-first century vision for toxicology involves a transition away from high-dose animal studies and into in vitro and computational models. This movement requires mapping pathways of toxicity through an understanding of how in vitro systems respond to chemical perturbation. Uncovering transcription factors responsible for gene expression patterns is essential for defining pathways of toxicity, and ultimately, for determining chemical mode of action, through which a toxicant acts. Traditionally this is achieved via chromatin immunoprecipitation studies and summarized by calculating, which transcription factors are statistically associated with the up-and down-regulated genes. These lists are commonly determined via statistical or fold-change cutoffs, a procedure that is sensitive to statistical power and may not be relevant to determining transcription factor associations. To move away from an arbitrary statistical or fold-change based cutoffs, we have developed in the context of the Mapping the Human Toxome project, a novel enrichment paradigm called Information Dependent Enrichment Analysis (IDEA) to guide identification of the transcription factor network. We used the test case of endocrine disruption of MCF-7 cells activated by 17{beta} estradiol (E2). Using this new approach, we were able to establish a time course for transcriptional and functional responses to E2. ER and ER{beta} are associated with short-term transcriptional changes in response to E2. Sustained exposure leads to the recruitment of an additional ensemble of transcription factors and alteration of cell-cycle machinery. TFAP2C and SOX2 were the transcription factors most highly correlated with dose. E2F7, E2F1 and Foxm1, which are involved in cell proliferation, were enriched only at 24h. IDEA is, therefore, a novel tool to identify candidate pathways of toxicity, clearly outperforming Gene-set Enrichment Analysis but with similar results as Weighted Gene Correlation Network Analysis, which helps to identify genes not annotated to pathways.

Measuring epistasis in fitness landscapes: the correlation of fitness effects of mutations

Luca Ferretti — 2016-03-01

Genotypic fitness landscapes are constructed by assessing the fitness of all possible combinations of a given number of mutations. In the last years, several experimental fitness landscapes have been completely resolved. As fitness landscapes are high-dimensional, simple measures of their structure are used as statistics in empirical applications. Epistasis is one of the most relevant features of fitness landscapes. Here we propose a new natural measure of the amount of epistasis based on the correlation of fitness effects of mutations. This measure has a natural interpretation, captures well the interaction between mutations and can be obtained analytically for most landscape models. We discuss how this measure is related to previous measures of epistasis (number of peaks, roughness/slope, fraction of sign epistasis, Fourier-Walsh spectrum) and how it can be easily extended to landscapes with missing data or with fitness ranks only. Furthermore, the dependence of the correlation of fitness effects on mutational distance contains interesting information about the patterns of epistasis. This dependence can be used to uncover the amount and nature of epistatic interactions in a landscape or to discriminate between different landscape models.

Drosophila CLAMP is an essential protein with sex-specific roles in males and females

Jennifer A. Urban — 2016-03-08

Dosage compensation is a fundamental mechanism in many species that corrects for the inherent imbalance in X-chromosome copy number between XY males and XX females. In Drosophila melanogaster, transcriptional output from the single male X-chromosome is equalized to that of XX females by recruitment of the Male Specific Lethal (MSL) complex to specific sequences along the length of the X-chromosome. The initial recruitment of MSL complex to the X-chromosome is dependent on a recently discovered zinc finger protein called Chromatin-Linked Adapter for MSL Proteins (CLAMP). However, further studies on the in vivo function of CLAMP remained difficult because the location of the gene in pericentric heterochromatin made it challenging to create null mutations or deficiencies. Using the CRISPR/Cas9 genome editing system, we generated the first null mutant in the clamp gene that eliminates expression of CLAMP protein. We show that CLAMP is necessary for both male and female viability. While females die at the third instar larval stage, males die earlier, likely due to the essential role of CLAMP in male dosage compensation. Moreover, we demonstrate that CLAMP promotes dosage compensation in males and represses key male-specific transcripts involved in sex-determination in females. Our results reveal that CLAMP is an essential protein with dual roles in males and females, which together assure that dosage compensation is a sex-specific process.

Computational Pan-Genomics: Status, Promises and Challenges

Tobias Marschall — 2016-03-12

Many disciplines, from human genetics and oncology to plant breeding, microbiology and virology, commonly face the challenge of analyzing rapidly increasing numbers of genomes. In case of Homo sapiens, the number of sequenced genomes will approach hundreds of thousands in the next few years. Simply scaling up established bioinformatics pipelines will not be sufficient for leveraging the full potential of such rich genomic datasets. Instead, novel, qualitatively different computational methods and paradigms are needed. We will witness the rapid extension of computational pan-genomics, a new sub-area of research in computational biology. In this paper, we generalize existing definitions and understand a pan-genome as any collection of genomic sequences to be analyzed jointly or to be used as a reference. We examine already available approaches to construct and use pan-genomes, discuss the potential benefits of future technologies and methodologies, and review open challenges from the vantage point of the above-mentioned biological disciplines. As a prominent example for a computational paradigm shift, we particularly highlight the transition from the representation of reference genomes as strings to representations as graphs. We outline how this and other challenges from different application domains translate into common computational problems, point out relevant bioinformatics techniques and identify open problems in computer science. With this review, we aim to increase awareness that a joint approach to computational pan-genomics can help address many of the problems currently faced in various domains.

Novel Covariance-Based Neutrality Test of Time-Series Data Reveals Asymmetries in Ecological and Economic Systems

Alex Washburne — 2016-03-18

Systems as diverse as the interacting species in a community, alleles at a genetic locus, and companies in a market are characterized by competition (over resources, space, capital, etc) and adaptation. Neutral theory, built around the hypothesis that individual performance is independent of group membership, has found utility across the disciplines of ecology, population genetics, and economics, both because of the success of the neutral hypothesis in predicting system properties and because deviations from these predictions provide information about the underlying dynamics. However, most tests of neutrality are weak, based on static system properties such as species-abundance distributions or the number of singletons in a sample. Time-series data provide a window onto a systems dynamics, and should furnish tests of the neutral hypothesis that are more powerful to detect deviations from neutrality and more informative about to the type of competitive asymmetry that drives the deviation.nnHere, we present a neutrality test for time-series data. We apply this test to several microbial time-series and financial time-series and find that most of these systems are not neutral. Our test isolates the covariance structure of neutral competition, thus facilitating further exploration of the nature of asymmetry in the covariance structure of competitive systems. Much like neutrality tests from population genetics that use relative abundance distributions have enabled researchers to scan entire genomes for genes under selection, we anticipate our time-series test will be useful for quick significance tests of neutrality across a range of ecological, economic, and sociological systems for which time-series data are available. Future work can use our test to categorize and compare the dynamic fingerprints of particular competitive asymmetries (frequency dependence, volatility smiles, etc) to improve forecasting and management of complex adaptive systems.nnAuthor SummaryFrom fisheries and forestries to game parks and gut microbes, managing a community of organisms is much like managing a portfolio. Managers care about diversity, and calculations of risk - for extinction or financial ruin - require accurate models of the covariance between the parts of the portfolio.nnTo model the covariances in portfolios or communities, it helps to start simple with a null model assuming the equivalence of species or companies relative to one another (termed "neutrality") and letting the data suggest otherwise. Researchers in biology and finance have independently entertained and tested neutral models, but the existing tests have used snapshots of communities or the variance of fluctuations of individual populations, whereas tests of the covariances between species can better inform the development of alternative models.nnWe develop a covariance-based neutrality test for time-series data and use it to show that the human microbiome, North American birds, and companies in the S&P 500 all have a similar deviation from neutrality. Understanding and incorporating this non-neutral covariance structure can yield more accurate alternative models of community dynamics which can improve our management of "portfolios" of multi-species systems.

Membrane state diagrams make electrophysiological models simple

Robert Law — 2016-05-05

Ion channels are ubiquitous in living systems. Through interactions with membrane potential, ion channels both control metabolic events and mediate cell communication. Consequentially, membrane bioelectricity bears on fields ranging from cancer etiology to computational neuro-science. Conductance models have proven successful in quantitatively capturing these dynamics but are often considered difficult, with interpretation relegated to specialists. To facilitate research in membrane dynamics, especially in fields where roles for ion channels are just beginning to be quantified, we must make these models easy to understand.nnHere, we show that the membrane differential equation central to conductance models can be understood using simple circular geometry. The membrane state diagrams we construct are compact, faithful representations of conductance model state, designed to look like circular "cells" with currents flowing in and out. Every feature of a membrane state diagram corresponds to a physiological variable, so that insight taken from a diagram can be translated back to the underlying model. The construction is elementary: we convert conductances to angles subtended on the circle and potentials to radii; currents are then areas of the enclosed annular sectors.nnOur method clarifies a powerful but prohibitive modeling approach and has the potential for widespread use in both electrophysiological research and pedagogy. We illustrate how membrane state diagrams can augment traditional methods in the stability analysis of voltage equilibria and in depicting the Hodgkin-Huxley action potential, and we use the diagrams to infer the possibility of nontrivial fixed-voltage channel population dynamics by visual inspection rather than linear algebra.

Understanding How Microbiomes Influence the Systems they Inhabit: Insight from Ecosystem Ecology

Ed Hall — 2016-07-21

Translating the ever-increasing wealth of information on microbiomes (environment, host, or built environment) to advance the understanding of system-level processes is proving to be an exceptional research challenge. One reason for this challenge is that relationships between characteristics of microbiomes and the system-level processes they influence are often evaluated in the absence of a robust conceptual framework and reported without elucidating the underlying causal mechanisms. The reliance on correlative approaches limits the potential to expand the inference of a single relationship to additional systems and advance the field. We propose that research focused on how microbiomes influence the systems they inhabit should work within a common framework and target known microbial processes that contribute to the system-level processes of interest. Here we identify three distinct categories of microbiome characteristics (microbial processes, microbial community properties, and microbial membership) and propose a framework to empirically link each of these categories to each other and the broader system level processes they affect. We posit that it is particularly important to distinguish microbial community properties that can be predicted from constituent taxa (community aggregated traits) from and those properties that are currently unable to be predicted from constituent taxa (emergent properties). Existing methods in microbial ecology can be applied to more explicitly elucidate properties within each of these categories and connect these three categories of microbial characteristics with each other. We view this proposed framework, gleaned from a breadth of research on environmental microbiomes and ecosystem processes, as a promising pathway with the potential to advance discovery and understanding across a broad range of microbiome science.

Habits without Values

Kevin Miller — 2016-08-03

Habits form a crucial component of behavior. In recent years, key computational models have conceptualized habits as arising from model-free reinforcement learning (RL) mechanisms, which typically select between available actions based on the future value expected to result from each. Traditionally, however, habits have been understood as behaviors that can be triggered directly by a stimulus, without requiring the animal to evaluate expected outcomes. Here, we develop a computational model instantiating this traditional view, in which habits develop through the direct strengthening of recently taken actions rather than through the encoding of outcomes. We demonstrate that this model accounts for key behavioral manifestations of habits, including insensitivity to outcome devaluation and contingency degradation, as well as the effects of reinforcement schedule on the rate of habit formation. The model also explains the prevalent observation of perseveration in repeated-choice tasks as an additional behavioral manifestation of the habit system. We suggest that mapping habitual behaviors onto value-free mechanisms provides a parsimonious account of existing behavioral and neural data. This mapping may provide a new foundation for building robust and comprehensive models of the interaction of habits with other, more goal-directed types of behaviors and help to better guide research into the neural mechanisms underlying control of instrumental behavior more generally.

Opponent surrounds explain diversity of contextual phenomena across visual modalities

David A. Mely — 2016-08-22

AsbtractContext is known to affect how a stimulus is perceived. A variety of illusions have been attributed to contextual processing -- from orientation tilt effects to chromatic induction phenomena, but their neural underpinnings remain poorly understood. Here, we present a recurrent network model of classical and extra-classical receptive fields that is constrained by the anatomy and physiology of the visual cortex. A key feature of the model is the postulated existence of two spatially disjoint near-vs. far-extra-classical regions with complementary facilitatory and suppressive contributions to the classical receptive field. The model accounts for a variety of contextual illusions, reveals commonalities between seemingly disparate phenomena, and helps organize them into a novel taxonomy. It explains how center-surround interactions may shift from attraction to repulsion in tilt effects, and from contrast to assimilation in induction phenomena. The model further explains enhanced perceptual shifts generated by a class of patterned background stimuli that activate the two opponent extra-classical regions cooperatively. Overall, the ability of the model to account for the variety and complexity of contextual illusions provides computational evidence for a novel canonical circuit that is shared across visual modalities.

Fossils matter: improved estimates of divergence times in Pinus reveal older diversification

Bianca Saladin — 2016-09-04

BackgroundThe taxonomy of the genus Pinus is widely accepted and a well-resolved phylogeny based on entire plastome sequences exists. However, there is a large discrepancy in estimated divergence times of major pine clades among existing studies mainly due to differences in fossil placement and dating methods used. We currently lack a dated molecular pine phylogeny that makes full usage of the rich fossil record in pines. This study is the first to estimate the divergence dates of pines based on a large number of fossils (21) evenly distributed across all major clades in combination with applying the most novel dating method.nnResultsWe present a range of molecular phylogenetic trees of Pinus generated within a Bayesian framework using both the novel fossilized birth-death and the traditional node dating method with different fossil sets. We find the origin of pines likely to be up to 30 Myr older (Early Cretaceous) than inferred in most previous studies (Late Cretaceous) and propose generally older divergence times for major clades within Pinus than previously thought. Our age estimates vary significantly between the different dating approaches but the results generally agree on older divergence times. We present a revised list of 21 fossils that are suitable to use in dating or comparative analyses of pines.nnConclusionsAn accurate timescale for the divergence times in pines is essential if we are to link diversification processes and functional adaptation of this genus to geological events or to changing climates. Next to older divergence times in Pinus, our results indicate that node age estimates in pines depend on dating approaches and fossil sets used due to different inherent characteristics of dating approaches. Our set of dated phylogenetic trees of pines presented herein provide the basis to account for uncertainties in age estimations when applying comparative phylogenetic methods, which will improve our understanding of the evolutionary and ecological history in pines.

A comparative study of metagenomics analysis pipelines at the species level

Teo, Y. V. — 2016-10-15

Many metagenomics classification tools have been developed with the rapid growth of the metagenomics field. However, the classification of closely related species remains a challenge for this field. Here, we compared MetaPhlAn2, kallisto and Kraken for their performances in two metagenomics settings, human metagenomics and environmental metagenomics. Our comparative study showed that kallisto demonstrated higher sensitivity than MetaPhlAn2 and Kraken and better quantification accuracy than Kraken at the species level. We also showed that classification tools that run on full reference genomes misidentified many species that were not truly present. In order to reduce false positives, we introduced marker genes from MetaPhlAn2 into our pipeline, which uses kallisto for the classification step, as an additional filtering step for species detection.

Adaptive, arousal-related adjustments of perceptual biases optimize perception in a dynamic environment

Krishnamurthy, K. — 2016-10-26

Prior expectations can be used to improve perceptual judgments about ambiguous stimuli. However, little is known about if and how these improvements are maintained in dynamic environments in which the quality of appropriate priors changes from one stimulus to the next. Using a novel sound-localization task, we show that changes in stimulus predictability lead to arousal-mediated adjustments in the magnitude of prior-driven biases that optimize perceptual judgments about each stimulus. These adjustments depend on task-dependent changes in the relevance and reliability of prior expectations, which subjects update using both normative and idiosyncratic principles. The resulting variations in biases across task conditions and individuals are reflected in modulations of pupil diameter, such that larger stimulus-evoked pupil responses correspond to smaller biases. These results suggest a critical role for the arousal system in adjusting the strength of perceptual biases with respect to inferred environmental dynamics to optimize perceptual judgements.

Anchored Phylogenomics of Angiosperms I: Assessing the Robustness of Phylogenetic Estimates

Budenhagen, C. — 2016-11-08

An important goal of the angiosperm systematics community has been to develop a shared approach to molecular data collection, such that phylogenomic data sets from different focal clades can be combined for meta-studies across the entire group. Although significant progress has been made through efforts such as DNA barcoding, transcriptome sequencing, and whole-plastid sequencing, the community current lacks a cost efficient methodology for collecting nuclear phylogenomic data across all angiosperms. Here, we leverage genomic resources from 43 angiosperm species to develop enrichment probes useful for collecting ~500 loci from non-model taxa across the diversity of angiosperms. By taking an anchored phylogenomics approach, in which probes are designed to represent sequence diversity across the group, we are able to efficiently target loci with sufficient phylogenetic signal to resolve deep, intermediate, and shallow angiosperm relationships. After demonstrating the utility of this resource, we present a method that generates a heat map for each node on a phylogeny that reveals the sensitivity of support for the node across analysis conditions, as well as different locus, site, and taxon schemes. Focusing on the effect of locus and site sampling, we use this approach to statistically evaluate relative support for the alternative relationships among eudicots, monocots, and magnoliids. Although the results from supermatrix and coalescent analyses are largely consistent across the tree, we find support for this deep relationship to be more sensitive to the particular choice of sites and loci when a supermatrix approach as employed. Averaged across analysis approaches and data subsampling schemes, our data support a eudicot-monocot sister relationship, which is supported by a number of recent angiosperm studies.

Quantitative Analysis of Dopamine Neuron Subtypes Generated from Mouse Embryonic Stem Cells

Dingle, Y.-T. L. — 2016-12-13

Dopamine (DA) neuron subtypes modulate specific physiological functions and are involved in distinct neurological disorders. Embryonic stem cell (ESC) derived DA neurons have the potential to aid in the study of disease mechanisms, drug discovery, and possibly cell replacement therapies. DA neurons can be generated from ESCs in vitro, but the subtypes of ESC-derived DA neurons have not been investigated in detail despite the diversity of DA neurons observed in vivo. Due to cell culture heterogeneity, sampling methods applied to ESC-derived cultures can be ambiguous and potentially biased. Therefore, we developed a quantification method to capture the depth of DA neuron production in vitro by estimating the error associated with systematic random sampling. Using this method, we quantified calbindin+ and calretinin+ subtypes of DA neurons generated from mouse ESCs. We found a higher production of the calbindin+ subtype (11-27%) compared to the calretinin+ subtype (2-13%) of DA neuron; in addition, DA neurons expressing neither subtype marker were also generated. We then examined whether exogenous sonic hedgehog (SHH) and fibroblast growth factor 8 (FGF8) affected subtype generation. Our results demonstrate that exogenous SHH and FGF8 did not alter DA neuron subtype generation in vitro. These findings suggest that a deeper understanding DA neuron derivation inclusive of mechanisms that govern the in vitro subtype specification of ESC-derived DA neurons is required.nnNoteAll research was planned and conducted while members were at Brown UniversitynnResearch fundingNIH/NCRR/NIGMS RI Hospital COBRE Center for Stem Cell Biology (8P20GM103468-04) (MZ) Brown Institute for Brain Science Pilot Grant (4-63662) (MZ/DHK)

Chunking as a rational strategy for lossy data compression in visual working memory tasks.

Nassar, M. R. — 2017-01-06

The nature of capacity limits for visual working memory has been the subject of an intense debate that has relied on models that assume items are encoded independently. Here we propose that instead, similar features are jointly encoded through a "chunking" process to optimize performance on visual working memory tasks. We show that such chunking can: 1) facilitate performance improvements for abstract capacity-limited systems, 2) be optimized through reinforcement, 3) be implemented by center-surround dynamics, and 4) increase effective storage capacity at the expense of recall precision. Human performance on a variant of a canonical working memory task demonstrated performance advantages, precision detriments, inter-item dependencies, and trial-to-trial behavioral adjustments diagnostic of performance optimization through center-surround chunking. Models incorporating center-surround chunking provided a better quantitative description of human performance in our study as well as in a meta-analytic dataset, and apparent differences in working memory capacity across individuals were attributable to individual differences in the implementation of chunking. Our results reveal a normative rationale for center-surround connectivity in working memory circuitry, call for re-evaluation of memory performance differences that have previously been attributed to differences in capacity, and support a more nuanced view of visual working memory capacity limitations: strategic tradeoff between storage capacity and memory precision through chunking contribute to flexible capacity limitations that include both discrete and continuous aspects.

Pairwise comparisons are problematic when analyzing functional genomic data across species

Dunn, C. W. — 2017-02-09

AbstractThere is considerable interest in comparing functional genomic data across species. One goal of such work is to provide an integrated understanding of genome and phenotype evolution. Most comparative functional genomic studies have relied on multiple pairwise comparisons between species, an approach that does not incorporate information about the evolutionary relationships among species. The statistical problems that arise from not considering these relationships can lead pairwise approaches to the wrong conclusions, and are a missed opportunity to learn about biology that can only be understood in an explicit phylogenetic context. Here we examine two recently published studies that compare gene expression across species with pairwise methods, and find reason to question the original conclusions of both. One study interpreted pairwise comparisons of gene expression as support for the ortholog conjecture, the hypothesis that orthologs tend to be more similar than paralogs. The other study interpreted pairwise comparisons of embryonic gene expression across distantly related animals as evidence for a distinct evolutionary process that gave rise to phyla. In each study, distinct patterns of pairwise similarity among species were originally interpreted as evidence of particular evolutionary processes, but instead we find they reflect species relationships. These reanalyses concretely demonstrate the inadequacy of pairwise comparisons for analyzing functional genomic data across species. It will be critical to adopt phylogenetic comparative methods in future functional genomic work. Fortunately, phylogenetic comparative biology is also a rapidly advancing field with many methods that can be directly applied to functional genomic data.nnSignificanceComparisons of genome function between species are providing important insight into the evolutionary origins of diversity. Here we demonstrate that comparative functional genomics studies can come to the wrong conclusions if they do not take the relationships of species into account and instead rely on pairwise comparisons between species, as is common practice. We re-examined two previously published studies and found problems with pairwise comparisons that draw both their original conclusions into question. One study had found support for the ortholog conjecture and the other had concluded that the evolution of gene expression was different between animal phyla than within them. Our results demonstrate that to answer evolutionary questions about genome function, it is critical to consider evolutionary relationships.

Variability in fitness effects and the limitations of lineage selection

Graves, C. J. — 2017-02-11

Evolutionary biologists commonly assess the evolutionary advantage of an allele based on its effects on the lifetime survival and reproduction of individuals. However, alleles affecting traits like sex, evolvability, and cooperation can cause fitness effects that depend heavily on differences in the environmental, genetic, and social context of individuals carrying the allele. This variability makes it difficult to summarize the evolutionary fate of an allele based solely on its effects on any one individual. In this review we show how attempts to average over variability in the fitness effects of an allele can sometimes cause misleading results. We then describe a number of intriguing new evolutionary phenomena that have emerged in studies that explicitly model the fate of alleles that influence long-term lineage dynamics. We conclude with prospects for generalizations of population genetics theory and discuss how this theory might be applied to the evolution of infectious diseases.

Natural selection can favor the evolution of ratchet robustness over evolution of mutational robustness

Lan, Y. — 2017-03-27

The vast majority of fitness-affecting mutations are deleterious. How natural populations evolve to cope is a question of fundamental interest. Previous studies have reported the evolution of mutational robustness, that is, natural selection favoring populations with less deleterious mutations. By definition, mutational robustness provides a short-term fitness advantage. However, this overlooks the fact that mutational robustness decreases finite asexual populations ability to purge recurrent deleterious mutations. Thus, mutational robustness also results in higher risk of long-term extinction by Mullers ratchet. Here, we explore the tension between short- and long- term response to deleterious mutations. We first show that populations can resist the ratchet if either the selection coefficient or the ratio of beneficial to deleterious mutations increases as fitness declines. We designate these properties as ratchet robustness, which fundamentally reflects a negative feedback between mutation rate and the tendency to accumulate more mutations. We also find in simulations that populations can evolve ratchet robustness when challenged by deleterious mutations. We conclude that mutational robustness cannot be selected for in the long term, but it can be favored in the short-term, purely because of temporary fitness advantage. We also discuss other potential causes of mutational robustness in nature.

Just above chance: is it harder to decode information from human prefrontal cortex BOLD signals?

Bhandari, A. — 2017-04-13

Understanding the nature and form of prefrontal cortex representations that support flexible behavior is an important open problem in cognitive neuroscience. In humans, multi-voxel pattern analysis (MVPA) of fMRI BOLD measurements has emerged as an important approach for studying neural representations. An implicit, untested assumption underlying many PFC MVPA studies is that the base rate of decoding information from PFC BOLD activity patterns is similar to that of other brain regions. Here we estimate these base rates from a meta-analysis of published MVPA studies and show that the PFC has a significantly lower base rate for decoding than visual sensory cortex. Our results have implications for the design and interpretation of MVPA studies of prefrontal cortex, and raise important questions about its functional organization.

The rate of transient beta frequency events predicts impaired function across tasks and species

Shin, H. — 2017-04-20

Beta frequency oscillations (15-29Hz) are among the most prominent signatures of brain activity. Beta power is predictive of many healthy and abnormal behaviors, including perception, attention and motor action. Recent evidence shows that in non-averaged signals, beta can emerge as transient high-power "events". As such, functionally relevant differences in averaged power across time and trials can reflect accumulated changes in the number, power, duration, and/or frequency span of the events. We show for the first time that functionally relevant differences in averaged prestimulus beta power in human sensory neocortex reflects a difference in the number of high-power beta events per trial, i.e., the rate of events. Further, high power beta events close to the time of the stimulus were more likely to impair perception. This result is consistent across detection and attention tasks in human magnetoencephalography (MEG) and is conserved in local field potential (LFP) recordings of mice performing a detection task. Our findings suggest transient brain rhythms are best viewed as a "rate metric" in their impact on function, and provides a new framework for understanding and manipulating functionally relevant rhythmic events.

Alternative cleavage of a bone morphogenetic protein (BMP) produces ligands with distinct developmental functions and receptor preference

Anderson, E. N. — 2017-04-26

TGF-{beta} and Bone Morphogenetic Protein (BMP) family proteins are made as proprotein dimers, which are cleaved by proprotein convertases to release the active C-terminal ligand dimer. Multiple proteolytic processing sites in Glass bottom boat (Gbb), the Drosophila BMP7 ortholog, can produce distinct forms of active ligand. Cleavage at the S1 or atypical S0 site produces Gbb15, the conventional small BMP ligand, while cleavage at the NS site produces the larger Gbb38 ligand (1, 2). Here, we found that blocking NS cleavage increased association of the full length prodomain with Gbb15 resulting in a concomitant decrease in signaling activity. NS cleavage is required in vivo for Gbb-Decapentaplegic (Dpp) heterodimer-mediated wing vein patterning but not in cell culture to enable Gbb15-Dpp het-erodimer activity. Gbb NS cleavage is also required in vivo for the regulation of pupal ecdysis and viability that is dependent on the type II receptor Wishful thinking (Wit). We found that the ability of Gbb38 to signal requires the expression of either Wit or the type I receptor, Saxophone (Sax). Finally, we discovered that the production of Gbb38 in 3rd instar larvae results when processing at the S1/S0 site is blocked by O-linked glycosylation. Our findings demonstrate that BMP prodomain cleavage can ensure that the mature ligand is not inhibited by the prodomain. Furthermore, alternative processing of BMP proproteins produces ligand types that signal preferentially through different receptors and exhibit specific developmental functions.

Modeling Of Biomechanics And Biorheology Of Red Blood Cells In Type-2 Diabetes Mellitus

Chang, H.-Y. — 2017-04-28

Erythrocytes in patients with type-2 diabetes mellitus (T2DM) are associated with reduced cell deformability and elevated blood viscosity, which contribute to impaired blood flow and other pathophysiological aspects of diabetes related vascular complications. In this study, by using a two-component red blood cell (RBC) model and systematic parameter variation, we perform detailed computational simulations to probe the alteration of the biomechanical, rheological and dynamic behavior of T2DM RBCs in response to morphological change and membrane stiffening. First, we examine the elastic response of T2DM RBCs subject to static tensile forcing and their viscoelastic relaxation response upon release of the stretching force. Second, we investigate the membrane fluctuations of T2DM RBCs and explore the effect of cell shape on the fluctuation amplitudes. Third, we subject the T2DM RBCs to shear flow and probe the effects of cell shape and effective membrane viscosity on their tank-treading movement. In addition, we model the cell dynamic behavior in a microfluidic channel with constriction and quantify the biorheological properties of individual T2DM RBCs. Finally, we simulate T2DM RBC suspensions under shear and compare the predicted viscosity with experimental measurements. Taken together these simulation results and their comparison with currently available experimental data are helpful in identifying a specific parametric model the first of its kind that best describes the main hallmarks of T2DM RBCs, which can be used in future simulation studies of hematologic complications of T2DM patients.

Opportunities And Obstacles For Deep Learning In Biology And Medicine

Ching, T. — 2017-05-28

Deep learning, which describes a class of machine learning algorithms, has recently showed impressive results across a variety of domains. Biology and medicine are data rich, but the data are complex and often ill-understood. Problems of this nature may be particularly well-suited to deep learning techniques. We examine applications of deep learning to a variety of biomedical problems--patient classification, fundamental biological processes, and treatment of patients--and discuss whether deep learning will transform these tasks or if the biomedical sphere poses unique challenges. We find that deep learning has yet to revolutionize or definitively resolve any of these problems, but promising advances have been made on the prior state of the art. Even when improvement over a previous baseline has been modest, we have seen signs that deep learning methods may speed or aid human investigation. More work is needed to address concerns related to interpretability and how to best model each problem. Furthermore, the limited amount of labeled data for training presents problems in some domains, as do legal and privacy constraints on work with sensitive health records. Nonetheless, we foresee deep learning powering changes at both bench and bedside with the potential to transform several areas of biology and medicine.

Learning And Transfer Of Working Memory Gating Policies

Bhandari, A. — 2017-05-31

Abstract knowledge about the tasks we encounter enables us to rapidly and flexibly adapt to novel task contexts. Previous research has focused primarily on abstract rules that leverage shared structure in stimulus-response (S-R) mappings as the basis of such task knowledge. Here we provide evidence that working memory (WM) gating policies - a type of control policy required for internal control of WM during a task - constitute a form of abstract task knowledge that can be transferred across contexts. In two experiments, we report specific evidence for the transfer of selective WM gating policies across changes of task context. We show that this transfer is not tied to shared structure in S-R mappings, but instead in the dynamic structure of the task. Collectively, our results highlight the importance of WM gating policies in particular, and control policies in general, as a key component of the task knowledge that supports flexible behavior and task generalization.

Targeted Enrichment of Large Gene Families for Phylogenetic Inference: Phylogeny and Molecular Evolution of Photosynthesis Genes in the Portullugo (Caryophyllales)

Moore, A. — 2017-06-04

Hybrid enrichment is an increasingly popular approach for obtaining hundreds of loci for phylogenetic analysis across many taxa quickly and cheaply. The genes targeted for sequencing are typically single-copy loci, which facilitate a more straightforward sequence assembly and homology assignment process. However, single copy loci are relatively uncommon elements of most genomes, and as such may provide a biased evolutionary history. Furthermore, this approach limits the inclusion of most genes of functional interest, which often belong to multi-gene families. Here we demonstrate the feasibility of including large gene families in hybrid enrichment protocols for phylogeny reconstruction and subsequent analyses of molecular evolution, using a new set of bait sequences designed for the "portullugo" (Caryophyllales), a moderately sized lineage of flowering plants (~2200 species) that includes the cacti and harbors many evolutionary transitions to C4 and CAM photosynthesis. Including multi-gene families allowed us to simultaneously infer a robust phylogeny and construct a dense sampling of sequences for a major enzyme of C4 and CAM photosynthesis, which revealed the accumulation of adaptive amino acid substitutions associated with C4 and CAM origins in particular paralogs. Our final set of matrices for phylogenetic analyses included 75-218 loci across 74 taxa, with ~50% matrix completeness across datasets. Phylogenetic resolution was greatly improved across the tree, at both shallow and deep levels. Concatenation and coalescent-based approaches both resolve with strong support the sister lineage of the cacti: Anacampserotaceae + Portulacaceae, two lineages of mostly diminutive succulent herbs of warm, arid regions. In spite of this congruence, BUCKy concordance analyses demonstrated strong and conflicting signals across gene trees for the resolution of the sister group of the cacti. Our results add to the growing number of examples illustrating the complexity of phylogenetic signals in genomic-scale data.

Genetics of the Research Domain Criteria (RDoC): genome-wide association study of delay discounting

Sanchez-Roige, S. — 2017-06-07

Delay discounting (DD), which is the tendency to discount the value of delayed versus current rewards, is elevated in a constellation of diseases and behavioral conditions. We performed a genome-wide association study of DD using 23,127 research participants of European ancestry. The most significantly associated SNP was rs6528024 (P = 2.40 x 10-8), which is located in an intron of the gene GPM6B. We also showed that 12% of the variance in DD was accounted for by genotype, and that the genetic signature of DD overlapped with attention-deficit/hyperactivity disorder, schizophrenia, major depression, smoking, personality, cognition, and body weight.

Genome-wide association study of Alcohol Use Disorder Identification Test (AUDIT) scores in 20,328 research participants of European ancestry

Sanchez-Roige, S. — 2017-06-15

Genetic factors contribute to the risk for developing alcohol use disorder (AUD). In collaboration with the genetics company 23andMe, Inc., we performed a genome-wide association (GWAS) study of the Alcohol Use Disorder Identification Test (AUDIT), an instrument designed to screen for alcohol misuse over the past year. Our final sample consisted of 20,328 research participants of European ancestry (55.3% females; mean age = 53.8, SD = 16.1) who reported ever using alcohol. Our results showed that the chip-heritability of AUDIT score, when treated as a continuous phenotype, was 12%. No loci reached genome-wide significance. The gene ADH1C, which has been previously implicated in AUD, was among our most significant associations (4.4 x 10-7; rs141973904). We also detected a suggestive association on chromosome 1 (2.1 x 10-7; rs182344113) near the gene KCNJ9, which has been implicated in mouse models of high ethanol drinking. Using LD score regression, we identified positive genetic correlations between AUDIT score and AUD, high alcohol consumption, and cigarette smoking. We also observed an unexpected positive genetic correlation between AUDIT and educational attainment, and additional unexpected negative correlations with BMI/obesity and attention-deficit/hyperactivity disorder (ADHD). We conclude that conducting a genetic study using data from a population unselected for AUD and responding to an online questionnaire may represent a cost-effective strategy for elucidating the etiology of AUD.

Persistent homology demarcates a leaf morphospace

Li, M. — 2017-06-20

Current morphometric methods that comprehensively measure shape cannot compare the disparate leaf shapes found in seed plants and are sensitive to processing artifacts. We explore the use of persistent homology, a topological method applied across the scales of a function, to overcome these limitations. The described method isolates subsets of shape features and measures the spatial relationship of neighboring pixel densities in a shape. We apply the method to the analysis of 182,707 leaves, both published and unpublished, representing 141 plant families collected from 75 sites throughout the world. By measuring leaves from throughout the seed plants using persistent homology, a defined morphospace comparing all leaves is demarcated. Clear differences in shape between major phylogenetic groups are detected and estimates of leaf shape diversity within plant families are made. This approach does not only predict plant family, but also the collection site, confirming phylogenetically invariant morphological features that characterize leaves from specific locations. The application of a persistent homology method to measure leaf shape allows for a unified morphometric framework to measure plant form, including shape and branching architectures.

Sign of selection on mutation rate modifiers depends on population size

Raynes, Y. — 2017-06-28

The influence of population size (N) on natural selection acting on alleles that affect fitness has been understood for over half a century1. As N declines, genetic drift overwhelms selection and alleles with direct fitness effects are rendered neutral. Often, though, alleles experience so called indirect selection, meaning they affect not the fitness of an individual but the fitness distribution of its offspring. Some of the best studied examples of indirect selection include alleles that modify aspects of the genetic system such as recombination2 and mutation3 rates. Here we use analytics, simulations and experimental populations of S. cerevisiae to show that modifiers that increase the genomic mutation rate (mutators) are favored by indirect selection in large populations but become disfavored as N declines. This surprising phenomenon of sign inversion in selective effect demonstrates that indirect selection on a mutator exhibits a qualitatively novel dependence on N. Sign inversion may help understand the relatively sporadic distribution of mutators in nature despite their frequent emergence in laboratory populations. More generally, sign inversion may be broadly applicable to other instances of indirect selection, suggesting a previously unappreciated but critical role of population size in evolution.

SHP2 Is Required for BCR-ABL1-Induced Hematologic Neoplasms

Gu, S. — 2017-06-30

BCR-ABL1-targeting tyrosine kinase inhibitors (TKIs) have revolutionized treatment of Philadelphia chromosome-positive (Ph+) hematologic neoplasms. Nevertheless, acquired TKI resistance remains a major problem in chronic myeloid leukemia (CML), and TKIs are less effective against Ph+ B-cell acute lymphoblastic leukemia (B-ALL). GAB2, a scaffolding adaptor that binds and activates SHP2, is essential for leukemogenesis by BCR-ABL1, and a GAB2 mutant lacking SHP2 binding cannot mediate leukemogenesis. Using a genetic loss-of-function approach and bone marrow transplantation (BMT) models for CML and BCR-ABL1+ B-ALL, we show that SHP2 is required for BCR-ABL1-evoked myeloid and lymphoid neoplasia. Ptpn11 deletion impairs initiation and maintenance of CML-like myeloproliferative neoplasm, and compromises induction of BCR-ABL1+ B-ALL. SHP2, and specifically, its SH2 domains, PTP activity and C-terminal tyrosines, is essential for BCR-ABL1+, but not WT, pre-B cell proliferation. The MEK/ERK pathway is regulated by SHP2 in WT and BCR-ABL1+ pre-B cells, but is only required for the proliferation of BCR-ABL1+ cells. SHP2 is required for SRC family kinase (SFK) activation only in BCR-ABL1+ pre-B cells. RNAseq reveals distinct SHP2-dependent transcriptional programs in BCR-ABL1+ and WT pre-B cells. Our results suggest that SHP2, via SFKs and ERK, represses MXD3/4 to facilitate a MYC-dependent proliferation program in BCR-ABL1-transformed pre-B cells.

The influence of higher-order epistasis on biological fitness landscape topography

Weinreich, D. M. — 2017-07-18

The effect of a mutation on the organism often depends on what other mutations are already present in its genome. Geneticists refer to such mutational interactions as epistasis. Pairwise epistatic effects have been recognized for over a century, and their evolutionary implications have received theoretical attention for nearly as long. However, pairwise epistatic interactions themselves can vary with genomic background. This is called higher-order epistasis, and its consequences for evolution are much less well understood. Here, we assess the influence that higher-order epistasis has on the topography of 16 published, biological fitness landscapes. We find that on average, their effects on fitness landscape declines with order, and suggest that notable exceptions to this trend may deserve experimental scrutiny. We explore whether natural selection may have contributed to this finding, and conclude by highlight opportunities for further work dissecting the influence that epistasis of all orders has on the efficiency of natural selection.

Drosophila Kruppel homolog 1 represses lipolysis through interaction with dFOXO

Kang, P. — 2017-07-18

Transcriptional coordination is a vital process contributing to metabolic homeostasis. As one of the key nodes in the metabolic network, the forkhead transcription factor FOXO has been shown to interact with diverse transcription co-factors and integrate signals from multiple pathways to control metabolism, oxidative stress response, and cell cycle. Recently, insulin/FOXO signaling has been implicated in the regulation of insect development via the interaction with insect hormones, such as ecdysone and juvenile hormone. In this study, we identified an interaction between dFOXO and the zinc finger transcription factor Kruppel homolog 1 (Kr-h1), one of the key players in juvenile hormone signaling in Drosophila. We found that Kr-h1 mutants have reduced triglyceride storage, decreased insulin signaling and delayed larval development. Notably, Kr-h1 physically and genetically interacts with dFOXO in vitro and in vivo to regulate the transcriptional activation of adipose lipase brummer (bmm). The transcriptional co-regulation by Kr-h1 and dFOXO may represent a broad mechanism by which Kruppel-like factors integrate with insulin signaling to maintain metabolic homeostasis and coordinate organism growth.

Widespread sampling biases in herbaria revealed from large-scale digitization

Daru, B. H. — 2017-07-18

O_LINon-random collecting practices may bias conclusions drawn from analyses of herbarium records. Recent efforts to fully digitize and mobilize regional floras offer a timely opportunity to assess commonalities and differences in herbarium sampling biases.nC_LIO_LIWe determined spatial, temporal, trait, phylogenetic, and collector biases in [~]5 million herbarium records, representing three of the most complete digitized floras of the world: Australia (AU), South Africa (SA), and New England (NE).nC_LIO_LIWe identified numerous shared and unique biases among these regions. Shared biases included specimens i) collected close to roads and herbaria; ii) collected more frequently during spring; iii) of threatened species collected less frequently; and iv) of close relatives collected in similar numbers. Regional differences included i) over-representation of graminoids in SA and AU and of annuals in AU; and ii) peak collection during the 1910s in NE, 1980s in SA, and 1990s in AU. Finally, in all regions, a disproportionately large percentage of specimens were collected by a few individuals. These mega-collectors, and their associated preferences and idiosyncrasies, may have shaped patterns of collection bias via founder effects.nC_LIO_LIStudies using herbarium collections should account for sampling biases and future collecting efforts should avoid compounding these biases.nC_LI

Dissociable neural mechanisms track evidence accumulation for selection of attention versus action

Shenhav, A. — 2017-08-02

Decision-making is typically studied as a sequential process from the selection of what to attend (e.g., between possible tasks, stimuli, or stimulus attributes) to the selection of which actions to take based on the attended information. However, people often gather information across these levels in parallel. For instance, even as they choose their actions, they may continue to evaluate how much to attend other tasks or dimensions of information within a task. We scanned participants while they made such parallel evaluations, simultaneously weighing how much to attend two dynamic stimulus attributes and which response to give based on the attended information. Regions of prefrontal cortex tracked information about the stimulus attributes in dissociable ways, related to either the predicted reward (ventromedial prefrontal cortex) or the degree to which that attribute was being attended (dorsal anterior cingulate, dACC). Within dACC, adjacent regions tracked uncertainty at different levels of the decision, regarding what to attend versus how to respond. These findings bridge research on perceptual and value-based decision-making, demonstrating that people dynamically integrate information in parallel across different levels of decision making.nnNaturalistic decisions allow an individual to weigh their options within a particular task (e.g., how best to word the introduction to a paper) while also weighing how much to attend other tasks (e.g., responding to e-mails). These different types of decision-making have a hierarchical but reciprocal relationship: Decisions at higher levels inform the focus of attention at lower levels (e.g., whether to select between citations or email addresses) while, at the same time, information at lower levels (e.g., the salience of an incoming email) informs decisions regarding which task to attend. Critically, recent studies suggest that decisions across these levels may occur in parallel, continuously informed by information that is integrated from the environment and from ones internal milieu1,2.nnResearch on cognitive control and perceptual decision-making has examined how responses are selected when attentional targets are clearly defined (e.g., based on instruction to attend a stimulus dimension), including cases in which responding requires accumulating information regarding a noisy percept (e.g., evidence favoring a left or right response)3-7. Separate research on value-based decision-making has examined how individuals select which stimulus dimension(s) to attend in order to maximize their expected rewards8-11. However, it remains unclear how the accumulation of evidence to select high-level goals and/or attentional targets interacts with the simultaneous accumulation of evidence to select responses according to those goals (e.g., based on the perceptual properties of the stimuli). Recent work has highlighted the importance of such interactions to understanding task selection12-15, multi-attribute decision-making16-18, foraging behavior19-21, cognitive effort22,23, and self-control24-27.nnWhile these interactions remain poorly understood, previous research has identified candidate neural mechanisms associated with multi-attribute value-based decision-making11,28,29 and with selecting a response based on noisy information from an instructed attentional target3-5. These research areas have implicated the ventromedial prefrontal cortex (vmPFC) in tracking the value of potential targets of attention (e.g., stimulus attributes)8,11 and the dorsal anterior cingulate cortex (dACC) in tracking an individuals uncertainty regarding which response to select30-32. It has been further proposed that dACC may differentiate between uncertainty at each of these parallel levels of decision-making (e.g., at the level of task goals or strategies vs. specific motor actions), and that these may be separately encoded at different locations along the dACCs rostrocaudal axis32,33. However, neural activity within and across these prefrontal regions has not yet been examined in a setting in which information is weighed at both levels within and across trials.nnHere we use a value-based perceptual decision-making task to examine how people integrate different dynamic sources of information to decide (a) which perceptual attribute to attend and (b) how to respond based on the evidence for that attribute. Participants performed a task in which they regularly faced a conflict between attending the stimulus attribute that offered the greater reward or the attribute that was more perceptually salient (akin to persevering in writing ones paper when an enticing email awaits). We demonstrate that dACC and vmPFC track evidence for the two attributes in dissociable ways. Across these regions, vmPFC weighs attribute evidence by the reward it predicts and dACC weighs it by its attentional priority (i.e., the degree to which that attribute drives choice). Within dACC, adjacent regions differentiated between uncertainty at the two levels of the decision, regarding what to attend (rostral dACC) versus how to respond (caudal dACC).

Dissociable components of the reward circuit are involved in appraisal versus choice

Shenhav, A. — 2017-08-03

People can evaluate a set of options as a whole, or they can approach those same options with the purpose of making a choice between them. A common network has been implicated across these two types of evaluations, including regions of ventromedial prefrontal cortex and the posterior midline. We test the hypothesis that sub-components of this reward circuit are differentially involved in triggering more automatic appraisal of ones options (Dorsal Value Network) versus explicitly comparing between those options (Ventral Value Network). Participants undergoing fMRI were instructed to appraise how much they liked a set of products (Like) or to choose the product they most preferred (Choose). Activity in the Dorsal Value Network consistently tracked set liking, across both task-relevant (Like) and task-irrelevant (Choose) trials. In contrast, the Ventral Value Network was sensitive to evaluation condition (more active during Choose than Like trials). Within vmPFC, anatomically distinct regions were dissociated in their sensitivity to choice (ventrally, in medial OFC) versus appraisal (dorsally, in pregenual ACC). Dorsal regions additionally tracked decision certainty across both types of evaluation. These findings suggest that separable mechanisms drive decisions about how good ones options are versus decisions about which option is best.

Seamless Multiscale Modeling of Coagulation Using Dissipative Particle Dynamics

Yazdani, A. — 2017-08-26

We propose a new multiscale framework that seamlessly integrates four key components of blood clotting namely, blood rheology, cell mechanics, coagulation kinetics and transport of species and platelet adhesive dynamics. We use transport dissipative particle dynamics (tDPD), which is the extended form of original DPD, as the base solver, while a coarse-grained representation of blood cells membrane accounts for its mechanics. Our results show the dominant effect of blood flow and high Peclet numbers on the reactive transport of the chemical species signifying the importance of membrane bound reactions on the surface of adhered platelets. This new multiscale particle-based methodology helps us probe synergistic mechanisms of thrombus formation, and can open new directions in addressing other biological processes from sub-cellular to macroscopic scales.

Within and across-trial dynamics of human EEG reveal cooperative interplay between reinforcement learning and working memory

Collins, A. — 2017-09-05

Learning from rewards and punishments is essential to survival, and facilitates flexible human behavior. It is widely appreciated that multiple cognitive and reinforcement learning systems contribute to behavior, but the nature of their interactions is elusive. Here, we leverage novel methods for extracting trial-by-trial indices of reinforcement learning (RL) and working memory (WM) in human electroencephalography to reveal single trial computations beyond that afforded by behavior alone. Within-trial dynamics confirmed that increases in neural expectation were predictive of reduced neural surprise in the following feedback period, supporting central tenets of RL models. Cross-trial dynamics revealed a cooperative interplay between systems for learning, in which WM contributes expectations to guide RL, despite competition between systems during choice. Together, these results provide a deeper understanding of how multiple neural systems interact for learning and decision making, and facilitate analysis of their disruption in clinical populations.nnOne sentence summaryDecoding of dynamical neural signals in humans reveals cooperation between cognitive and habit learning systems.

Exome chip meta-analysis elucidates the genetic architecture of rare coding variants in smoking and drinking behavior

Liu, D. J. — 2017-09-12

BackgroundSmoking and alcohol use behaviors in humans have been associated with common genetic variants within multiple genomic loci. Investigation of rare variation within these loci holds promise for identifying causal variants impacting biological mechanisms in the etiology of disordered behavior. Microarrays have been designed to genotype rare nonsynonymous and putative loss of function variants. Such variants are expected to have greater deleterious consequences on gene function than other variants, and significantly contribute to disease risk.nnMethodsIn the present study, we analyzed [~]250,000 rare variants from 17 independent studies. Each variant was tested for association with five addiction-related phenotypes: cigarettes per day, pack years, smoking initiation, age of smoking initiation, and alcoholic drinks per week. We conducted single variant tests of all variants, and gene-based burden tests of nonsynonymous or putative loss of function variants with minor allele frequency less than 1%.nnResultsMeta-analytic sample sizes ranged from 70,847 to 164,142 individuals, depending on the phenotype. Known loci tagged by common variants replicated, but there was no robust evidence for individually associated rare variants, either in gene based or single variant tests. Using a modified method-of-moment approach, we found that all low frequency coding variants, in aggregate, contributed 1.7% to 3.6% of the phenotypic variation for the five traits (p<.05).nnConclusionsThe findings indicate that rare coding variants contribute to phenotypic variation, but that much larger samples and/or denser genotyping of rare variants will be required to successfully identify associations with these phenotypes, whether individual variants or gene- based associations.

Data-driven Modeling of Thrombus Size and Shape in Aortic Dissections: Role of Hemodynamics

Yazdani, A. — 2017-09-12

Aortic dissection is a pathology that manifests due to micro-structural defects in the aortic wall. Blood enters the damaged wall through an intimal tear, thereby creating a so-called false lumen and exposing the blood to thrombogenic intramural constituents such as collagen. The natural history of this acute vascular injury thus depends, in part, on thrombus formation, maturation, and possible healing within the false lumen. A key question is: Why do some false lumens thrombose completely while other thrombose partially or little at all? An ability to predict the location and extent of thrombus in subjects with dissection could contribute significantly to clinical decision-making, including interventional design. We develop, for the first time, a data-driven particle-continuum model for thrombus formation in a murine model of aortic dissection. In the proposed model, we simulate a final-value problem in lieu of the original initial-value problem with significantly fewer particles that may grow in size upon activation, representing the local concentration of blood-borne species. Numerical results confirm that geometry and local hemodynamics play significant roles in the acute progression of thrombus. Despite geometrical differences between murine and human dissections, mouse models can provide considerable insight and have gained in popularity owing to their reproducibility. Our results for three classes of geometrically different false lumens show that thrombus forms and extends to a greater extent in regions with lower bulk shear rates. Dense thrombi are less likely to form in high-shear zones and in the presence of strong vortices. The present data-driven study suggests that the proposed model is robust and can be employed to assess thrombus formation in human aortic dissections.

Discordant association of the CREBRF rs373863828 minor allele with increased body mass index and protection from type 2 diabetes in Maori and Pacific (Polynesian) people living in New Zealand

Krishnan, M. — 2017-09-13

Aim/HypothesesThe minor allele of CREBRF rs373863828 associates with increased body mass index (BMI) and reduced risk of type 2 diabetes (T2D) in the Samoan population of Samoa and American Samoa. Our aim was to test rs373863828 for association with BMI and odds of T2D, gout and chronic kidney disease (CKD) in M[a]ori and Pacific (Polynesian) people living in Aotearoa New Zealand in 2,286 adults.nnMethodsAssociation analyses were performed by linear and logistic regression with BMI, log-transformed BMI, waist circumference, T2D, gout and CKD. Analyses were adjusted for age, sex, the first four genome-wide principal components, and (when appropriate) BMI, waist circumference and T2D.nnResultsFor the minor allele of rs373863828 the effect size for log-transformed BMI was 0.038 (95% CI [0.022-0.055], P=4.8x10-6) and for T2D was OR=0.59 (95% CI [0.47-0.73], P=1.9x10-6). There was no evidence for association of genotype with variance in BMI (P=0.13). Nor was there evidence for association with serum urate ({beta}=0.012 mmol/L, Pc=0.10), gout (OR=1.00, P=0.98) or CKD (OR=0.91, P=0.59).nnConclusions/interpretationOur results replicated, with very similar effect sizes, association of the minor allele of rs373863828 with higher BMI but lower odds of T2D among New Zealand Polynesian adults, as in Samoan adults living in Samoa and American Samoa.

The Microbiomes of Pancreatic Tissue in Pancreatic Cancer and Non-Cancer Subjects

del Castillo, E. — 2017-09-15

ObjectiveTo determine whether bacteria are present in the pancreas of pancreatic cancer and non-cancer subjects and examine whether bacterial profiles vary by site and disease phenotype.nnDesign77 patients requiring surgery for pancreatic diseases, or diseases of the foregut, at the Rhode Island Hospital (RIH) were recruited into this study between 2014 and 2016. In addition, 36 whole pancreas were obtained from the National Disease Research Interchange (NDRI) from subjects who were of similar age as the RIH patients and had not died of cancer. The primary exposure of interest was the measurement of the relative abundance of bacterial taxa in all tissue specimens using 16S rRNA gene sequencing.nnResultsNumber of bacterial reads per sample varied substantially across sample type and patients, but all demonstrated the presence of diverse gastrointestinal bacteria, including bacterial taxa typically identified in the oral cavity. Bacterial profiles were noted to be more similar within individuals across sites in the pancreas, than between individuals by site, suggesting that the pancreas as a whole has its own microbiome. Comparing the mean relative abundance of bacterial taxa in pancreatic cancer patients to those without cancer revealed differences in bacterial taxa previously linked to periodontal disease, including Porphyromonas.nnConclusionsBacterial taxa known to inhabit the oral cavity, as well as the intestine, were identified in pancreatic tissue of cancer and non-cancer subjects. Whether any of these bacteria play a causal role in pancreatic carcinogenesis, or are simply opportunistic in nature, needs to be further examined.

Identifying structural variants using linked-read sequencing data

Elyanow, R. — 2017-09-18

Structural variation, including large deletions, duplications, inversions, translocations, and other rearrangements, is common in human and cancer genomes. A number of methods have been developed to identify structural variants from Illumina short-read sequencing data. However, reliable identification of structural variants remains challenging because many variants have breakpoints in repetitive regions of the genome and thus are difficult to identify with short reads. The recently developed linked-read sequencing technology from 10X Genomics combines a novel barcoding strategy with Illumina sequencing. This technology labels all reads that originate from a small number (~5-10) DNA molecules ~50Kbp in length with the same molecular barcode. These barcoded reads contain long-range sequence information that is advantageous for identification of structural variants. We present Novel Adjacency Identification with Barcoded Reads (NAIBR), an algorithm to identify structural variants in linked-read sequencing data. NAIBR predicts novel adjacencies in a individual genome resulting from structural variants using a probabilistic model that combines multiple signals in barcoded reads. We show that NAIBR outperforms several existing methods for structural variant identification - including two recent methods that also analyze linked-reads - on simulated sequencing data and 10X whole-genome sequencing data from the NA12878 human genome and the HCC1954 breast cancer cell line. Several of the novel somatic structural variants identified in HCC1954 overlap known cancer genes.

Perimenstrual Symptom Exacerbation in Borderline Personality Disorder: Evidence from Multilevel Models and the Carolina Premenstrual Assessment Scoring System

Eisenlohr-Moul, T. — 2017-09-24

BackgroundIndividuals with borderline personality disorder (BPD) suffer from a constellation of rapidly shifting emotional, interpersonal, and behavioral symptoms. The menstrual cycle may contribute to symptom instability among females with this disorder.nnMethodsFifteen healthy, unmedicated females with BPD and without dysmenorrhea reported daily symptoms across 35 days. Urine luteinizing hormone (LH) and salivary progesterone (P4) were used to confirm ovulation and cycle phase. Cyclical worsening of symptoms was evaluated using (1) phase contrasts in multilevel models and (2) the Carolina Premenstrual Assessment Scoring System (C-PASS; Eisenlohr-Moul et al., 2017b), a protocol for evaluating clinically significant cycle effects on symptoms.nnResultsMost symptoms demonstrated midluteal worsening, a perimenstrual peak, and resolution of symptoms in the follicular or ovulatory phase. Post-hoc correlations with person-centered progesterone revealed negative correlations with most symptoms. Depressive symptoms showed an unexpected delayed pattern in which baseline levels of symptoms were observed in the ovulatory and midluteal phases, and exacerbations were observed during both the perimenstrual and follicular phases. The majority of participants met C-PASS criteria for clinically significant (>=30%) symptom exacerbation. All participants met the emotional instability criterion of BPD, and no participant met DSM-5 criteria for premenstrual dysphoric disorder (PMDD).nnConclusionsFemales with BPD may be at elevated risk for perimenstrual worsening of emotional symptoms. Longitudinal studies with fine-grained hormonal measurement as well as hormonal experiments are needed to determine the pathophysiology of perimenstrual exacerbation in BPD.

Compositional clustering in task structure learning

Franklin, N. T. — 2017-10-02

Humans are remarkably adept at generalizing knowledge between experiences in a way that can be difficult for computers. Often, this entails generalizing constituent pieces of experiences that do not fully overlap, but nonetheless share useful similarities with, previously acquired knowledge. However, it is often unclear how knowledge gained in one context should generalize to another. Previous computational models and data suggest that rather than learning about each individual context, humans build latent abstract structures and learn to link these structures to arbitrary contexts, facilitating generalization. In these models, task structures that are more popular across contexts are more likely to be revisited in new contexts. However, these models predict that structures are either re - used as a whole or created from scratch, prohibiting the ability to generalize constituent parts of learned structures. This contrasts with ecological settings, where some aspects of task structure, such as the transition function, will be shared between context separately from other aspects, such as the reward function. Here, we develop a novel non - parametric Bayesian agent that forms independent latent clusters for transition and reward functions that may have different popularity across contexts. We compare this agent to an agent that jointly clusters both across a range of task domains. We show that relative performance of the two agents depends on the statistics of the task domain, including the mutual information between transition and reward functions in the environment, and the stochasticity of the observations. We formalize our analysis through an information theoretic account of the priors, and develop a meta learning agent that can dynamically arbitrate between strategies across task domains. We argue that this provides a first step in allowing for compositional structures in reinforcement learners, which should be provide a better model of human learning and additional flexibility for artificial agents.nnAuthor summaryA musician may learn to generalize behaviors across instruments for different purposes, for example, reusing hand motions used when playing classical on the flute to play jazz on the saxophone. Conversely, she may learn to play a single song across many instruments that require completely distinct physical motions, but nonetheless transfer knowledge between them. This degree of compositionality is often absent from computational frameworks of learning, forcing agents either to generalize entire learned policies or to learn new policies from scratch. Here, we propose a solution to this problem that allows an agent to generalize components of a policy independently and compare it to an agent that generalizes components as a whole. We show that the degree to which one form of generalization is favored over the other is dependent on the features of task domain, with independent generalization of task components favored in environments with weak relationships between components or high degrees of noise and joint generalization of task components favored when there is a clear, discoverable relationship between task components. Furthermore, we show that the overall meta structure of the environment can be learned and leveraged by an agent that dynamically arbitrates between these forms of structure learning.

Cross-task contributions of fronto-basal ganglia circuitry in response inhibition and conflict-induced slowing

Jahfari, S. — 2017-10-06

Why are we so slow in choosing the lesser of two evils? We considered whether such slowing relates to uncertainty about the value of these options, which arises from the tendency to avoid them during learning, and whether such slowing relates to fronto-subthalamic inhibitory control mechanisms. 49 participants performed a reinforcement-learning task and a stop-signal task while fMRI was recorded. A reinforcement-learning model was used to quantify learning strategies. Individual differences in lose-lose slowing related to information uncertainty due to sampling, and independently, to less efficient response inhibition in the stop-signal task. Neuroimaging analysis revealed an analogous dissociation: subthalamic nucleus (STN) BOLD activity related to variability in stopping latencies, whereas weaker fronto-subthalamic connectivity related to slowing and information sampling. Across tasks, fast inhibitors increased STN activity for successfully cancelled responses in the stop task, but decreased activity for lose-lose choices. These data support the notion that fronto-STN communication implements a rapid but transient brake on response execution, and that slowing due to decision uncertainty could result from an inefficient release of this "hold your horses" mechanism.

Cell-specific exon methylation and CTCF binding in neurons regulates calcium ion channel splicing and function

Lopez Soto, E. J. — 2019-12-15

Cell-specific alternative splicing modulates myriad cell functions and this process is disrupted in disease. The mechanisms governing alternative splicing are known for relatively few genes and typically focus on RNA splicing factors. In sensory neurons, cell-specific alternative splicing of the presynaptic voltage-gated calcium channel Cacna1b gene modulates opioid sensitivity. How this splicing is regulated has remained unknown. We find that cell-specific exon DNA hypomethylation permits binding of CTCF, the master regulator of chromatin structure in mammals, which, in turn, controls splicing in noxious heat-sensing nociceptors. Hypomethylation of an alternative exon specifically in nociceptors allows for CTCF binding, and expression of CaV2.2 channels with increased opioid sensitivity. Following nerve injury, exon methylation is increased, and splicing is disrupted. Our studies define the molecular mechanisms of cell-specific alternative splicing of a functionally validated exon in normal and disease states - and reveal a potential target for the treatment of chronic pain. HighlightsO_LIThe molecular basis of cell-specific splicing of a synaptic calcium channel gene. C_LIO_LISplicing controlled by cell-specific exon hypomethylation and CTCF binding. C_LIO_LIPeripheral nerve injury disrupts exon hypomethylation and splicing. C_LIO_LITargeted demethylation of exon by dCAS9-TET modifies alternative splicing. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=158 SRC="FIGDIR/small/876185v2_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@11b592eorg.highwire.dtl.DTLVardef@7c0eddorg.highwire.dtl.DTLVardef@b403bborg.highwire.dtl.DTLVardef@611882_HPS_FORMAT_FIGEXP M_FIG Cell-specific epigenetic modifications in a synaptic calcium ion channel gene controls cell-specific splicing in normal and neuropathic pain. In naive animals, in most neurons, Cacna1b e37a locus is hipermethylated (5-mC) and CTCF does not bind this locus. During splicing, e37a is skipped and Cacna1b mRNAs include e37b. In contrast, in Trpv1-lineage neurons, Cacna1b e37a locus is hypomethylated and is permissive for CTCF binding. CTCF promotes e37a inclusion and both Cacna1b e37a and e37b mRNAs are expressed. E37a confers strong sensitivity to the Cav2.2 channel to inhibition by -opioid receptors (OR). Morphine is more effective at inhibiting e37a-containing Cav2.2 channels. After peripheral nerve injury that results in pathological pain, methylation level of Cacna1b e37a locus is increased, CTCF binding is impaired, and Cacna1b e37a mRNA levels are decreased. This disrupted splicing pattern is associated with reduced efficacy of morphine in vivo. C_FIG

Differential involvement of EEG oscillatory components in sameness vs. spatial-relation visual reasoning tasks

Alamia, A. — 2019-12-16

The development of deep convolutional networks (DCNs) has recently led to great successes in computer vision and have become de facto computational models of vision. However, a growing body of work suggests that they exhibit critical limitations beyond image categorization. Here, we study a fundamental limitation of DCNs for judging whether two items are the same or different (SD) compared to a baseline assessment of their spatial relationship (SR). We test the prediction that SD tasks recruit additional cortical mechanisms which underlie critical aspects of visual cognition that are not explained by current computational models. We thus recorded EEG signals from 14 participants engaged in the same tasks as the computational models. Importantly, the two tasks were matched in terms of difficulty by an adaptive psychometric procedure: yet, on top of a modulation of evoked potentials, our results revealed higher activity in the low beta (13-20Hz) band in the SD compared to the SR conditions, which we surmise as reflecting the crucial involvement of recurrent mechanisms sustaining working memory and attention. Author SummaryDespite the impressive progress of deep convolutional networks (DCNs) in object recognition, recent studies demonstrated that state-of-the-art vision algorithms encounter severe limitations when performing certain visual reasoning tasks: for instance, convolutional networks can easily solve problems involving spatial relations, but fail in identifying whether two items are identical or different (same-different task). This conclusion led us to test the hypothesis that different computational mechanisms are needed to successfully perform these tasks also in the visual system. First, we confirmed in our simulations that DCNs can successfully perform spatial relationship tasks but struggle with same-different ones. Then, we tested 14 participants on the same experimental design while recording their EEG signals. Remarkably, our results revealed a significant difference between the tasks in the occipital brain regions both in evoked potentials and in the oscillatory dynamics. Specifically, an increase of activity was found when performing the SD over the SR condition. We interpret these results as reflecting the fundamental involvement of recurrent mechanisms implementing cognitive functions such as working memory and attention.

When, why and how clonal diversity predicts future tumour growth

Noble, R. J. — 2019-12-18

Intratumour heterogeneity holds promise as a prognostic biomarker in multiple cancer types. However, the relationship between this marker and its clinical impact is mediated by an evolutionary process that is not well understood. Here we employ a spatial computational model of tumour evolution to assess when, why and how intratumour heterogeneity can be used to forecast tumour growth rate, an important predictor of clinical progression. We identify three conditions that can lead to a positive correlation between clonal diversity and subsequent growth rate: diversity is measured early in tumour development; selective sweeps are rare; and/or tumours vary in the rate at which they acquire driver mutations. Opposite conditions typically lead to negative correlation. Our results further suggest that prognosis can be better predicted on the basis of both clonal diversity and genomic instability than either factor alone. Nevertheless, we find that, for predicting tumour growth, clonal diversity is likely to perform worse than conventional measures of tumour stage and grade. We thus offer explanations - grounded in evolutionary theory - for empirical findings in various cancers. Our work informs the search for new prognostic biomarkers and contributes to the development of predictive oncology.

Odor Modulates the Temporal Dynamics of Fear Memory Consolidation

Grella, S. L. — 2019-12-19

Systems consolidation (SC) theory proposes that recent, contextually rich memories are stored in the hippocampus (HPC). As these memories become remote, they are believed to rely more heavily on cortical structures within the prefrontal cortex (PFC), where they lose much of their contextual detail and become schematized. Odor is a particularly evocative cue for intense remote memory recall and despite these memories being remote, they are highly contextual. In instances such as post-traumatic stress disorder (PTSD), intense remote memory recall can occur years after trauma, which seemingly contradicts SC. We hypothesized that odor may shift the organization of salient or fearful memories such that when paired with an odor at the time of encoding, they are delayed in the de-contextualization process that occurs across time, and retrieval may still rely on the HPC, where memories are imbued with contextually rich information, even at remote time points. We investigated this by tagging odor- and non-odor-associated fear memories in male c57BL/6 mice and assessed recall and c-Fos expression in the dorsal CA1 (dCA1) and prelimbic cortex (PL) 1 d or 21 d later. In support of SC, our data showed that recent memories were more dCA1-dependent whereas remote memories were more PL-dependent. However, we also found that odor influenced this temporal dynamic biasing the memory system from the PL to the dCA1 when odor cues were present. Behaviorally, inhibiting the dCA1 with activity-dependent DREADDs had no effect on recall at 1 d and unexpectedly caused an increase in freezing at 21 d. Together, these findings demonstrate that odor can shift the organization of fear memories at the systems level.

Core-2 O-glycans are required for Galectin-3 interaction with the osteoarthritis related protein lubricin

Flowers, S. A. — 2019-12-21

Synovial fluid lubricin (proteoglycan 4) is a mucin-type O-linked glycosylated (60% of the mass) biological lubricant involved in osteoarthritis (OA) development. Lubricin has been reported to be cross-linked by synovial galectin-3 on the lubricating articular surface. Here, we confirm that binding to galectin-3 depended on core-2 O-linked glycans, where surface plasmon resonance of a recombinant lubricin (rhPRG4) devoid of core-2 structures lacked binding capacity to recombinant galectin-3. Both galectin-3 levels and interactions with synovial lubricin were found to be decreased in late-stage OA patients coinciding with an increase of truncated and less sialylated core 1 O-glycans. These data suggest a defect cross-linking of surface active molecules in OA and provides novel insights into OA molecular pathology.

An instrument to assess HIV-related knowledge and adjustment to HIV+ status, and their association with anti-retroviral adherence

Laws, M. B. — 2019-12-30

BackgroundFindings on the association between health literacy and anti-retroviral (ARV) adherence are inconsistent. Health literacy is usually operationalized with simple tests of basic literacy, but more complex conceptions of health literacy include content knowledge. People living with chronic illness also conceptualize and experience illness in ways other than biomedical or mechanistic models of disease. ObjectiveThere are no instruments that comprehensively assess knowledge of people living with HIV concerning HIV disease and treatment; or psychological adjustment to being HIV+. Little is known about the relationship between factual knowledge, or positive identification as HIV+, and anti-retroviral (ARV) adherence. MethodsFormative work with in-depth semi-structured interviews, and cognitive testing, to develop a structured instrument assessing HIV-related knowledge, and personal meanings of living with HIV. Pilot administration of the instrument to a convenience sample of 101 respondents. Key ResultsRespondents varied considerably in their expressed need for in-depth knowledge, the accuracy of their understanding of relevant scientific concepts and facts about ARV treatment, and psychological adjustment and acceptance of HIV+ status. Most knowledge domains were not significantly related to self-reported ARV adherence, but accurate knowledge specifically about ARV treatment was (r=0.25, p=.02), as was an adapted version of the Need for Cognition scale (r=.256, p=.012). Negative feelings about living with HIV (r=.33, p=.0012), and medication taking (r=.276, p=.008) were significantly associated with non-adherence. ConclusionThe instrument may be useful in diagnosing addressable reasons for non-adherence, as a component of psychoeducational interventions, and for evaluation of such interventions. PLAIN LANGUAGE SUMMARYWe developed a questionnaire for people with HIV. General knowledge about HIV wasnt related to whether people took their medications, but specific information about treatment was. People with lots of bad feelings about living with HIV were less likely to take medications as prescribed. People who liked to think more about their decisions were more likely to take their medications.

The stability flexibility tradeoff and the dark side of detail

Nassar, M. R. — 2020-01-03

Learning in dynamic environments requires integrating over stable fluctuations to minimize the impact of noise (stability) but rapidly responding in the face of fundamental changes (flexibility). Achieving one of these goals often requires sacrificing the other to some degree, producing a stability-flexibility tradeoff. Individuals navigate this tradeoff in different ways, with some people learning rapidly (emphasizing flexibility) and others relying more heavily on historical information (emphasizing stability). Despite the prominence of such individual differences in learning tasks, the degree to which they relate to broader characteristics of real-world behavior or pathologies has not been well explored. Here we relate individual differences in learning behavior to self-report measures thought to collectively capture characteristics of the Autism spectrum. We show that that young adults who learn most slowly tend to integrate more effective samples into their beliefs about the world making them more robust to noise (more stability), but are more likely to integrate information from previous contexts (less flexibility). We show that individuals who report paying more attention to detail tend to use high flexibility and low stability information processing strategies. We demonstrate the robustness of this inverse relationship between attention to detail and formation of stable beliefs in a heterogeneous population of children that includes a high proportion of Autism diagnoses. Together, our results highlight that attention to detail reflects an information processing policy that comes with a substantial downside, namely the ability to integrate data to overcome environmental noise.

Identifying Sequence Perturbations to an Intrinsically Disordered Protein that Determine Its Phase Separation Behavior

Schuster, B. — 2020-01-06

Phase separation of intrinsically disordered proteins (IDPs) commonly underlies the formation of membraneless organelles, which compartmentalize molecules intracellularly in the absence of a lipid membrane. Identifying the protein sequence features responsible for IDP phase separation is critical for understanding physiological roles and pathological consequences of biomolecular condensation, as well as for harnessing phase separation for applications in bio-inspired materials design. To expand our knowledge of sequence determinants of IDP phase separation, we characterized variants of the intrinsically disordered RGG domain from LAF-1, a model protein involved in phase separation and a key component of P granules. Based on a predictive coarse-grained IDP model, we identified a region of the RGG domain that has high contact probability and is highly conserved between species; deletion of this region significantly disrupts phase separation in vitro and in vivo. We determined the effects of charge patterning on phase behavior through sequence shuffling. By altering the wild-type sequence, which contains well-mixed charged residues, to increase charge segregation, we designed sequences with significantly increased phase separation propensity. This result indicates the natural sequence is under negative selection to moderate this mode of interaction. We measured the contributions of tyrosine and arginine residues to phase separation experimentally through mutagenesis studies and computationally through direct interrogation of different modes of interaction using all-atom simulations. Finally, we show that in spite of these sequence perturbations, the RGG-derived condensates remain liquid-like. Together, these studies advance a predictive framework and identify key biophysical principles of sequence features important to phase separation. Significance StatementMembraneless organelles are assemblies of highly concentrated biomolecules that form through a liquid-liquid phase separation process. These assemblies are often enriched in intrinsically disordered proteins, which play an important role in driving phase separation. Understanding the sequence-to-phase behavior relationship of these disordered proteins is important for understanding the biochemistry of membraneless organelles, as well as for designing synthetic organelles and biomaterials. In this work, we explore a model protein, the disordered N-terminal domain of LAF-1, and highlight how three key features of the sequence control the proteins propensity to phase separate. Combining predictive simulations with experiments, we find that phase behavior of this model IDP is dictated by the presence of a short conserved domain, charge patterning, and arginine-tyrosine interactions.

"Pain in my heart": Understanding perinatal depression among women living with HIV in Malawi

LeMasters, K. — 2020-01-06

BackgroundPerinatal depression (PND) can interfere with HIV care engagement and outcomes. We examined experiences of PND among women living with HIV (WLWH) in Malawi. MethodsWe screened 73 WLWH presenting for perinatal care in Lilongwe, Malawi using the Edinburgh Postnatal Depression Scale (EPDS). We conducted interviews with 24 women experiencing PND and analyzed data using inductive and deductive coding and narrative analysis. ResultsWomen experienced a double burden of physical and mental illness, expressed as pain in ones heart. Receiving an HIV diagnosis unexpectedly during antenatal care was a key contributor to developing PND. This development was influenced by stigmatization and social support. ConclusionsThese findings highlight the need to recognize the mental health implications of routine screening for HIV and to routinely screen and treat PND among WLWH. Culturally appropriate mental health interventions are needed in settings with a high HIV burden.

Early life adversity decreases fear expression in pre-adolescence by accelerating amygdalar parvalbumin cell development

Manzano Nieves, G. — 2020-01-07

Resource insecurity (e.g., poverty) can be a significant source of stress. Decreased resources during childhood has been associated with increased risk for developing stress-related disorders, including major depressive disorder and anxiety Although the link between early life adversity and increased risk for psychopathology has been well established, the developmental mechanisms remain unclear. Using a mouse model of poverty-like rearing, limited bedding and nesting materials (LB), we tested the effects of LB on the development of fear learning and of key neuronal structures involved in emotional regulation, the medial prefrontal cortex (mPFC) and basolateral amygdala (BLA). LB delayed the ability of pre-adolescent mice to express, but not form, an auditory conditioned fear memory. LB disrupted typical fear circuit development, accelerating parvalbumin positive (PV+) inhibitory interneuron maturation in the BLA and delaying the maturation of connections between the mPFC and BLA. The decreased fear expression in LB reared mice during early development was rescued through optogenetic inactivation of PV+ cells in the BLA. Together our data demonstrate that LB has profound and deleterious effects on mPFC and BLA development, decreasing threat-associated behavior expression, but not learning, in childhood. The current results provide a model of transiently blunt emotional reactivity in childhood, with fear-associated memories emerging later in adolescence, and possibly contributing to later pathology development.

Population sequencing data reveal a compendium of mutational processes in human germline

Seplyarskiy, V. B. — 2020-01-11

Mechanistic processes underlying human germline mutations remain largely unknown. Variation in mutation rate and spectra along the genome is informative about the biological mechanisms. We statistically decompose this variation into separate processes using a blind source separation technique. The analysis of a large-scale whole genome sequencing dataset (TOPMed) reveals nine processes that explain the variation in mutation properties between loci. Seven of these processes lend themselves to a biological interpretation. One process is driven by bulky DNA lesions that resolve asymmetrically with respect to transcription and replication. Two processes independently track direction of replication fork and replication timing. We identify a mutagenic effect of active demethylation primarily acting in regulatory regions. We also demonstrate that a recently discovered mutagenic process specific to oocytes can be localized solely from population sequencing data. This process is spread across all chromosomes and is highly asymmetric with respect to the direction of transcription, suggesting a major role of DNA damage.

Recessive deleterious variation has a limited impact on signals of adaptive introgression in human populations

Zhang, X. — 2020-01-14

Admixture with archaic hominins has altered the landscape of genomic variation in modern human populations. Several gene regions have been previously identified as candidates of adaptive introgression (AI) that facilitated human adaptation to specific environments. However, simulation-based studies have suggested that population genetics processes other than adaptive mutations, such as heterosis from recessive deleterious variants private to populations before admixture, can also lead to patterns in genomic data that resemble adaptive introgression. The extent to which the presence of deleterious variants affect the false-positive rate and the power of current methods to detect AI has not been fully assessed. Here, we used extensive simulations to show that recessive deleterious mutations can increase the false positive rates of tests for AI compared to models without deleterious variants. We further examined candidates of AI in modern humans identified from previous studies and show that, although deleterious variants may hinder the performance of AI detection in modern humans, most signals remained robust when deleterious variants are included in the null model. While deleterious variants may have a limited impact on detecting signals of adaptive introgression in humans, we found that at least two AI candidate genes, HYAL2 and HLA, are particularly susceptible to high false positive rates due to the recessive deleterious mutations. By quantifying parameters that affect heterosis, we show that the high false positives are largely attributed to the high exon densities together with low recombination rates in the genomic regions, which can further be exaggerated by the population growth in recent human evolution. Although the combination of such parameters is rare in the human genome, caution is still warranted in other species with different genomic composition and demographic histories.

Time course and shared neurocognitive mechanisms of mental imagery and visual perception

Maier, M. — 2020-01-15

When we imagine an object and when we actually see that object, similar brain regions become active. Yet, the time course and mechanisms with which imagery engages perceptual networks remain to be better understood. An emerging view holds that imagery and perception follow distinct dynamics during early visual processing with similarities arising only during later, high-level visual processing. However, confounds of visual stimulation and paradigms favoring observation of high-level processes associated with subjective imagery strength may have precluded evidence of earlier shared mechanisms. We therefore manipulated prior knowledge that informs early-stage top-down predictions and tracked electrophysiological brain responses while fully controlling visual stimulation. Participants saw and imagined objects associated with varying amounts of semantic knowledge. Imagery and perception were equally influenced by knowledge at an early stage (P1 component), revealing shared mechanisms that support low-level visual processing. This finding complements previous research by showing that imagery is not merely perception in reverse. Instead, in line with the predictive processing framework, both, perception and imagery are active and constructive processes that share top-down mechanisms already in early phases of processing.

Sox2 modulation increases naive pluripotency plasticity

Tremble, K. — 2020-01-15

Induced pluripotency provides a tool to explore the molecular mechanisms underlying the establishment, maintenance and differentiation of naive pluripotent stem cells (nPSCs). Here, we report that self-renewal of nPSCs requires minimal Sox2 expression (Sox2-low). Sox2-low nPSCs do not show impaired neuroectoderm specification and differentiate efficiently in vitro into all embryonic germ lineages. Strikingly, Sox2-low cells also differentiate towards the trophoblast lineage both in vitro and in vivo. At the single-cell level self-renewing Sox2-low nPSCs exhibit a homogeneous naive molecular signature. However, they also display a basal trophoblast molecular signature and decreased ability of Oct4 to bind naive-associated regulatory sequences compared to control cells. These features underlie observed enhanced cell potency upon the removal of self-renewing cues. In sum, this work defines Sox2 as a restrictor of developmental potential and suggests perturbation of the naive pluripotent network as an underlying cause of increased cell potency. HighlightsO_LILow Sox2 expression is sufficient for naive pluripotent stem cell self-renewal C_LIO_LILow Sox2 expression does not impair neurectoderm differentiation in vitro C_LIO_LILow Sox2 expression impairs Oct4 genomic occupancy C_LIO_LILow Sox2 expression increases naive pluripotent cell plasticity in vitro and in vivo C_LI

Selective Postnatal Excitation of Neocortical Pyramidal Neurons Results in Distinctive Behavioral and Circuit Deficits in Adulthood

Medendorp, W. E. — 2020-01-18

In leading models of Autism Spectrum Disorder, and in human data, the efficacy of outgoing cortical connectivity transitions from overly exuberant to languid from early development to adulthood. This transition begs the question of whether the early enhancement in excitation might be a common driver, across etiologies, of these symptoms. We directly tested this concept by chemogenetically driving neuronal activity in neocortical neurons during postnatal days 4-14. Hyperexcitation of Emx1-, but not dopamine transporter-, parvalbumin-, or Dlx5/6-expressing neurons led to decreased social interaction and increased grooming activity in adult animals. In vivo optogenetic interrogation in adults revealed decreased baseline but increased stimulus-evoked firing rates of pyramidal neurons, impaired recruitment of inhibitory neurons and reduced cortico-striatal communication. These results directly support the prediction that changed firing in developing circuits irreversibly alters adult circuit function that leads to maladaptive changes in behaviors. This experimental approach offers a valuable platform to study the impact of disruption of developmental neural activity on the formation and function of adult neural circuits and behavior.

NetMix: A network-structured mixture model for reduced-bias estimation of altered subnetworks

Reyna, M. A. — 2020-01-19

A classic problem in computational biology is the identification of altered subnetworks: subnetworks of an interaction network that contain genes/proteins that are differentially expressed, highly mutated, or otherwise aberrant compared to other genes/proteins. Numerous methods have been developed to solve this problem under various assumptions, but the statistical properties of these methods are often unknown. For example, some widely-used methods are reported to output very large subnetworks that are difficult to interpret biologically. In this work, we formulate the identification of altered subnetworks as the problem of estimating the parameters of a class of probability distributions which we call the Altered Subset Distribution (ASD). We derive a connection between a popular method, jActiveModules, and the maximum likelihood estimator (MLE) of the ASD. We show that the MLE is statistically biased, explaining the large subnetworks output by jActiveModules. We introduce NetMix, an algorithm that uses Gaussian mixture models to obtain less biased estimates of the parameters of the ASD. We demonstrate that NetMix outperforms existing methods in identifying altered subnetworks on both simulated and real data, including the identification of differentially expressed genes from both microarray and RNA-seq experiments and the identification of cancer driver genes in somatic mutation data. AvailabilityNetMix is available online at https://github.com/raphael-group/netmix. Contactbraphael@princeton.edu

Changing and stable chromatin accessibility supports transcriptional overhaul during neural stem cell activation

Maybury-Lewis, S. Y. — 2020-01-29

Adult neural stem cells are largely quiescent, and require transcriptional reprogramming to reenter the cell cycle and undergo neurogenesis. However, the precise mechanisms that underlie the rapid transcriptional overhaul during NSC activation remain undefined. Here, we identify the genome-wide chromatin accessibility differences between primary neural stem and progenitor cells in quiescent and activated states. We show that these distinct cellular states exhibit both shared and unique chromatin profiles, which are both associated with gene regulation. Interestingly, we find that accessible chromatin states specific to quiescent or activated cells are active enhancers bound by pro-neurogenic and quiescence factors, ASCL1 and NFI. In contrast, shared sites are gene promoters harboring constitutively accessible chromatin enriched for particular core promoter elements that are functionally associated with translation and metabolic functions. Together, our findings reveal how accessible chromatin states regulate a transcriptional overhaul and drive the switch between quiescence and proliferation in NSC activation.

Recombinant human MG53 protein preserves mitochondria integrity in cardiomyocytes during ischemia reperfusion-induced oxidative stress

Gumpper, K. — 2020-02-07

Ischemic injury to the heart causes a loss of mitochondria function due to an increase in oxidative stress. MG53, also known as TRIM72, is highly expressed in striated muscle and is essential to repair damage to plasma membrane. We have shown that mg53-/- mice are more susceptible to ischemia-reperfusion injury, whereas treatment with exogenous recombinant human MG53 (rhMG53) reduces both infarct damage and restores cardiac function. This study assesses whether MG53 protects and repairs mitochondria injury after oxidative stress associated with myocardial infarction. We hypothesize that in addition to known cell membrane repair function, MG53 acts as a myokine to protect cardiomyocytes by maintaining mitochondrial function. A combination of in vivo and in vitro ischemia/reperfusion models were used to assess MG53s effect on mitochondria using biochemical assays and confocal microscopic imaging. Treatment with rhMG53 allowed cells to maintain a healthy mitochondrial membrane potential, reduced release of mitochondrial reactive oxygen species, and mitigated mitophagy. Mitochondrial localization of rhMG53 is mediated by exposure of and interaction with cardiolipin on the mitochondrial membrane. Our data demonstrates that rhMG53 protein preserves mitochondria integrity in cardiomyocytes during ischemia reperfusion-induced oxidative stress.

Discriminating between sleep and exercise-induced fatigue using computer vision and behavioral genetics

Schuch, K. N. — 2020-02-07

Following prolonged swimming, Caenorhabditis elegans cycle between active swimming bouts and inactive quiescent bouts. Swimming is exercise for C. elegans and here we suggest that inactive bouts are a recovery state akin to fatigue. Previously, analysis of exercise-induced quiescent (EIQ) bouts relied on laborious manual observation, as existing automated analysis methods for C. elegans swimming either cannot analyze EIQ bouts or fail to accurately track animal posture during these bouts. It is known that cGMP-dependent kinase (PKG) activity plays a conserved role in sleep, rest, and arousal. Using C. elegans EGL-4 PKG, we first validate a novel learning-based computer vision approach to automatically analyze C. elegans locomotory behavior and distinguish between activity and inactivity during swimming for long periods of time. We find that C. elegans EGL-4 PKG function predicts EIQ first bout timing, fractional quiescence, bout number, and bout duration, suggesting that previously described pathways are engaged during EIQ bouts. However, EIQ bouts are likely not sleep as animals are feeding during the majority of EIQ bouts. We find that genetic perturbation of neurons required for other C. elegans sleep states also does not alter EIQ dynamics. Additionally, we find that EIQ onset is sensitive to age and DAF-16 FOXO function. In summary, we have validated a new behavioral analysis software that enabled a quantitative and detailed assessment of swimming behavior, including EIQ. We found novel EIQ defects in aged animals and animals with mutations in a gene involved in stress tolerance. We anticipate that further use of this software will facilitate the analysis of genes and pathways critical for fatigue and other C. elegans behaviors.

Abstract sequential task control is facilitated by practice and embedded motor sequences

Trach, J. E. — 2020-02-07

Everyday task sequences, such as cooking, contain overarching goals (completing the meal), sub-goals (prepare vegetables), and motor actions (chopping). Such tasks generally are considered hierarchical because superordinate levels (e.g., goals) affect performance at subordinate levels (e.g., sub-goals and motor actions). However, there is debate as to whether this hierarchy is "strict" with unidirectional, top-down influences, and it is unknown if and how practice affects performance at the superordinate levels. To investigate these questions, we manipulated practice with sequences at the goal and motor action levels using an abstract, or non-motor, task sequence paradigm (Desrochers et al., 2015; Schneider & Logan, 2006). In three experiments, participants performed memorized abstract task sequences composed of simple tasks (e.g., color/shape judgements), where some contained embedded motor response sequences. We found that practice facilitated performance and reduced control costs for abstract task sequences and subordinate tasks. The interrelation was different between the hierarchical levels, demonstrating a strict relationship between abstract task sequence goals and sub-goals and a non-strict relationship between sub-goal and motor response levels. Under some conditions, the motor response level influenced the abstract task sequence level in a non-strict manner. Further, manipulating the presence or absence of a motor sequence after learning indicated that these effects were not the result of an integrated representation produced by practice. These experiments provide evidence for a mixed hierarchical model of task sequences and insight into the distinct roles of practice and motor processing in efficiently executing task sequences in daily life.

Chronic wireless streaming of invasive neural recordings at home for circuit discovery and adaptive stimulation

Gilron, R. — 2020-02-14

Invasive neural recording in humans shows promise for understanding the circuit basis of brain disorders. Most recordings have been done for short durations from externalized brain leads in hospital settings, or from first-generation implantable sensing devices that offer only intermittent brief streaming of time series data. Here we report the first human use of an implantable neural interface for wireless multichannel streaming of field potentials over long periods, with and without simultaneous therapeutic neurostimulation, untethered to receiving devices. Four Parkinsons disease patients streamed bilateral 4-channel motor cortical and basal ganglia field potentials at home for over 500 hours, paired with wearable monitors that behaviorally categorize states of inadequate or excessive movement. Motor state during normal home activities was efficiently decoded using either supervised learning or unsupervised clustering algorithms. This platform supports adaptive deep brain stimulation, and may be widely applicable to brain disorders treatable by invasive neuromodulation.

Behavioral features of motivated response to alcohol in Drosophila

Catalano, J. L. — 2020-02-17

Animals avoid predators and find the best food and mates by learning from the consequences of their behavior. However, reinforcers are not always uniquely appetitive or aversive but can have complex properties. Most intoxicating substances fall within this category; provoking aversive sensory and physiological reactions while simultaneously inducing overwhelming appetitive properties. Here we describe the subtle behavioral features associated with continued seeking for alcohol despite aversive consequences. We developed an automated runway apparatus to measure how Drosophila respond to consecutive exposures of a volatilized substance. Behavior within this Behavioral Expression of Ethanol Reinforcement Runway (BEER Run) demonstrated a defined shift from aversive to appetitive responses to volatilized ethanol. Behavioral metrics attained by combining computer vision and machine learning methods, reveal that a subset of 9 classified behaviors and component behavioral features associate with this shift. We propose this combination of 9 behaviors can be used to navigate the complexities of operant learning to reveal motivated goal-seeking behavior.

ASH1L REGULATES THE STRUCTURAL DEVELOPMENT OF NEURONAL CIRCUITRY BY MODULATING BDNF/TrkB SIGNALING IN HUMAN NEURONS

Cheon, S. H. — 2020-02-19

Autism spectrum disorders (ASD) are associated with defects in neuronal connectivity and are highly heritable. Genetic findings suggest that there is an overrepresentation of chromatin regulatory genes among the genes associated with ASD. ASH1 like histone lysine methyltransferase (ASH1L) was identified as a major risk factor for autism. ASH1L methylates Histone H3 on Lysine 36, which is proposed to result primarily in transcriptional activation. However, how mutations in ASH1L lead to deficits in neuronal connectivity associated with autism pathogenesis is not known. We report that ASH1L regulates neuronal morphogenesis by counteracting the catalytic activity of Polycomb Repressive complex 2 group (PRC2) in stem cell-derived human neurons. Depletion of ASH1L decreases neurite outgrowth and decreases expression of the gene encoding the neurotrophin receptor TrkB whose signaling pathway is linked to neuronal morphogenesis. This is overcome by inhibition of PRC2 activity, indicating a balance between the Trithorax group protein ASH1L and PRC2 activity determines neuronal morphology and connectivity. Thus, ASH1L epigenetically regulates neuronal connectivity by modulating the BDNF-TrkB signaling pathway, which likely contributes to the neurodevelopmental pathogenesis associated with ASD in patients with ASH1L mutations. eTOC BLURBCheon et al. report a novel epigenetic mechanism that implicates the counteracting activities of the evolutionarily conserved Trithorax (ASH1L) and Polycomb (PRC2) chromatin regulators, in the modulation of human neuronal connectivity by regulating the developmentally important TrkB-BDNF signaling pathway. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/954586v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@738ee5org.highwire.dtl.DTLVardef@aa8085org.highwire.dtl.DTLVardef@1dbc315org.highwire.dtl.DTLVardef@e4bfa2_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIASH1L regulates neuronal morphogenesis by modulating neurotrophin signaling C_LIO_LICounteracting activities of Trithorax (ASH1L) and Polycomb (PRC2) affect neuronal arborization C_LIO_LILoss of ASH1L modulates growth cone size in human neurons C_LI

Genetic Variants and Functional Pathways Associated with Resilience to Alzheimer's Disease

Dumitrescu, L. — 2020-02-21

Approximately 30% of older adults exhibit the neuropathologic features of Alzheimers disease (AD) without signs of cognitive impairment. Yet, little is known about the genetic factors that allow these potentially resilient individuals to remain cognitively normal in the face of substantial neuropathology. We performed a large, genome-wide association study (GWAS) of two previously validated metrics of cognitive resilience quantified using a latent variable modeling approach and representing better-than-predicted cognitive performance for a given level of neuropathology. Data were harmonized across 5,108 participants from a clinical trial of AD and three longitudinal cohort studies of cognitive aging. All analyses were run across all participants and repeated restricting the sample to individuals with normal cognition to identify variants at the earliest stages of disease. As expected, all resilience metrics were genetically correlated with cognitive performance and education attainment traits (p-values<2.5x10-20), and we observed novel correlations with neuropsychiatric conditions (p-values<7.9x10-4). Notably, neither resilience metric was genetically correlated with clinical AD (p-values>0.42) nor associated with APOE (p-values>0.13). In single variant analyses, we observed a genome-wide significant locus among participants with normal cognition on chromosome 18 upstream of ATP8B1 (index SNP rs2571244, MAF=0.08, p=2.3x10-8). The top variant at this locus (rs2571244) was significantly associated with methylation in prefrontal cortex tissue at multiple CpG sites, including one just upstream of ATPB81 (cg19596477; p=2x10-13). Overall, this comprehensive genetic analysis of resilience implicates a putative role of vascular risk, metabolism, and mental health in protection from the cognitive consequences of neuropathology, while also providing evidence for a novel resilience gene along the bile acid metabolism pathway. Furthermore, the genetic architecture of resilience appears to be distinct from that of clinical AD, suggesting that a shift in focus to molecular contributors to resilience may identify novel pathways for therapeutic targets.

Automated identification of multinucleated germ cells with U-Net

Bell, S. — 2020-02-20

Phthalic acid esters (phthalates) are male reproductive toxicants, which exert their most potent toxicity during a critical window of sensitivity in fetal development. In the fetal rat, exposure to phthalates reduces testosterone biosynthesis, alters the development of seminiferous cords and other male reproductive tissues, and induces the formation of abnormal multinucleated germ cells (MNGs). Identification of MNGs is a time-intensive process, and it requires specialized training to identify MNGs in histological sections. As a result, MNGs are not routinely quantified in phthalate toxicity experiments. In order to speed up and standardize this process, we have developed an improved method for automated detection of MNGs. Using hand-labeled histological section images with human-identified MNGs, we trained a convolutional neural network with a U-Net architecture to identify MNGs on unlabeled images. With unseen hand-labeled images not used in model training, we assessed the performance of the model, using five different configurations of the data. On average, the model reached near human accuracy, and in the best model, it exceeded it. The use of automated image analysis will allow data on this histopathological endpoint to be more readily collected for analysis of phthalate toxicity. Our trained model application code is available for download at github.com/brown-ccv/mngcount.

Single-molecule sequencing of long DNA molecules allows high contiguity de novo genome assembly for the fungus fly, Sciara coprophila

Urban, J. M. — 2020-02-25

The lower Dipteran fungus fly, Sciara coprophila, has many unique biological features. For example, Sciara undergoes paternal chromosome elimination and maternal X chromosome nondisjunction during spermatogenesis, paternal X elimination during embryogenesis, intrachromosomal DNA amplification of DNA puff loci during larval development, and germline-limited chromosome elimination from all somatic cells. Paternal chromosome elimination in Sciara was the first observation of imprinting, though the mechanism remains a mystery. Here, we present the first draft genome sequence for Sciara coprophila to take a large step forward in aiding these studies. We approached assembling the Sciara genome using multiple sequencing technologies: PacBio, Oxford Nanopore MinION, and Illumina. To find an optimal assembly using these datasets, we generated 44 Illumina assemblies using 7 short-read assemblers and 50 long-read assemblies of PacBio and MinION sequence data using 6 long-read assemblers. We ranked assemblies using a battery of reference-free metrics, and scaffolded a subset of the highest-ranking assemblies using BioNano Genomics optical maps. RNA-seq datasets from multiple life stages and both sexes facilitated genome annotation. Moreover, we anchored nearly half of the Sciara genome sequence into chromosomes. Finally, we used the signal level of both the PacBio and Oxford Nanopore data to explore the presence or absence of DNA modifications in the Sciara genome since DNA modifications may play a role in imprinting in Sciara, as they do in mammals. These data serve as the foundation for future research by the growing community studying the unique features of this emerging model system.

Sensitive, highly multiplexed sequencing of microhaplotypes from the Plasmodium falciparum heterozygome

Tessema, S. K. — 2020-02-26

BackgroundTargeted next generation sequencing offers the potential for consistent, deep coverage of information rich genomic regions to characterize polyclonal Plasmodium falciparum infections. However, methods to identify and sequence these genomic regions are currently limited. MethodsA bioinformatic pipeline and multiplex methods were developed to identify and simultaneously sequence 100 targets and applied to dried blood spot (DBS) controls and field isolates from Mozambique. For comparison, WGS data were generated for the same controls. ResultsUsing publicly available genomes, 4465 high diversity genomic regions suited for targeted sequencing were identified, representing the P. falciparum heterozygome. For this study, 93 microhaplotypes with high diversity (median HE = 0.7) were selected along with 7 drug resistance loci. The sequencing method achieved very high coverage (median 99%), specificity (99.8%) and sensitivity (90% for haplotypes with 5% within sample frequency in DBS with 100 parasites/{micro}L). In silico analyses revealed that microhaplotypes provided much higher resolution to discriminate related from unrelated polyclonal infections than biallelic SNP barcodes. DiscussionThe bioinformatic and laboratory methods outlined here provide a flexible tool for efficient, low-cost, high throughput interrogation of the P. falciparum genome, and can be tailored to simultaneously address multiple questions of interest in various epidemiological settings.

In search of the RNA world on Mars

Mojarro, A. — 2020-02-28

Advances in origins of life research and prebiotic chemistry suggest that life as we know it may have emerged from an earlier RNA World. However, it has been difficult to reconcile the conditions used in laboratory experiments with real-world geochemical environments that may have existed on the early Earth and hosted the origin(s) of life. This challenge is in part due to geologic resurfacing and recycling that have erased the overwhelming majority of the Earths prebiotic history. We therefore propose that Mars, a planet frozen in time, comprised of many surfaces that have remained relatively unchanged since their formation >4 Gya, is the best alternative to search for environments consistent with geochemical requirements imposed by the RNA world. In this study we synthesize in situ and orbital observations of Mars and modeling of its early atmosphere into solutions containing a range of pHs and concentrations of prebiotically relevant metals (Fe2+, Mg2+, and Mn2+), spanning various candidate aqueous environments. We then experimentally determine RNA degradation kinetics due to metal-catalyzed hydrolysis and evaluate whether early Mars could have been permissive towards the accumulation of long-lived RNA polymers. Our results indicate that a Mg2+-rich basalt sourcing metals to a slightly acidic (pH 5.4) aqueous environment mediates the slowest rates of metal-catalyzed RNA hydrolysis, though geologic evidence and modeling of basalt weathering suggest that aquifers on Mars would be near neutral (pH [~]7). Moreover, oxidizing conditions on Mars have major consequences regarding the availability oxygen-sensitive prebiotic metals (i.e., Fe2+ and Mn2+) very early in its history due to increased RNA degradation rates and precipitation. Overall, 1) low pH better preserves RNA than basic conditions at high concentrations; 2) acidic to neutral pH environments with Fe2+ or Mn2+ will hydrolyze more RNA; and 3) alkaline environments with Mg2+ dramatically hydrolyze more RNA.

SLC45A2 protein stability and regulation of melanosome pH determine melanocyte pigmentation

Marks, M. S. — 2020-03-04

SLC45A2 encodes a putative transporter expressed primarily in pigment cells. SLC45A2 mutations and polymorphisms cause oculocutaneous albinism (OCA) and pigmentation variation, but neither SLC45A2 localization and function nor how gene variants affect these properties are known. We show that SLC45A2 localizes to mature melanosomes that only partially overlap with a cohort expressing the chloride channel OCA2. SLC45A2 expressed ectopically in HeLa cells localizes to lysosomes and raises lysosomal pH, suggesting that, like OCA2, SLC45A2 in melanocytes de-acidifies maturing melanosomes to support melanin synthesis. Analyses of SLC45A2- and OCA2-deficient mouse melanocytes show that SLC45A2 functions later during melanosome maturation than OCA2, and that OCA2 overexpression compensates for loss of SLC45A2 expression in pigmentation. The light skin-associated SLC45A2 allelic F374 variant restores only moderate pigmentation to SLC45A2-deficient melanocytes because of low level expression in melanosomes due to rapid proteasome-independent degradation. Our data indicate that SLC45A2 maintains melanosome neutralization - initially orchestrated by transient OCA2 activity - to support melanization at late stages of melanosome maturation, and that a common variant imparts reduced activity due to protein instability.

Differentiation drives widespread rewiring of the neural stem cell chaperone network

Vonk, W. I. M. — 2020-03-06

Neural stem and progenitor cells (NSPCs) are critical for continued cellular replacement in the adult brain. Life-long maintenance of a functional NSPC pool necessitates stringent mechanisms to preserve a pristine proteome. We find that the NSPCs chaperone network robustly maintains misfolded protein solubility and stress resilience through high levels of the ATP-dependent chaperonin TRiC/CCT. Strikingly, NSPC differentiation rewires the cellular chaperone network, reducing TRiC/CCT levels and inducing those of the ATP-independent small heat shock proteins (sHSPs). This switches the proteostasis strategy in neural progeny cells to promote sequestration of misfolded proteins into protective inclusions. The chaperone network of NSPCs is more effective than that of differentiated cells, leading to improved management of proteotoxic stress and amyloidogenic proteins. However, NSPC proteostasis is impaired by brain aging. The less efficient chaperone network of differentiated neural progeny may contribute to their enhanced susceptibility to neurodegenerative diseases characterized by aberrant protein misfolding and aggregation.

A Method for Large Scale Implantation of 3D Microdevice Ensembles into Brain and Soft Tissue

Sigurdsson, S. A. — 2020-03-08

Wireless networks of implantable electronic sensors and actuators on the microscale (sub-mm) are being explored for monitoring and modulation of physiological activity for medical diagnostics and therapeutic purposes. Beyond the requirement of integrating multiple electronic or chemical functions within small device volumes, a key challenge is the development of high-throughput methods for implantation of large numbers of microdevices into soft tissues with minimal damage. To that end, we have developed a method for high-throughput implantation of ∼100-200 μm size devices which are here simulated by proxy microparticle ensembles. While generally applicable to subdermal tissue, our main focus and experimental testbed is the implantation of microparticles into the brain. The method deploys a scalable delivery tool composed of a 2-dimensional array of polyethylene glycol tipped microneedles which confine the microparticle payloads. Upon dissolution of the bioresorbable polyethylene glycol, the supporting array structure is retrieved and the microparticles remain embedded in the tissue, distributed spatially and geometrically according to the design of the microfabricated delivery tool. We first evaluated the method in an agarose testbed in terms of spatial precision and throughput for up to 1000 passive spherical and planar microparticles acting as proxy devices. We then performed the same evaluations of particles implanted into the rat cortex under acute conditions and assessed the tissue injury produced by our method of implantation under chronic conditions.Competing Interest StatementThe authors have declared no competing interest.View Full Text

Subventricular zone adult mouse neural stem cells and human glioblastoma stem cells require insulin receptor for self-renewal

Chidambaram, S. — 2020-03-11

The insulin receptor (IR) is an evolutionarily conserved signaling protein that regulates development and cellular metabolism. IR signaling regulates neurogenesis in Drosophila; however, a specific role for the IR in maintaining adult neural stem cells (NSCs) in mammals has not been investigated. We show that conditionally deleting the IR reduces adult NSCs of the subventricular zone by [~]70% accompanied by a corresponding increase in progenitors. IR deletion produced hyposmia due to aberrant olfactory bulb neurogenesis. Interestingly, hippocampal neurogenesis was not perturbed nor were hippocampal dependent behaviors. Highly aggressive proneural and mesenchymal glioblastomas (GBMs) had high IR/insulin-like growth factor (IGF) pathway gene expression, and isolated glioma stem cells had an aberrantly high ratio of IR:IGF1R receptors. Moreover, IR knockdown inhibited proneural and mesenchymal GBM tumorsphere growth. Altogether, these data demonstrate that the IR is essential for a subset of normal NSCs as well as for brain tumor cancer stem cell self-renewal.

VarStack: a Web Tool for Data Retrieval to Interpret Somatic Variants in Cancer

Howard, M. F. — 2020-03-11

Background and objectiveAdvances in tumor genome sequencing created an urgent need for bioinformatics tools to support the interpretation of the clinical significance of the variants detected. VarStack is a web tool which is a base to retrieve somatic variant data in cancer from existing databases. MethodsVarStack incorporates data from several publicly available databases and presents them with an easy-to-navigate user-interface. It currently supports data from the Catalogue of Somatic Mutations in Cancer (COSMIC), gnomAD, cBioPortal, ClinVar, OncoKB and UCSC Genome browser. It retrieves the data from these databases and returns back to the user in a fraction of the time it would take to manually navigate each site independently. ResultsUsers submit a variant with gene symbol, peptide change, and coding sequence change. They may select a variety of tumor specific studies in cBioportal to search through in addition to their original query. The results from the databases are presented in tabs. Users can export the results as a CSV file. VarStack also has the batch search feature in which user submits a list of variants and download a CSV file with the data from the databases. With the batch search and data download options users can easily incorporate VarStack into their workflow or tools. VarStack saves time by providing variant data to the user from multiple databases in an easy-to-export and interpretable format. AvailabilityVarStack is freely available under https://varstack.brown.edu.

Mapping the Structural and Dynamic Determinants of pH-sensitive Heparin Binding to Granulocyte Macrophage-colony Stimulating Factor

Cui, J. Y. — 2020-03-11

GMCSF is an immunomodulatory cytokine that is harnessed as a therapeutic. GMCSF is known to interact with other clinically important molecules, such as heparin, suggesting that endogenous and administered GMCSF has the potential to modulate orthogonal treatment outcomes. Thus, molecular level characterization of GMCSF and its interactions with other biologically active compounds is critical to understanding these mechanisms and predicting clinical outcomes. Here, we dissect the molecular motions and structural contributions that facilitate the GMCSF-heparin interaction, previously shown to be pH-dependent, using NMR spectroscopy, SPR, and molecular docking. We find that GMCSF and heparin binding is related to a change in flexibility reflected in the dynamic profile of GMCSF at acidic pH. The molecular motions driving this interaction largely occur on the ms-{micro}s timescale. Interestingly, we find that GMCSF and heparin binding is not only pH-dependent but is also heparin chain length dependent. We propose a mechanism where a positive binding pocket that is not fully solvent accessible at neutral pH becomes more accessible at acidic pH, allowing heparin to dock with the protein.

Identification of a molecular fingerprint for synaptic glia

Castro, R. — 2020-03-14

The inability to specifically identify and manipulate synaptic glial cells remains a major obstacle to understanding fundamental aspects of synapse formation, stability and repair. Using a combinatorial gene expression approach, we discovered molecular markers that allow us to specifically label perisynaptic Schwann cells (PSCs), glial cells at neuromuscular synapses. Using these markers, we demonstrate that PSCs fully-differentiate postnatally and have a unique molecular signature that includes genes predicted and known to play critical roles at synapses. These findings will serve as a springboard for unprecedented approaches for studying molecular determinants of PSC differentiation and function at neuromuscular synapses and possibly synapse-associated glia throughout the CNS.

Tyrosine phosphorylation regulates hnRNPA2 granule protein partitioning & reduces neurodegeneration

Ryan, V. H. — 2020-03-15

mRNA transport in neurons is a ubiquitous process but has been often overlooked as a contributor to disease. Mutations of transport granule protein hnRNPA2 cause hereditary proteinopathy of neurons, myocytes, and bone. Here, we examine transport granule component specificity, assembly/disassembly, and the link to neurodegeneration. hnRNPA2 transport granule components hnRNPF and ch-TOG interact weakly with hnRNPA2 yet they each partition specifically into hnRNPA2 liquid phases. hnRNPA2 tyrosine phosphorylation dissociates granule interactions by reducing hnRNPA2 phase separation and preventing partitioning of hnRNPF and ch-TOG; tyrosine phosphorylation also decreases aggregation of hnRNPA2 disease mutants. A C. elegans model of hnRNPA2 D290V-associated neurodegeneration exhibits TDP-43 ortholog-dependent glutamatergic neurodegeneration. Expression of the tyrosine kinase that phosphorylates hnRNPA2 reduces glutamatergic neurodegeneration. The evidence for specific partitioning of granule components as well as disruption of these interactions and reduction of neurodegeneration by tyrosine phosphorylation suggest transport granule biology has a role in the pathogenesis of neurodegeneration.

Quantitative properties of the creation and activation of a cell-intrinsic engram

Gallistel, C. R. — 2020-03-18

The conditional pause in the spontaneous firing of the cerebellar Purkinje, which determines the timing of the conditional eyeblink response, is mediated by a cell-intrinsic engram (Johansson, et al. 2014) that encodes the interstimulus interval. Our trial-by-trial analysis of the pause parameters reveals that it consists of a single unusually long interspike interval, whose onset and offset latencies are stochastically independent scalar functions of the interstimulus interval. The coefficients of variation are comparable to those observed in the timing of the overt conditional eyeblink. The onsets of the long interspike interval are step changes; there is no prior build-up of inhibition. A single spike volley in the parallel fiber input triggers the read-out of the engram into the long interspike interval; subsequent volleys have no effect on the pause. The high spontaneous firing rate on which the one-interval firing pause supervenes is markedly non-stationary (Fano factors >> 1).

The microRNA, miR-133b, functions to slow Duchenne muscular dystrophy pathogenesis

Taetzsch, T. — 2020-03-20

Duchenne muscular dystrophy (DMD) is characterized by progressive degeneration of skeletal muscles. To date, there are no treatments available to slow or prevent the disease. Hence, it remains essential to identify molecular factors that promote muscle biogenesis since they could serve as therapeutic targets for treating DMD. While the muscle enriched microRNA, miR-133b, has been implicated in the biogenesis of muscle fibers, its role in DMD remains unknown. To assess the role of miR-133b in DMD-affected skeletal muscles, we genetically ablated miR-133b in the mdx mouse model of DMD. In the absence of miR-133b, the tibialis anterior muscle of juvenile and adult mdx mice is populated by small muscle fibers with centralized nuclei, exhibits increased fibrosis, and thickened interstitial space. Additional analysis revealed that loss of miR-133b exacerbates DMD-pathogenesis partly by altering the number of satellite cells and levels of protein-encoding genes, including previously identified miR-133b targets as well as genes involved in cell proliferation and fibrosis. Altogether, our data demonstrate that skeletal muscles utilize miR-133b to mitigate the deleterious effects of DMD.

Channel Embedding for Informative Protein Identification from Highly Multiplexed Images

Magid, S. A. — 2020-03-25

Interest is growing rapidly in using deep learning to classify biomedical images, and interpreting these deep-learned models is necessary for life-critical decisions and scientific discovery. Effective interpretation techniques accelerate biomarker discovery and provide new insights into the etiology, diagnosis, and treatment of disease. Most interpretation techniques aim to discover spatially-salient regions within images, but few techniques consider imagery with multiple channels of information. For instance, highly multiplexed tumor and tissue images have 30-100 channels and require interpretation methods that work across many channels to provide deep molecular insights. We propose a novel channel embedding method that extracts features from each channel. We then use these features to train a classifier for prediction. Using this channel embedding, we apply an interpretation method to rank the most discriminative channels. To validate our approach, we conduct an ablation study on a synthetic dataset. Moreover, we demonstrate that our method aligns with biological findings on highly multiplexed images of breast cancer cells while outperforming baseline pipelines.

Assessment of a weak mode of bacterial adhesion by applying an electric field

Tang, J. X. — 2020-04-02

Microbial attachment to surfaces is ubiquitous in nature. Most species of bacteria attach and adhere to surfaces via special appendages such as pili and fimbriae, the roles of which have been extensively studied. Here we report an experiment on pilus-less mutants of Caulobacter crescentus weakly attached to a plastic surface and subjected to an electric field parallel to the surface. We find that some individual cells transiently but repeatedly adhere to the surface in a stick-slip fashion in the presence of an electric field. Even while transiently detached, these bacteria move significantly slower than the unattached ones in the same field of view undergoing electrophoretic motion. We refer this behavior of repeated and transient attachment as "quasi-attachment". The speed of the quasi-attached bacteria exhibits large variations, frequently dropping close to zero for short intervals of time. This study suggests applying electric field as a useful method to investigate bacteria-surface interaction, which is significant in broader contexts such as infection and environmental control. SignificanceInteraction between bacteria and surfaces occur widely in nature, including those in industrial, environmental, and medical settings. It is therefore important to understand various mechanisms and factors that affect numerous forms of bacterium-surface interaction, particularly those resulting in adhesion or attachment, be they strong or weak, permanent or transient. This work takes a unique approach to identify a transient and reversible mode of bacterial attachment to a solid surface, by applying an electric field to exert a force for detachment. The force thus exerted proves to reach the amplitude required to detach bacteria of a pilus-less strain that weakly attach to a plastic surface. The method may be applied broadly to investigate bacteria-surface interaction.

Integrating XMALab and DeepLabCut for high-throughput XROMM

Laurence-Chasen, J. D. — 2020-04-13

Marker tracking is a major bottleneck in studies involving X-ray Reconstruction of Moving Morphology (XROMM). Here, we tested whether DeepLabCut, a new deep learning package built for markerless tracking, could be applied to videoradiographic data to improve data processing throughput. Our novel workflow integrates XMALab, the existing XROMM marker tracking software, and DeepLabCut while retaining each programs utility. XMALab is used for generating training datasets, error correction, and 3D reconstruction, whereas the majority of marker tracking is transferred to DeepLabCut for automatic batch processing. In the two case studies that involved an in vivo behavior, our workflow achieved a 6 to 13-fold increase in data throughput. In the third case study, which involved an acyclic, post mortem manipulation, DeepLabCut struggled to generalize to the range of novel poses and did not surpass the throughput of XMALab alone. Deployed in the proper context, this new workflow facilitates large scale XROMM studies that were previously precluded by software constraints.

Selective enhancement of low-gamma activity by tACS improves phonemic processing and reading accuracy in dyslexia

Marchesotti, S. — 2020-04-17

The phonological deficit in dyslexia is associated with altered low-gamma oscillatory function in left auditory cortex, but a causal relationship between oscillatory function and phonemic processing has never been established. After confirming a deficit at 30 Hz with electroencephalography (EEG), we applied 20 minutes of transcranial alternating current stimulation (tACS) to transiently restore this activity in adults with dyslexia. The intervention significantly improved phonological processing and reading accuracy as measured immediately after tACS. The effect was selective to 30 Hz stimulation, and proportional to dyslexia severity. Importantly, we observed that the focal intervention on the left auditory cortex also decreased 30 Hz activity in the right superior temporal cortex, resulting in reinstating a left dominance for the oscillatory response, as present in controls. These findings formally establish a causal role of neural oscillations in phonological processing, and offer solid neurophysiological grounds for a potential correction of low-gamma anomalies, and for alleviating of the phonological deficit in dyslexia.

Predicting the Viability of Beta-Lactamase: How Folding and Binding Free Energies Correlate with Beta-Lactamase Fitness

Yang, J. — 2020-04-16

One of the long-standing holy grails of molecular evolution has been the ability to predict an organisms fitness directly from its genotype. With such predictive abilities in hand, researchers would be able to more accurately forecast how organisms will evolve and how proteins with novel functions could be engineered, leading to revolutionary advances in medicine and biotechnology. In this work, we assemble the largest reported set of experimental TEM-1 {beta}-lactamase folding free energies and use this data in conjunction with previously acquired fitness data and computational free energy predictions to determine how much of the fitness of {beta}-lactamase can be directly predicted by thermodynamic folding and binding free energies. We focus upon {beta}-lactamase because of its long history as a model enzyme and its central role in antibiotic resistance. Based upon a set of 21 {beta}-lactamase single and double mutants expressly designed to influence protein folding, we first demonstrate that modeling software such as FoldX and PyRosetta designed to compute folding free energies can meaningfully, although not perfectly, predict the experimental folding free energies of single mutants. Interestingly, while these techniques also yield sensible double mutant free energies, we show that they do so for the wrong physical reasons. We then go on to assess how well both experimental and computational folding free energies explain single mutant fitness. We find that folding free energies account for, at most, 24% of the variance in {beta}-lactamase fitness values according to linear models and, somewhat surprisingly, complementing folding free energies with computationally-predicted binding free energies of residues near the active site only increases the folding-only figure by a few percent. This strongly suggests that the majority of {beta}-lactamases fitness is controlled by factors other than free energies. Overall, our results shed a bright light on to what extent the community is justified in using thermodynamic measures to infer protein fitness as well as how applicable modern computational techniques for predicting free energies will be to the large data sets of multiply-mutated proteins forthcoming.

Two dynamically distinct circuits driving inhibition in sensory thalamus

Cruikshank, S. J. — 2020-04-18

Most sensory information destined for the neocortex is relayed through the thalamus, where considerable transformation occurs1,2. One powerful means of transformation involves interactions between excitatory thalamocortical neurons that carry data to cortex and inhibitory neurons of the thalamic reticular nucleus (TRN) that regulate flow of those data3-6. Despite enduring recognition of its importance7-9, understanding of TRN cell types, their organization, and their functional properties has lagged that of the thalamocortical systems they control. Here we address this, investigating somatosensory and visual circuits of the TRN. In the somatosensory TRN we observed two groups of genetically defined neurons that are topographically segregated, physiologically distinct, and connect reciprocally with independent thalamocortical nuclei via dynamically divergent synapses. Calbindin-expressing cells, located in the central core, connect with the ventral posterior nucleus (VP), the primary somatosensory thalamocortical relay. In contrast, somatostatin-expressing cells, residing along the surrounding edges of TRN, synapse with the posterior medial thalamic nucleus (POM), a higher-order structure that carries both top-down and bottom-up information10-12. The two TRN cell groups process their inputs in pathway-specific ways. Synapses from VP to central TRN cells transmit rapid excitatory currents that depress deeply during repetitive activity, driving phasic spike output. Synapses from POM to edge TRN cells evoke slower, less depressing excitatory currents that drive more persistent spiking. Differences in intrinsic physiology of TRN cell types, including state-dependent bursting, contribute to these output dynamics. Thus, processing specializations of two somatosensory TRN subcircuits appear to be tuned to the signals they carry--a primary central subcircuit to discrete sensory events, and a higher-order edge subcircuit to temporally distributed signals integrated from multiple sources. The structure and function of visual TRN subcircuits closely resemble those of the somatosensory TRN. These results provide fundamental insights about how subnetworks of TRN neurons may differentially process distinct classes of thalamic information.

Taxonomy of neural oscillation events in primate auditory cortex

Neymotin, S. A. — 2020-04-18

Electrophysiological oscillations in the brain have been shown to occur as multi-cycle events, with onset and offset dependent on behavioral and cognitive state. To provide a baseline for state-related and task-related events, we quantified oscillation features in resting-state recordings. We used two invasively-recorded electrophysiology datasets: one from human, and one from non-human primate auditory system. After removing incidentally occuring event related potentials, we used a wavelet transform based method to quantify oscillation features. We identified about 2 million oscillation events, classified within traditional frequency bands: delta, theta, alpha, beta, low gamma, gamma, high gamma. Oscillation events of 1-44 cycles were present in at least one frequency band in 90% of the time in human and non-human primate recordings. Individual oscillation events were characterized by non-constant frequency and amplitude. This result naturally contrasts with prior studies which assumed such constancy, but is consistent with evidence from event-associated oscillations. We measured oscillation event duration, frequency span, and waveform shape. Oscillations tended to exhibit multiple cycles per event, verifiable by comparing filtered to unfiltered waveforms. In addition to the clear intra-event rhythmicity, there was also evidence of inter-event rhythmicity within bands, demonstrated by finding that coefficient of variation of interval distributions and Fano Factor measures differed significantly from a Poisson distribution assumption. Overall, our study demonstrates that rhythmic, multi-cycle oscillation events dominate auditory cortical dynamics.

Rapid 'multi-directed' cholinergic transmission at central synapses

Sethuramanujam, S. — 2020-04-20

Acetylcholine (ACh) is a key neurotransmitter that plays diverse roles in many parts of the central nervous system, including the retina. However, assessing the precise spatiotemporal dynamics of ACh is technically challenging and whether ACh transmits signals via rapid, point-to-point synaptic mechanisms, or broader-scale non-synaptic mechanisms has been difficult to ascertain. Here, we examined the properties of cholinergic transmission at individual contacts made between direction-selective starburst amacrine cells and downstream ganglion cells in the retina. Using a combination of electrophysiology, serial block-face electron microscopy, and two-photon ACh imaging, we demonstrate that ACh signaling bears the hallmarks of both non-synaptic and synaptic forms of transmission. ACh co-activates nicotinic ACh receptors located on the intersecting dendrites of pairs of ganglion cells, with equal efficiency (non-synaptic)-- and yet retains the ability to generate rapid miniature currents ([~]1 ms rise times: synaptic). Fast cholinergic signals do not appear to depend on anatomically well-defined synaptic structures. We estimate that ACh spread is limited to [~]1-2 {micro}m from its sites of release, which may help starbursts drive local direction-selective cholinergic responses in ganglion cell dendrites. Together, our results establish the functional architecture for cholinergic signaling at a central synapse and propose a novel motif whereby single presynaptic sites can co-transmit information to multiple neurons on a millisecond timescale.

FastSK: Fast Sequence Analysis with Gapped String Kernels

Blakely, D. — 2020-04-23

Gapped k-mer kernels with Support Vector Machines (gkm-SVMs) have achieved strong predictive performance on regulatory DNA sequences on modestly-sized training sets. However, existing gkm-SVM algorithms suffer from slow kernel computation time, as they depend exponentially on the sub-sequence feature-length, number of mismatch positions, and the tasks alphabet size. In this work, we introduce a fast and scalable algorithm for calculating gapped k-mer string kernels. Our method, named FastSK, uses a simplified kernel formulation that decomposes the kernel calculation into a set of independent counting operations over the possible mismatch positions. This simplified decomposition allows us to devise a fast Monte Carlo approximation that rapidly converges. FastSK can scale to much greater feature lengths, allows us to consider more mismatches, and is performant on a variety of sequence analysis tasks. On 10 DNA transcription factor binding site (TFBS) prediction datasets, FastSK consistently matches or outperforms the state-of-the-art gkmSVM-2.0 algorithms in AUC, while achieving average speedups in kernel computation of [~]100x and speedups of [~]800x for large feature lengths. We further show that FastSK outperforms character-level recurrent and convolutional neural networks across all 10 TFBS tasks. We then extend FastSK to 7 English-language medical named entity recognition datasets and 10 protein remote homology detection datasets. FastSK consistently matches or outperforms these baselines. Our algorithm is available as a Python package and as C++ source code1.

Convergent evolution of p38/MAPK activation in hormone resistant prostate cancer mediates pro-survival, immune evasive, and metastatic phenotypes

Ware, K. E. — 2020-04-24

Adaptation of cancer cells to targeted therapy follows ecological paradigms observed in natural populations that encounter resource depletion and changing environments, including activation of pro-survival mechanisms, migration to new locations, and escape of predation. We identified the p38 MAPK pathway as a common molecular driver of these three responses during the adaptation to hormone therapy resistance in prostate cancer. The p38 pathway is activated in therapy-resistant cells and mechanistically drives these three convergent responses through sustained AR activity, enhanced invasion and metastasis, and immune evasion. Targeting p38 signaling may represent a new therapeutic strategy to treat men with metastatic, hormone therapy-resistant prostate cancer.

Gromov-Wasserstein optimal transport to align single-cell multi-omics data

Demetci, P. — 2020-04-29

Data integration of single-cell measurements is critical for understanding cell development and disease, but the lack of correspondence between different types of measurements makes such efforts challenging. Several unsupervised algorithms can align heterogeneous single-cell measurements in a shared space, enabling the creation of mappings between single cells in different data domains. However, these algorithms require hyperparameter tuning for high-quality alignments, which is difficult in an unsupervised setting without correspondence information for validation. We present Single-Cell alignment using Optimal Transport (SCOT), an unsupervised learning algorithm that uses Gromov Wasserstein-based optimal transport to align single-cell multi-omics datasets. We compare the alignment performance of SCOT with state-of-the-art algorithms on four simulated and two real-world datasets. SCOT performs on par with state-of-the-art methods but is faster and requires tuning fewer hyperparameters. Furthermore, we provide an algorithm for SCOT to use Gromov Wasserstein distance to guide the parameter selection. Thus, unlike previous methods, SCOT aligns well without using any orthogonal correspondence information to pick the hyperparameters. Our source code and scripts for replicating the results are available at https://github.com/rsinghlab/SCOT.

Sniff-synchronized, gradient-guided olfactory search by freely-moving mice

Findley, T. M. — 2020-05-01

For many organisms, searching for relevant targets such as food or mates entails active, strategic sampling of the environment. Finding odorous targets may be the most ancient search problem that motile organisms evolved to solve. While chemosensory navigation has been well characterized in micro-organisms and invertebrates, spatial olfaction in vertebrates is poorly understood. We have established an olfactory search assay in which freely-moving mice navigate noisy concentration gradients of airborne odor. Mice solve this task using concentration gradient cues and do not require stereo olfaction for performance. During task performance, respiration and nose movement are synchronized with tens of milliseconds precision. This synchrony is present during trials and largely absent during inter-trial intervals, suggesting that sniff-synchronized nose movement is a strategic behavioral state rather than simply a constant accompaniment to fast breathing. To investigate the spatiotemporal structure of these active sensing movements, we used machine learning methods to parse motion trajectories into elementary movement motifs. Motifs fall into two clusters, which correspond to investigation and approach states. Investigation motifs lock precisely to sniffing, such that the individual motifs preferentially occur at specific phases of the sniff cycle. This work clarifies sensorimotor strategies for mouse olfactory search and guides ongoing work into the underlying neural mechanisms.

The Role of Conjunctive Representations in Controlling Actions

Kikumoto, A. — 2020-05-02

Action selection appears to rely on conjunctive representations that nonlinearly integrate task-relevant features (Kikumoto & Mayr, 2020). We test here the corollary hypothesis that such representations are also intricately involved during attempts to stop an action--a key aspect of action regulation. We tracked both conjunctive representations and those of constituent rule, stimulus, or response features through trial-by-trial representational similarity analysis of the EEG signal in a combined, rule-selection and stop-signal paradigm. Across two experiments with student participants (N = 57), we found (a) that the strength of decoded conjunctive representations prior to the stop signal uniquely predicted trial-by-trial stopping success (Exp. 1) and (b) that these representations were selectively suppressed following the onset of the stop signal (Exp. 1 and 2). We conclude that conjunctive representations are key to successful action execution and therefore need to be suppressed when an intended action is no longer appropriate. Statement of RelevanceSome theorists have posited that as a necessary step during action selection, action-relevant features need to be combined within a conjunctive representation that is more than the sum if its basic features. Consequently, such representations should also play a critical role when trying to stop an intended action--a key aspect of self-regulation. However direct evidence of conjunctive representations has been elusive. Using a method for tracking both conjunctive and basic-feature representations on a trial-by-trial basis in the EEG signal, we show that the stronger the conjunctive representations, the harder it was to stop the intended action. Furthermore, the stopping process also selectively reduced the strength of conjunctive representations. These results further our knowledge about action regulation by showing that conjunctive representations are a necessary precursor for carrying out actions successfully and for that reason also need to be the target of self-regulatory stopping attempts.

No Difference in 2D:4D Ratio between Youth with Elevated Prenatal Androgen Exposure due to Congenital Adrenal Hyperplasia and Controls

Nave, G. — 2020-05-08

The second-to-fourth digit ratio (2D:4D) has been associated with sexual dimorphism, with a lower 2D:4D in males. A large body of research has relied on the 2D:4D as a proxy for prenatal androgen exposure, and includes reports of relationships between 2D:4D and a wide range of human traits. Here, we examine the validity of the 2D:4D proxy by studying the association between 2D:4D and classical Congenital Adrenal Hyperplasia (CAH) due to 21-hydroxylase deficiency, a condition characterized by excessive prenatal exposure to androgens during most of the gestational period. To this end, we retrospectively examine 513 serial radiographs of the left hand obtained clinically in 90 youth with classical CAH (45 female) and 70 control youth (31 female). Replicating previous reports, we observe associations of the 2D:4D with sex (lower 2D:4D in males) and age (increase of 2D:4D through development). However, we find no evidence for differences in 2D:4D between CAH and controls (full sample: {square} = -0.001 (-0.008, 0.006)]; females: {square} = -0.004 [-0.015, 0.007]; males: {square} = 0.001, [-0.008, 0.011]). Although our findings do not rule out a small association between the 2D:4D and CAH, they cast doubt on the usefulness of the 2D:4D as a biomarker for prenatal androgen exposure in behavioral research.

Pulmonary infection interrupts acute cutaneous wound healing through disruption of chemokine signals

Crane, M. J. — 2020-05-10

Studies of the immune response typically focus on single-insult systems, with little known about how multi-insult encounters are managed. Pneumonia in patients recovering from surgery is a clinical situation that exemplifies the need for the patient to mount two distinct immune responses. Examining this, we have determined that poor wound healing is an unreported complication of pneumonia in laparotomy patients. Using mouse models, we found that lung infection suppressed the trafficking of innate leukocytes to wounded skin, while pulmonary resistance to the bacterial infection was maintained. The dual insults caused distinct systemic and local changes to the inflammatory response, the most striking being a rapid and sustained decrease in chemokine levels at the wound site of mice with pneumonia. Remarkably, replenishing wound chemokine levels completely rescued the wound-healing rate in mice with a pulmonary infection. These findings have broad implications for understanding the mechanisms guiding the innate immune system to prioritize inflammatory sites. One Sentence SummaryChemokine-mediated signaling drives the prioritization of innate immune responses to bacterial pulmonary infection over cutaneous wound healing. HighlightsO_LIHuman laparotomy patients with pneumonia have an increased rate of incision dehiscence, and this observation can be recapitulated in mouse models of bacterial lung infections and skin wounds. C_LIO_LILung infection causes rapid and sustained suppression of skin wound chemokine and inflammatory cytokine production as well as leukocyte recruitment. C_LIO_LIUnique systemic shifts in the immune compartment occur with two inflammatory insults, including the cytokine/chemokine signature and the mobilization, recruitment, and phenotype of innate leukocytes. C_LIO_LIRestoration of chemokine signaling in the wounds of mice that have a lung infection results in increased neutrophil trafficking to the wound site and rescues the rate of healing. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/084442v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@861000org.highwire.dtl.DTLVardef@1847251org.highwire.dtl.DTLVardef@6d6438org.highwire.dtl.DTLVardef@1ce99c6_HPS_FORMAT_FIGEXP M_FIG C_FIG

L-bodies are novel RNA-protein condensates driving RNA transport in Xenopus oocytes

Neil, C. R. — 2020-05-10

RNP granules are membrane-less compartments within cells, formed by phase separation, that function as regulatory hubs for diverse biological processes. However, the mechanisms by which RNAs and proteins interact to promote RNP granule structure and function in vivo remain unclear. In Xenopus laevis oocytes, maternal mRNAs are transported as large RNPs to the vegetal hemisphere of the developing oocyte, where local translation is critical for proper embryonic patterning. Here, we demonstrate that vegetal transport RNPs represent a new class of cytoplasmic RNP granule, termed Localization-bodies (L-bodies). We show that L-bodies are multiphase RNP granules, containing a dynamic protein-containing phase surrounding a non-dynamic RNA-containing substructure. Our results support a role for RNA as a critical scaffold component within these RNP granules and suggest that cis-elements within localized mRNAs may drive subcellular RNA localization through control over phase behavior.

Describing the current status of Plasmodium falciparum population structure and drug resistance within mainland Tanzania using molecular inversion probes

Moser, K. A. — 2020-05-09

High-throughput Plasmodium genomic data is increasingly useful in assessing prevalence of clinically important mutations and malaria transmission patterns. Understanding parasite diversity is important for identification of specific human or parasite populations that can be targeted by control programs, and to monitor the spread of mutations associated with drug resistance. An up-to-date understanding of regional parasite population dynamics is also critical to monitor the impact of control efforts. However, this data is largely absent from high-burden nations in Africa, and to date, no such analysis has been conducted for malaria parasites in Tanzania country-wide. To this end, over 1,000 P. falciparum clinical isolates were collected in 2017 from 13 sites in seven administrative regions across Tanzania, and parasites were genotyped at 1,800 variable positions genome-wide using molecular inversion probes. Population structure was detectable among Tanzanian P. falciparum parasites, roughly separating parasites from the northern and southern districts and identifying genetically admixed populations in the north. Isolates from geographically close districts were more likely to be genetically related compared to parasites sampled from more distant districts. Known drug resistance mutations were seen at increased frequency in northern districts, and additional variants with undetermined significance for antimalarial resistance also varied by geography. Malaria Indicator Survey (2017) data corresponded with genetic findings, including average region-level complexity-of-infection and malaria prevalence estimates. The parasite populations identified here provide important information on extant spatial patterns of genetic diversity of Tanzanian parasites, to which future surveys of genetic relatedness can be compared. SIGNIFICANCEDocumenting dynamics of malaria parasite genomics in high-transmission settings at scale in sub-Saharan Africa is critical for policy and decision making to support ongoing malaria elimination initiatives. Using molecular inversion probes, we genotyped over 1,000 Tanzanian Plasmodium falciparum samples collected country-wide in 2017 at hundreds of variable polymorphic positions across the genome. Frequencies of known drug resistance mutations were higher in northern districts of the country compared to the south. Results also showed a distinct isolation-by-distance pattern (whereby increasing geographic distance was correlated with decreasing genetic relatedness), as well as signals of higher genetic sharing between several southern districts. These results provide, for the first time, a picture of current within-country diversity of Tanzanian P. falciparum populations.

Fronto-parietal, cingulo-opercular and striatal contributions to learning and implementing control policies

Bhandari, A. — 2020-05-11

Efficient task performance requires co-ordination of internal cognitive processes by implementing control policies adapted to the dynamic structure of task demands. The cognitive and neural basis of control policy implementation remains poorly characterized, in part because it is typically confounded with implementing new stimulus-response rules. To disambiguate these processes, we asked participants to perform multiple novel variants of a working memory control task. Each variant had a unique, novel sequential trial structure, but all shared common stimulusresponse rules, enabling us to test control policy implementation separate from rule learning. Behaviorally, we found evidence for two adaptive processes tied to control policy implementation. One process was reflected in slower responses on the first trial with a novel sequential trial structure, followed by rapid speeding on subsequent trials. A second process was reflected in the diminishing size of the first trial cost as participants accommodated different variants of the task over many blocks. Using fMRI, we observed that the striatum and a cingulo-opercular cortical network increased activity to the first trial, tracking the fast adjustment. This pattern of activity dissociated these regions from a fronto-parietal network including dorsolateral PFC, inferior frontal junction, inferior parietal sulcus, and rostrolateral PFC, which showed a slower decline in activity across trials, mirroring findings in rule implementation studies, but in the absence of rule implementation demands. Our results reveal two adaptive processes underlying the implementation of efficient, generalizable control policies, and suggest a broader account of the role of a cortico-striatal network in control policy implementation. Significance statementRapid adaptation to novel tasks is a hallmark of human behavior. Understanding how human brains achieve this is of critical importance in neuroscience. Here we broaden the scope of this problem, going beyond task rules to more broadly consider the cognitive control demands produced by novel task dynamics. We propose that humans rely on two adaptive processes to rapidly implement efficient, generalizable control policies as task dynamics change, even when task rules remain unchanged. One process unfolds rapidly and underlies efficient adaptation. A second process unfolds slowly with experience across task conditions and underlies generalization of control policies. Using fMRI, we identify cingulo-opercular cortex, fronto-parietal cortex and striatum as dissociable components of a cortico-striatal network that contribute to control implementation.

When effort matters: Expectations of reward and efficacy guide cognitive control allocation

Froemer, R. — 2020-05-16

The amount of mental effort we invest in a task is influenced by the reward we can expect if we perform that task well. However, some of the rewards that have the greatest potential for driving these efforts (e.g., jobs, grants) are partly determined by factors beyond ones control. In such cases, effort has more limited efficacy for obtaining rewards. We have proposed that people integrate information about the expected reward and efficacy for effort to determine the expected value of control, and then adjust their control allocation (i.e. mental effort) accordingly. Here we test this theorys key behavioral and neural predictions. We show that participants invest more control when this control is more rewarding and more efficacious, and that these incentive components separately modulate EEG signatures of incentive evaluation and proactive control allocation. Our findings support the prediction that people combine worth and worthwhileness to determine how much effort to invest.

Membrane bending by protein phase separation

Yuan, F. — 2020-05-22

Membrane bending is a ubiquitous cellular process that is required for membrane traffic, cell motility, organelle biogenesis, and cell division. Proteins that bind to membranes using specific structural features, such as wedge-like amphipathic helices and crescent-shaped scaffolds, are thought to be the primary drivers of membrane bending. However, many membrane-binding proteins have substantial regions of intrinsic disorder, which lack a stable three-dimensional structure. Interestingly, many of these disordered domains have recently been found to form networks stabilized by weak, multi-valent contacts, leading to assembly of protein liquid phases on membrane surfaces. Here we ask how membrane-associated protein liquids impact membrane curvature. We find that protein phase separation on the surfaces of synthetic and cell-derived membrane vesicles creates a substantial compressive stress in the plane of the membrane. This stress drives the membrane to bend inward, creating protein-lined membrane tubules. A simple mechanical model of this process accurately predicts the experimentally measured relationship between the rigidity of the membrane and the diameter of the membrane tubules. Discovery of this mechanism, which may be relevant to a broad range of cellular protrusions, illustrates that membrane remodeling is not exclusive to structured scaffolds, but can also be driven by the rapidly emerging class of liquid-like protein networks that assemble at membranes. Significance StatementCellular membranes take on an elaborate set of highly curved and bent shapes, which are essential to diverse cellular functions from endocytosis to cell division. The prevailing view has been that membrane bending is driven by proteins with curved shapes, which assemble at the membrane surface to form solid scaffolds. In contrast, here we show that proteins which form liquid-like assemblies on membranes are also potent drivers of bending. These "liquid scaffolds" apply compressive stress to the membrane surface, generating a diverse and dynamic family of membrane shapes. These data, which come at a time when liquid-like protein assemblies are being identified throughout the cell, suggest that liquid-like protein assemblies may play an important role in shaping cellular membranes.

Natural variation in the regulation of neurodevelopmental genes modifies flight performance in Drosophila

Spierer, A. — 2020-05-27

The winged insects of the order Diptera are colloquially named for their most recognizable phenotype: flight. These insects rely on flight for a number of important life history traits, like dispersal, foraging, and courtship. Despite the importance of flight, relatively little is known about the genetic architecture of variation for flight performance. Accordingly, we sought to uncover the genetic modifiers of flight using a measure of flies reaction and response to an abrupt drop in a vertical flight column. We conducted an association study using 197 of the Drosophila Genetic Reference Panel (DGRP) lines, and identified a combination of additive and marginal variants, epistatic interactions, whole genes, and enrichment across interaction networks. We functionally validated 13 of these candidate genes (Adgf-A/Adgf-A2/CG32181, bru1, CadN, CG11073, CG15236, CG9766, CREG, Dscam4, form3, fry, Lasp/CG9692, Pde6, Snoo) contribution to flight, two of which (fry and Snoo) also validate a whole gene analysis we introduce for the DGRP: PEGASUS_flies. Overall, our results suggest modifiers of muscle and wing morphology, and peripheral and central nervous system assembly and function are all important for flight performance. Additionally, we identified ppk23, an Acid Sensing Ion Channel (ASIC) homolog, as an important hub for epistatic interactions. These results represent a snapshot of the genetic modifiers affecting drop-response flight performance in Drosophila, with implications for other insects. It also draws connections between genetic modifiers of performance and BMP signaling and ASICs as targets for treating neurodegeneration and neurodysfunction. Author summaryInsect flight is a widely recognizable phenotype of winged insects, hence the name: flies. While fruit flies, or Drosophila melanogaster, are a genetically tractable model, flight performance is a highly integrative phenotype, making it challenging to comprehensively identify the genetic modifiers that contribute to its genetic architecture. Accordingly, we screened 197 Drosophila Genetic Reference Panel lines for their ability to react and respond to an abrupt drop. Using several computational tools, we successfully identified several additive, marginal, and epistatic variants, as well as whole genes and altered sub-networks of gene-gene and protein-protein interaction networks, demonstrating the benefits of using multiple methodologies to elucidate the genetic architecture of complex traits more generally. Many of these significant genes and variants mapped to regions of the genome that affect development of sensory and motor neurons, wing and muscle development, and regulation of transcription factors. We also introduce PEGASUS_flies, a Drosophila-adapted version of the PEGASUS platform first used in human studies, to infer gene-level significance of association based on the distribution of individual variant P-values. Our results contribute to the debate over the relative importance of individual, additive factors and epistatic, or higher order, interactions, in the mapping of genotype to phenotype.

The role of matrix metalloproteinase-9 (MMP-9) in neurodevelopmental deficits and experience-dependent structural plasticity in Xenopus laevis tadpoles

Gore, S. — 2020-05-31

Matrix metalloproteinase-9 (MMP-9) is a secreted endopeptidase targeting extracellular matrix proteins, creating permissive environments for neuronal development and plasticity. Developmental dysregulation of MMP-9 is also associated with neurodevelopmental disorders (ND). Here we test the hypothesis that chronically elevated MMP-9 activity during early neurodevelopment is responsible for neural circuit hyperconnectivity observed after early exposure to valproic acid (VPA), a known teratogen associated with autism spectrum disorder in humans. In Xenopus tadpoles, VPA exposure results in excess local synaptic connectivity, disrupted social behavior and increased seizure susceptibility. We found that overexpressing MMP-9 in the brain copies effects of VPA on synaptic connectivity, and blocking MMP-9 activity pharmacologically or genetically reverses effects of VPA on physiology and behavior. We further show that during normal neurodevelopment MMP-9 levels are tightly regulated by neuronal activity and required for structural plasticity. These studies show a critical role for MMP-9 in both normal and abnormal development.

Lynx1 modulates the activity of nAChRs to slow NMJ and muscle fiber degeneration during aging

Vaughan, S. K. — 2020-05-31

The authors have withdrawn their manuscript to revisit some of the data, interpretations and conclusions. Therefore, the authors do not wish this work to be cited as a reference. If you have any questions, please contact the corresponding author.

Spatially displaced excitation contributes to the encoding of interrupted motion by the retinal direction-selective circuit

Ding, J. — 2020-05-31

Spatially distributed excitation and inhibition collectively shape a visual neurons receptive field (RF) properties. In the direction-selective circuit of the mammalian retina, the role of strong null-direction inhibition of On-Off direction-selective ganglion cells (ON-OFF DSGCs) on their direction selectivity is well-studied. However, how excitatory inputs influence the On-Off DSGCs visual response is underexplored. Here, we report that On-Off DSGCs have a spatially displaced glutamatergic receptive field along their preferred-null motion axis. This displaced receptive field contributes to DSGC null-direction spiking during interrupted motion trajectories. Theoretical analyses indicate that population responses during interrupted motion may help populations of On-Off DSGCs signal the spatial location of moving objects in complex, naturalistic visual environments. Our study highlights that the direction-selective circuit exploits separate sets of mechanisms under different stimulus conditions, and these mechanisms may help encode multiple visual features.

Validation and Performance Comparison of Three SARS-CoV-2 Antibody Assays

Lu, S. — 2020-05-30

Serology testing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is increasingly being used during the current pandemic of Coronavirus Disease 2019 (COVID-19). The clinical and epidemiologic utilities of antibody-based SARS-CoV-2 testing are under debate. Characterizing these assays helps to understand the disease and provides scientific basis for deciding how to best use these assays. The study assessed one chemiluminescent assay (Abbott COVID-2 IgG) and two lateral flow assays (STANDARD Q [SQ] IgM/IgG Duo and Wondfo Total Antibody Test). Validation included 113 blood samples from 71 PCR-confirmed COVID-19 patients and 1182 samples from negative controls with potential interferences/cross-reactions, including 1063 pre-pandemic samples. IgM antibodies against SARS-CoV-2 were detected as early as post-symptom onset days 3-4. IgG antibodies were first detected post-onset days 5-6 by SQ assays. The detection rates increased gradually, and SQ IgG, Abbott IgG and Wondfo Total detected antibodies from all the PCR-confirmed patients 14 days after symptom onset. Overall agreements between SQ IgM/IgG and Wondfo Total was 88.5% and between SQ IgG and Abbott IgG was 94.6% (Kappa = 0.75, 0.89). No cross-reaction with other endemic coronavirus infections were identified. Viral hepatitis and autoimmune samples were the main cross-reactions observed. However, the interferences/cross-reactions were low. The specificities were 100% for SQ IgG and Wondfo Total and 99.62% for Abbott IgG and 98.87% for SQ IgM. These findings demonstrate high sensitivity and specificity of appropriately validated antibody-based SARS-CoV-2 assays with implications for clinical use and epidemiological seroprevalence studies.View Full Text

The neural representation of force across grasp types in motor cortex of humans with tetraplegia

Rastogi, A. — 2020-06-02

Intracortical brain-computer interfaces (iBCIs) have the potential to restore hand grasping and object interaction to individuals with tetraplegia. Optimal grasping and object interaction require simultaneous production of both force and grasp outputs. However, since overlapping neural populations are modulated by both parameters, grasp type could affect how well forces are decoded from motor cortex in a closed-loop force iBCI. Therefore, this work quantified the neural representation and offline decoding performance of discrete hand grasps and force levels in two participants with tetraplegia. Participants attempted to produce three discrete forces (light, medium, hard) using up to five hand grasp configurations. A two-way Welch ANOVA was implemented on multiunit neural features to assess their modulation to force and grasp. Demixed principal component analysis was used to assess for population-level tuning to force and grasp and to predict these parameters from neural activity. Three major findings emerged from this work: 1) Force information was neurally represented and could be decoded across multiple hand grasps (and, in one participant, across attempted elbow extension as well); 2) Grasp type affected force representation within multi-unit neural features and offline force classification accuracy; and 3) Grasp was classified more accurately and had greater population-level representation than force. These findings suggest that force and grasp have both independent and interacting representations within cortex, and that incorporating force control into real-time iBCI systems is feasible across multiple hand grasps if the decoder also accounts for grasp type. Significance StatementIntracortical brain-computer interfaces (iBCIs) have emerged as a promising technology to potentially restore hand grasping and object interaction in people with tetraplegia. This study is among the first to quantify the degree to which hand grasp affects force-related - or kinetic - neural activity and decoding performance in individuals with tetraplegia. The study results enhance our overall understanding of how the brain encodes kinetic parameters across varying kinematic behaviors -- and in particular, the degree to which these parameters have independent versus interacting neural representations. Such investigations are a critical first step to incorporating force control into human-operated iBCI systems, which would move the technology towards restoring more functional and naturalistic tasks.

Instantaneous amplitude and shape of postrhinal theta oscillations differentially encode running speed

Ghosh, M. — 2020-06-04

Hippocampal theta oscillations have a temporally asymmetric waveform shape, but it is not known if this theta asymmetry extends to all other cortical regions involved in spatial navigation and memory. Here, using both established and improved cycle-by-cycle analysis methods, we show that theta waveforms in the postrhinal cortex are also temporally asymmetric. On average, the falling phase of postrhinal theta cycles lasts longer than the subsequent rising phase. There are, however, rapid changes in both the instantaneous amplitude and instantaneous temporal asymmetry of postrhinal theta cycles. These rapid changes in amplitude and asymmetry are very poorly correlated, indicative of a mechanistic disconnect between these theta cycle features. We show that the instantaneous amplitude and asymmetry of postrhinal theta cycles differentially encode running speed. Although theta amplitude continues to increase at the fastest running speeds, temporal asymmetry of the theta waveform shape plateaus after medium speeds. Our results suggest that the amplitude and waveform shape of individual postrhinal theta cycles may be governed by partially independent mechanisms and emphasize the importance of employing a single cycle approach to understanding the genesis and behavioral correlates of cortical theta rhythms.

An expanding bacterial colony forms a depletion zone with growing droplets

Tang, J. X. — 2020-06-03

Many species of bacteria have developed means to spread on solid surfaces. This study focuses on the expansion of Pseudomonas aeruginosa on an agar gel surface. We report the occurrence and spread of a depletion zone, where the layer of bacteria on the agar becoming thinner. The depletion zone occurs within an expanded colony under conditions of minimal water evaporation. It is colocalized with a higher concentration of rhamnolipids, the biosurfactants that are produced by the bacteria and accumulate in the older region of the colony. With continued growth in bacterial population, dense droplets occur and coalesce in the depletion zone, displaying remarkable fluid dynamic behavior. Whereas expansion of a central depletion zone requires activities of live bacteria, new zones can be seeded by adding rhamnolipids. These depletion zones due to the added surfactants expand quickly, even on plates covered by bacteria that have been killed by ultraviolet light. We propose a model to account for the observed properties, taking into consideration bacterial growth and secretion, osmotic swelling, fluid volume expansion, cell-cell interaction, and interfacial fluid dynamics involving Marangoni flow. SignificanceBacterial growth and pattern formation have strong bearing on their biological functions, such as their spread and accumulation, biofilm growth & its effects on infection and antibiotic resistance. The bacterial species of this study, Pseudomonas aeruginosa, is a human pathogen responsible for frequent infections in wounds, airways, and urinary tract, particularly when involving the use of catheters. The findings of this study and the mechanisms we propose offer new insights on the important behaviors of bacterial collective motility, pattern dynamics, and biofilm growth.

Latent motives guide structure learning during adaptive social choice

van Baar, J. M. — 2020-06-08

Predicting the behavior of others is an essential part of human cognition that enables strategic social behavior (e.g., cooperation), and is impaired in multiple clinical populations. Despite its ubiquity, social prediction poses a generalization problem that remains poorly understood: We can neither assume that others will simply repeat their past behavior in new settings, nor that their future actions are entirely unrelated to the past. Here we demonstrate that humans solve this challenge using a structure learning mechanism that uncovers other people’s latent, unobservable motives, such as greed and risk aversion. In three studies, participants were tasked with predicting the decisions of another player in multiple unique economic games such as the Prisoner’s Dilemma. Participants achieved accurate social prediction by learning the hidden motivational structure underlying the player’s actions to cooperate or defect (e.g., that greed led to defecting in some cases but cooperation in others). This motive-based abstraction enabled participants to attend to information diagnostic of the player’s next move and disregard irrelevant contextual cues. Moreover, participants who successfully learned another’s motives were more strategic in a subsequent competitive interaction with that player, reflecting that accurate social structure learning can lead to more optimal social behaviors. These findings demonstrate that advantageous social behavior hinges on parsimonious and generalizable mental models that leverage others’ latent intentions.Significance statement A hallmark of human cognition is being able to predict the behavior of others. How do we achieve social prediction given that we routinely encounter others in a dizzying array of social situations? We find people achieve accurate social prediction by inferring another’s hidden motives—motives that do not necessarily have a one-to-one correspondence with observable behaviors. Participants were able to infer another’s motives using a structure learning mechanism that enabled generalization. Individuals used what they learned about others in one setting to predict their actions in an entirely new setting. This cognitive process can explain a wealth of social behaviors, ranging from strategic economic decisions to stereotyping and racial bias.Competing Interest StatementThe authors have declared no competing interest.View Full Text

Striatal dopamine synthesis capacity reflects smartphone social activity

Westbrook, A. — 2020-06-08

Striatal dopamine has been implicated in social behavior across humans, rodents, and non-human primates in artificial laboratory settings with highly-practiced tasks and fixed reward contingencies. Whether striatal dopamine drives naturalistic, spontaneous social behavior remains unclear. Here, we leverage day-to-day logs of unconstrained smartphone behavior and establish a novel link between smartphone social activity and individual differences in striatal dopamine synthesis capacity using [18F]-DOPA PET in (N=22) healthy adult humans. We find a strong relationship such that a higher proportion of social app interactions correlates with lower dopamine synthesis capacity in the bi-lateral putamen. Permutation tests and penalized regressions provide evidence that this link between dopamine synthesis capacity and social versus non-social smartphone taps is specific. These observations provide a key empirical grounding for current speculations about dopamines role in digital social behavior.

Neural systems underlying the learning of cognitive effort costs

Sayali, C. — 2020-06-08

People balance the benefits of cognitive work against the costs of cognitive effort. Models that incorporate prospective estimates of the costs of cognitive effort into decision making require a mechanism by which these costs are learned. However, it remains open what brain systems are important for this learning, particularly when learning is not tied explicitly to a decision about what task to perform. In this fMRI experiment, we parametrically manipulated the level of effort a task requires by increasing task switching frequency across six task contexts. In a scanned learning phase, participants implicitly learned about the task switching frequency in each context. In a subsequent test phase, participants made selections between pairs of these task contexts. We modeled learning within a reinforcement learning framework, and found that effort expectations that derived from task-switching probability and response time (RT) during learning were the best predictors of later choice behavior. Prediction errors (PE) from these two models were associated with FPN during distinct learning epochs. Specifically, PE derived from expected RT was most correlated with the fronto-parietal network early in learning, whereas PE derived from expected task switching frequency was correlated with the fronto-parietal network late in learning. These results suggest that multiple task-related factors are tracked by the brain while performing a task that can drive subsequent estimates of effort costs.

The epidemic consequences of virulence and free-living survival relationships in models of emerging outbreaks

Gomez, L. M. — 2020-06-09

The relationship between parasite virulence and transmission is a pillar of evolutionary theory that has specific implications for public health. Part of this canon involves the idea that virulence and free-living survival (a key component of transmission) may have different relationships in different host-parasite systems. Most examinations of the evolution of virulence-transmission relationships--theoretical or empirical in nature--tend to focus on the evolution of virulence, with transmission a secondary consideration. And even within transmission studies, the focus on free-living survival is a smaller subset, though recent studies have examined its importance in the ecology of infectious diseases. Few studies have examined the epidemic-scale consequences of variation in survival across different virulence-survival relationships. In this study, we utilize a mathematical model motivated by aspects of SARS-CoV-2 natural history to investigate how evolutionary changes in survival may influence several aspects of disease dynamics at the epidemiological scale. Across virulence-survival relationships (where these traits are positively or negatively correlated), we found that small changes (5% above and below the nominal value) in survival can have a meaningful effect on certain outbreak features, including the R0, and the size of the infectious peak in the population. These results highlight the importance of properly understanding the mechanistic relationship between virulence and parasite survival, as evolution of increased survival across different relationships with virulence will have considerably different epidemiological signatures.

SARS-CoV-2 nucleocapsid protein undergoes liquid-liquid phase separation stimulated by RNA and partitions into phases of human ribonucleoproteins

Perdikari, T. M. — 2020-06-10

Tightly packed complexes of nucleocapsid protein and genomic RNA form the core of viruses and may assemble within viral factories, dynamic compartments formed within the host cells. Here, we examine the possibility that the multivalent RNA-binding nucleocapsid protein (N) from the severe acute respiratory syndrome coronavirus (SARS-CoV-2) compacts RNA via protein-RNA liquid-liquid phase separation (LLPS) and that N interactions with host RNA-binding proteins are mediated by phase separation. To this end, we created a construct expressing recombinant N fused to a N-terminal maltose binding protein tag which helps keep the oligomeric N soluble for purification. Using in vitro phase separation assays, we find that N is assembly-prone and phase separates avidly. Phase separation is modulated by addition of RNA and changes in pH and is disfavored at high concentrations of salt. Furthermore, N enters into in vitro phase separated condensates of full-length human hnRNPs (TDP-43, FUS, and hnRNPA2) and their low complexity domains (LCs). However, N partitioning into the LC of FUS, but not TDP-43 or hnRNPA2, requires cleavage of the solubilizing MBP fusion. Hence, LLPS may be an essential mechanism used for SARS-CoV-2 and other RNA viral genome packing and host protein co-opting, functions necessary for viral replication and hence infectivity.

A predictive coarse-grained model for position-specific effects of post-translational modifications on disordered protein phase separation

Perdikari, T. M. — 2020-06-12

Biomolecules undergo liquid-liquid phase separation (LLPS) resulting in the formation of multicomponent protein-RNA membraneless organelles in cells. However, the physiological and pathological role of post translational modifications (PTMs) on the biophysics of phase behavior is only beginning to be probed. To study the effect of PTMs on LLPS in silico, we extend our transferable coarse-grained model of intrinsically disordered proteins to include phosphorylated and acetylated amino acids. Using the parameters for modified amino acids available for fixed charge atomistic forcefields, we parameterize the size and atomistic hydropathy of the coarse-grained modified amino acid beads, and hence the interactions between the modified and natural amino acids. We then elucidate how the number and position of phosphorylated and acetylated residues alter the proteins single chain compactness and its propensity to phase separate. We show that both the number and the position of phosphorylated threonines/serines or acetylated lysines can serve as a molecular on/off switch for phase separation in the well-studied disordered regions of FUS and DDX3X, respectively. We also compare modified residues to their commonly used PTM mimics for their impact on chain properties. Importantly, we show that the model can predict and capture experimentally measured differences in the phase behavior for position-specific modifications, showing that the position of modifications can dictate phase separation. In sum, this model will be useful for studying LLPS of post-translationally modified intrinsically disordered proteins and predicting how modifications control phase behavior with position-specific resolution. Statement of SignificancePost-translational modifications are important regulators of liquid-liquid phase separation (LLPS) which drives the formation of biomolecular condensates. Theoretical methods can be used to characterize the biophysical properties of intrinsically disordered proteins (IDPs). Our recent framework for molecular simulations using a C-centered coarse-grained model can predict the effect of various perturbations such as mutations (Dignon et al. PloS Comput. Biol, 2018) and temperature (Dignon et al, ACS Cent. Sci., 2019) on LLPS. Here, we expand this framework to incorporate modified residues like phosphothreonine, phosphoserine and acetylysine. This model will prove useful for simulating the phase separation of post-translationally modified IDPs and predicting how position-specific modifications can control phase behavior across the large family of proteins known to be phosphorylated and acetylated.

Unsupervised manifold alignment for single-cell multi-omics data

Singh, R. — 2020-06-15

Integrating single-cell measurements that capture different properties of the genome is vital to extending our understanding of genome biology. This task is challenging due to the lack of a shared axis across datasets obtained from different types of single-cell experiments. For most such datasets, we lack corresponding information among the cells (samples) and the measurements (features). In this scenario, unsupervised algorithms that are capable of aligning single-cell experiments are critical to learning an in silico co-assay that can help draw correspondences among the cells. Maximum mean discrepancy-based manifold alignment (MMD-MA) is such an unsupervised algorithm. Without requiring correspondence information, it can align single-cell datasets from different modalities in a common shared latent space, showing promising results on simulations and a small-scale single-cell experiment with 61 cells. However, it is essential to explore the applicability of this method to larger single-cell experiments with thousands of cells so that it can be of practical interest to the community. In this paper, we apply MMD-MA to two recent datasets that measure transcriptome and chromatin accessibility in ~2000 single cells. To scale the runtime of MMD-MA to a more substantial number of cells, we extend the original implementation to run on GPUs. We also introduce a method to automatically select one of the user-defined parameters, thus reducing the hyperparameter search space. We demonstrate that the proposed extensions allow MMD-MA to accurately align state-of-the-art single-cell experiments.

Miniaturized Devices for Bioluminescence Imaging in Freely Behaving Animals

Celinskis, D. — 2020-06-16

Fluorescence miniature microscopy in vivo has recently proven a major advance, enabling cellular imaging in freely behaving animals. However, fluorescence imaging suffers from autofluorescence, phototoxicity, photobleaching and non-homogeneous illumination artifacts. These factors limit the quality and time course of data collection. Bioluminescence provides an alternative kind of activity-dependent light indicator. Bioluminescent calcium indicators do not require light input, instead generating photons through chemiluminescence. As such, limitations inherent to the requirement for light presentation are eliminated. Further, bioluminescent indicators also do not require excitation light optics: the removal of this component should make lighter and lower cost microscope with fewer assembly parts. While there has been significant recent progress in making brighter and faster bioluminescence indicators, parallel advances in imaging hardware have not yet been realized. A hardware challenge is that despite potentially higher signal-to-noise of bioluminescence, the signal strength is lower than that of fluorescence. An open question we address in this report is whether fluorescent miniature microscopes can be rendered sensitive enough to detect bioluminescence. We demonstrate this possibility in vitro and in vivo by implementing optimizations of the UCLA fluorescent miniscope. These optimizations yielded a miniscope (BLmini) which is 22% lighter in weight, has 45% fewer components, is up to 58% less expensive, offers up to 15 times stronger signal (as dichroic filtering is not required) and is sensitive enough to capture spatiotemporal dynamics of bioluminescence in the brain with a signal-to-noise ratio of 34 dB.

Stick Stippling for Joint 3D Visualization of Diffusion MRI Fiber Orientations and Density

Cabeen, R. P. — 2020-06-16

This paper investigates a stick stippling approach for glyph-based visualization of complex neural fiber architecture derived from diffusion magnetic resonance imaging. The presence of subvoxel crossing fibers in the brain has prompted the development of advanced modeling techniques; however, there remains a need for improved visualization techniques to more clearly convey their rich structure. While tractography can illustrate large scale anatomy, visualization of diffusion models can provide a more complete picture of local anatomy without the known limitations of tracking. We identify challenges and evaluate techniques for visualizing multi-fiber models and identified techniques that improve on existing methods. We conducted experiments to compare these representations and evaluated them with in vivo diffusion MR datasets that vary in voxel resolution and anisotropy. We found that stick rendering as 3D tubes increased legibility of fiber orientation and that encoding fiber density by tube radius reduced clutter and reduced dependence on viewing orientation. Furthermore, we identified techniques to reduce the negative perceptual effects of voxel gridding through a jittering and re-sampling approach to produce a stippling effect. Looking forward, this approach provides a new way to explore diffusion MRI datasets that may aid in the visual analysis of white matter fiber architecture and microstructure. Our software implementation is available in the Quantitative Imaging Toolkit (QIT).

A system for phenotype harmonization in the NHLBI Trans-Omics for Precision Medicine (TOPMed) Program

Stilp, A. M. — 2020-06-20

Genotype-phenotype association studies often combine phenotype data from multiple studies to increase power. Harmonization of the data usually requires substantial effort due to heterogeneity in phenotype definitions, study design, data collection procedures, and data set organization. Here we describe a centralized system for phenotype harmonization that includes input from phenotype domain and study experts, quality control, documentation, reproducible results, and data sharing mechanisms. This system was developed for the National Heart, Lung and Blood Institutes Trans-Omics for Precision Medicine (TOPMed) program, which is generating genomic and other omics data for >80 studies with extensive phenotype data. To date, 63 phenotypes have been harmonized across thousands of participants from up to 17 TOPMed studies per phenotype. We discuss the challenges faced in this undertaking and how they were addressed. The harmonized phenotype data and associated documentation have been submitted to National Institutes of Health data repositories for controlled-access by the scientific community. We also provide materials to facilitate future harmonization efforts by the community, which include (1) the code used to generate the 63 harmonized phenotypes, enabling others to reproduce, modify or extend these harmonizations to additional studies; and (2) results of labeling thousands of phenotype variables with controlled vocabulary terms.

Mapping Drosophila insulin receptor structure to the regulation of aging through analysis of amino acid substitutions

Yamamoto, R. — 2020-07-01

Genetic manipulations of the Drosophila insulin/IGF signaling system slow aging, but it remains unknown how the insulin/IGF receptor acts to modulate lifespan or differentiate this control from that of growth, reproduction and metabolism. With homologous recombination we produced an allelic series of single amino acid substitutions in the fly insulin receptor (InR). Based on emerging biochemical and structural data, we map amino acid substitutions to receptor function to longevity and fecundity. We propose InR mutants generate bias in the process of asymmetric transphosphorylation when the receptor is activated. This induces specific kinase subdomains that modulate lifespan by additive processes, one involving survival costs of reproduction and the other involving reproduction-independent systems of longevity assurance. We identify a mutant in the kinase insert domain that robustly extends lifespan without affecting growth or reproduction, suggesting this element controls aging through unique mechanisms of longevity assurance.

Chronic activation of fear engrams induces extinction-like behavior in ethanol-exposed mice

Cincotta, C. — 2020-07-02

Alcohol withdrawal directly impacts the brains stress and memory systems, which may underlie individual susceptibility to persistent drug and alcohol-seeking behaviors. Numerous studies demonstrate that forced alcohol abstinence, which may lead to withdrawal, can impair fear-related memory processes in rodents such as extinction learning, however the underlying neural circuits mediating these impairments remain elusive. Here, we tested an optogenetic strategy aimed at mitigating fear extinction impairments in male c57BL/6 mice following exposure to alcohol (i.e., ethanol) and forced abstinence. In the first experiment, extensive behavioral extinction training in a fear-conditioned context was impaired in ethanol-exposed mice compared to controls. In the second experiment, neuronal ensembles processing a contextual fear memory in the dorsal hippocampus were tagged and optogenetically reactivated repeatedly in a distinct context in ethanol-exposed and control mice. Chronic activation of these cells resulted in a context-specific, extinction-like reduction in fear responses in both control and ethanol-exposed mice. These findings suggest that while ethanol can impair fear extinction learning, optogenetic manipulation of a fear engram is sufficient to induce an extinction-like reduction in fear responses.

Multi-scale Genomic Inference using Biologically Annotated Neural Networks

Demetci, P. — 2020-07-03

In this article, we present Biologically Annotated Neural Networks (BANNs), a nonlinear probabilistic framework for association mapping in genome-wide association (GWA) studies. BANNs are feedforward models with partially connected architectures that are based on biological annotations. This setup yields a fully interpretable neural network where the input layer encodes SNP-level effects, and the hidden layer models the aggregated effects among SNP-sets. We treat the weights and connections of the network as random variables with prior distributions that reflect how genetic effects manifest at different genomic scales. The BANNs software uses variational inference to provide posterior summaries which allow researchers to simultaneously perform (i) mapping with SNPs and (ii) enrichment analyses with SNP-sets on complex traits. Through simulations, we show that our method improves upon state-of-the-art association mapping and enrichment approaches across a wide range of genetic architectures. We then further illustrate the benefits of BANNs by analyzing real GWA data assayed in approximately 2,000 heterogenous stock of mice from the Wellcome Trust Centre for Human Genetics and approximately 7,000 individuals from the Framingham Heart Study. Lastly, using a random subset of individuals of European ancestry from the UK Biobank, we show that BANNs is able to replicate known associations in high and low-density lipoprotein cholesterol content. Author SummaryA common goal in genome-wide association (GWA) studies is to characterize the relationship between genotypic and phenotypic variation. Linear models are widely used tools in GWA analyses, in part, because they provide significance measures which detail how individual single nucleotide polymorphisms (SNPs) are statistically associated with a trait or disease of interest. However, traditional linear regression largely ignores non-additive genetic variation, and the univariate SNP-level mapping approach has been shown to be underpowered and challenging to interpret for certain trait architectures. While nonlinear methods such as neural networks are well known to account for complex data structures, these same algorithms have also been criticized as "black box" since they do not naturally carry out statistical hypothesis testing like classic linear models. This limitation has prevented nonlinear regression approaches from being used for association mapping tasks in GWA applications. Here, we present Biologically Annotated Neural Networks (BANNs): a flexible class of feedforward models with partially connected architectures that are based on biological annotations. The BANN framework uses approximate Bayesian inference to provide interpretable probabilistic summaries which can be used for simultaneous (i) mapping with SNPs and (ii) enrichment analyses with SNP-sets (e.g., genes or signaling pathways). We illustrate the benefits of our method over state-of-the-art approaches using extensive simulations. We also demonstrate the ability of BANNs to recover novel and previously discovered genomic associations using quantitative traits from the Wellcome Trust Centre for Human Genetics, the Framingham Heart Study, and the UK Biobank.

FreeClimber: Automated quantification of climbing performance in Drosophila, with examples from mitonuclear genotypes

Spierer, A. — 2020-07-04

Negative geotaxis (climbing) performance is a useful metric for quantifying Drosophila health and vigor. Manual methods to quantify climbing performance are slow, tedious, and may be systematically biased, while available computational methods have inflexible hardware or software requirements. We present an alternative: FreeClimber. This open source, Python-based pipeline subtracts a video’s static background to improve spot detection for moving flies in heterogeneous backgrounds. FreeClimber calculates a cohort’s velocity as the slope of the most linear portion of a mean-vertical position vs. time plot. It can run from a graphical user interface for parameter optimization or a command line interface for high-throughput and automated batch processing. It outputs calculated slopes, spot locations for follow up analyses such as tracking, and several visualizations and diagnostic plots. We demonstrate FreeClimber’s utility in a longitudinal study for endurance exercise performance in Drosophila using six distinct mitochondrial haplotypes paired with a common w1118 nuclear background.Summary statement FreeClimber quantifies the climbing velocity for a group of flies, eliminating systematic biases associated with traditional manual methods in a high throughput and automated (graphical and/or command line-based) platform.List of Symbols and AbbreviationsecceccentricityGUIGraphical User Interfacemito-nuclearMitochondrial-nuclearmitotypesMitochondrial haplotypeOreROregon RROIRegion of InterestyakyakubaZimZimbabwe53View Full Text

Unique transcriptional changes in coagulation cascade genes in SARS-CoV-2-infected lung epithelial cells: A potential factor in COVID-19 coagulopathies

FitzGerald, E. S. — 2020-07-07

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly become a global pandemic. In addition to the acute pulmonary symptoms of COVID-19 (the disease associated with SARS-CoV-2 infection), pulmonary and distal coagulopathies have caused morbidity and mortality in many patients. Currently, the molecular pathogenesis underlying COVID-19 associated coagulopathies are unknown. While there are many theories for the cause of this pathology, including hyper inflammation and excess tissue damage, the cellular and molecular underpinnings are not yet clear. By analyzing transcriptomic data sets from experimental and clinical research teams, we determined that changes in the gene expression of genes important in the extrinsic coagulation cascade in the lung epithelium may be important triggers for COVID-19 coagulopathy. This regulation of the extrinsic blood coagulation cascade is not seen with influenza A virus (IAV)-infected NHBEs suggesting that the lung epithelial derived coagulopathies are specific to SARS-Cov-2 infection. This study is the first to identify potential lung epithelial cell derived factors contributing to COVID-19 associated coagulopathy. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/182972v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@93cfb7org.highwire.dtl.DTLVardef@2a23c9org.highwire.dtl.DTLVardef@93623borg.highwire.dtl.DTLVardef@161e25_HPS_FORMAT_FIGEXP M_FIG C_FIG AUTHOR SUMMARYO_ST_ABSWhy was this study done?C_ST_ABSO_LISevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly become a global pandemic. C_LIO_LIIn addition to the acute pulmonary symptoms of COVID-19 (the disease associated with SARS-CoV-2 infection), pulmonary and distal coagulopathies have caused morbidity and mortality in many patients. C_LIO_LICurrently, the molecular pathogenesis underlying COVID-19 associated coagulopathies are unknown. Understanding the molecular basis of dysregulated blood coagulation during SARS-CoV-2 infection may help promote new therapeutic strategies to mitigate these complications in COVID-19 patients. C_LI What did the researchers do and find?O_LIWe analyzed three publicly available RNA sequencing datasets to identify possible molecular etiologies of COVID-19 associated coagulopathies. These data sets include sequencing libraries from clinically isolated samples of bronchoalveolar lavage fluid (BALF) and peripheral blood mononuclear cells (PBMCs) from SARS-CoV-2 positive patients and healthy controls. We also analyzed a publicly available RNA sequencing dataset derived from in vitro SARS-CoV-2 infected primary normal human bronchial epithelial (NHBE) cells and mock infected samples. C_LIO_LIPathway analysis of both NHBE and BALF differential gene expression gene sets. We found that SARS-CoV-2 infection induces the activation of the extrinsic blood coagulation cascade and suppression of the plasminogen activation system in both NHBEs and cells isolated from the BALF. PBMCs did not differentially express genes regulating blood coagulation. C_LIO_LIComparison with influenza A virus (IAV)-infected NHBEs revealed that the regulation of the extrinsic blood coagulation cascade is unique to SARS-CoV-2, and not seen with IAV infection. C_LI What do these findings mean?O_LIThe hyper-activation of the extrinsic blood coagulation cascade and the suppression of the plasminogen activation system in SARS-CoV-2 infected epithelial cells may drive diverse coagulopathies in the lung and distal organ systems. C_LIO_LIThe gene transcription pattern in SARS-CoV-2 infected epithelial cells is distinct from IAV infected epithelial cells with regards to the regulation of blood coagulation. C_LI

CLAMP and Zelda function together as pioneer transcription factors to promote Drosophila zygotic genome activation

Duan, J. E. — 2020-07-15

During the essential and conserved process of zygotic genome activation (ZGA), chromatin accessibility must increase to promote transcription. Drosophila is a well-established model for defining mechanisms that drive ZGA. Zelda (ZLD) is a key pioneer transcription factor (TF) that promotes ZGA in the Drosophila embryo. However, many genomic loci that contain GA-rich motifs become accessible during ZGA independent of ZLD. Therefore, we hypothesized that other early TFs that function with ZLD have not yet been identified, especially those that are capable of binding to GA-rich motifs such as CLAMP. Here, we demonstrate that Drosophila embryonic development requires maternal CLAMP to: 1) activate zygotic transcription; 2) increase chromatin accessibility at promoters of specific genes that often encode other essential TFs; 3) enhance chromatin accessibility to facilitate ZLD occupancy at a subset of key embryonic promoters. Thus, maternal CLAMP functions with ZLD in a pioneer-like role to drive zygotic genome activation.

Ecological load and balancing selection in circumboreal barnacles

Nunez, J. C. B. — 2020-07-19

Acorn barnacle adults experience environmental heterogeneity at various spatial scales of their circumboreal habitat, raising the question of how adaptation to high environmental variability is maintained in the face of strong juvenile dispersal and mortality. Here we show that 4% of genes in the barnacle genome experience balancing selection across the entire range of the species. Many of these genes harbor mutations maintained across 2 million years of evolution between the Pacific and Atlantic oceans. These genes are involved in ion regulation, pain reception, and heat tolerance, functions which are essential in highly variable ecosystems. The data also reveal complex population structure within and between basins, driven by the trans-Arctic interchange and the last glaciation. Divergence between Atlantic and Pacific populations is high, foreshadowing the onset of allopatric speciation, and suggesting that balancing selection is strong enough to maintain functional variation for millions of years in the face of complex demography.

HIV coinfection is associated with low fitness rpoB variants in rifampicin-resistant Mycobacterium tuberculosis

Loiseau, C. — 2020-07-22

We analysed 312 drug-resistant genomes of Mycobacterium tuberculosis (Mtb) collected from HIV coinfected and HIV negative TB patients from nine countries with a high tuberculosis burden. We found that rifampicin-resistant Mtb strains isolated from HIV coinfected patients carried disproportionally more resistance-conferring mutations in rpoB that are associated with a low fitness in the absence of the drug, suggesting these low fitness rpoB variants can thrive in the context of reduced host immunity.

Neural representation of abstract task structure during generalization

Vaidya, A. R. — 2020-07-21

Cognitive models in psychology and neuroscience widely assume that the human brain maintains an abstract representation of tasks. This assumption is fundamental to theories explaining how we learn quickly, think creatively, and act flexibly. However, neural evidence for a verifiably generative abstract task representation has been lacking. Here, we report an experimental paradigm that requires forming such a representation to act adaptively in novel conditions without feedback. Using functional magnetic resonance imaging, we observed that abstract task structure was represented within left mid-lateral prefrontal cortex, bilateral precuneus and inferior parietal cortex. These results provide support for the neural instantiation of the long-supposed abstract task representation in a setting where we can verify its influence. Such a representation can afford massive expansions of behavioral flexibility without additional experience, a vital characteristic of human cognition.

Synergid calcium ion oscillations define a new feature of pollen tube reception critical for blocking interspecific hybridization

Ponvert, N. D. — 2020-07-24

Reproductive isolation leads to the evolution of distinct new species, however, the molecular mechanisms that promote and maintain reproductive barriers are elusive. In flowering plants sperm cells are immotile and are delivered to female gametes by the pollen grain which germinates a polarized extension, the pollen tube, into floral tissue. After growing via polar extension to the female gametophyte, the pollen tube signals with receptive female cells flanking the egg in two distinct steps. If signaling is successful, the pollen tube releases sperm cells for fusion with female gametes. To better understand cell-cell recognition during reproduction, we investigated calcium ion dynamics associated with interspecific cell signaling between mating partners. We observed that interspecific pollen tubes successfully complete initial cell-cell signaling, but fail during later phases of pollen tube reception. Our work refines our understanding of pollen tube reception as a critical block to interspecific hybridization. Our results also shed light on the functional significance of the two phases of pollen tube reception, and implicate the second step as being the stage during which the pollen tube presents its genetic identity.

ABO genetic variation in Neanderthals and Denisovans

Fox, K. — 2020-07-27

1Variation at the ABO locus was one of the earliest sources of data in the study of human population identity and history, and to this day remains widely genotyped due to its importance in blood and tissue transfusions. Here, we look at ABO blood type variants in our archaic relatives: Neanderthals and Denisovans. Our goal is to understand the genetic landscape of the ABO gene in archaic humans, and how it relates to modern human ABO variation. We found two derived Neanderthal variants of the O allele in the Siberian Neanderthals (O1 and O2), one of these variants is shared with an European Neanderthal, who is a heterozygote for this O1 variant and a rare cis-AB variant. The Denisovan individual is heterozygous for two ancestral variant of the O1 allele, similar to variants found widely in modern humans. Perhaps more surprisingly, the O2 allele variant found in Siberian Neanderthals can be found at low frequencies in modern Europeans and Southeast Asians, and the O1 allele variant found in Siberian and European Neanderthal is also found at very low frequency in modern East Asians. Our genetic distance analyses suggest both alleles survive in modern humans due to inbreeding with Neanderthals. We find that the sequence backgrounds of the surviving Neanderthal-like O alleles in modern humans retain a higher sequence divergence than other surviving Neanderthal genome fragments, supporting a view of balancing selection operating in the Neanderthal ABO alleles by retaining highly diverse haplotypes compared to portions of the genome evolving neutrally.

Peripheral voltage-gated calcium channels in skin are essential for transient neurogenic thermal hyperalgesia in mice

DuBreuil, D. M. — 2020-07-29

Voltage-gated CaV2.2 calcium channels are expressed in nociceptors, at pre-synaptic terminals, soma, and axons. CaV2.2 channel inhibitors applied to the spinal cord relieve pain in humans and rodents, especially during pathological pain, but a biological function of nociceptor CaV2.2 channels in processing of nociception, outside pre-synaptic terminals, is not explored. Here, we demonstrate that functional CaV2.2 channels in skin are required for thermal hyperalgesia following intraplantar capsaicin exposure. We provide evidence that CaV2.2 channels at nociceptor free endings release inflammatory signals, ATP and IL-1{beta}. We assess the role of CaV2.2 splice isoforms to capsaicin-induced hyperalgesia measured by thermal and mechanical stimuli. Our data reveal a critical role for peripheral CaV2.2 channels in skin in neurogenic thermal hyperalgesia but not in mechanical hypersensitivity. Inhibition, or the complete lack, of peripheral CaV2.2 channels blunts the hyperalgesia response in vivo.

Natural Killer cell activation, reduced ACE2, TMPRSS2, cytokines G-CSF, M-CSF and SARS-CoV-2-S pseudovirus infectivity by MEK inhibitor treatment of human cells

Zhou, L. — 2020-08-03

COVID-19 affects vulnerable populations including elderly individuals and patients with cancer. Natural Killer (NK) cells and innate-immune TRAIL suppress transformed and virally-infected cells. ACE2, and TMPRSS2 protease promote SARS-CoV-2 infectivity, while inflammatory cytokines IL-6, or G-CSF worsen COVID-19 severity. We show MEK inhibitors (MEKi) VS-6766, trametinib and selumetinib reduce ACE2 expression in human cells. In some human cells, remdesivir increases ACE2-promoter luciferase-reporter expression, ACE2 mRNA and protein, and ACE2 expression is attenuated by MEKi. In serum-deprived and stimulated cells treated with remdesivir and MEKi we observed correlations between pRB, pERK, and ACE2 expression further supporting role of proliferative state and MAPK pathway in ACE2 regulation. We show elevated cytokines in COVID-19-(+) patient plasma (N=9) versus control (N=11). TMPRSS2, inflammatory cytokines G-CSF, M-CSF, IL-1, IL-6 and MCP-1 are suppressed by MEKi alone or with remdesivir. We observed MEKi stimulation of NK-cell killing of target-cells, without suppressing TRAIL-mediated cytotoxicity. Pseudotyped SARS-CoV-2 virus with a lentiviral core and SARS-CoV-2 D614 or G614 SPIKE (S) protein on its envelope infected human bronchial epithelial cells, small airway epithelial cells, or lung cancer cells and MEKi suppressed infectivity of the pseudovirus. We show a drug class-effect with MEKi to stimulate NK cells, inhibit inflammatory cytokines and block host-factors for SARS-CoV-2 infection leading also to suppression of SARS-CoV-2-S pseudovirus infection of human cells. MEKi may attenuate SARS-CoV-2 infection to allow immune responses and antiviral agents to control disease progression.

Adaptive learning through temporal dynamics of state representation

Razmi, N. — 2020-08-04

People adjust their learning rate rationally according to local environmental statistics and calibrate such adjustments based on the broader statistical context. To date, no theory has captured the observed range of adaptive learning behaviors or the complexity of its neural correlates. Here, we attempt to do so using a neural network model that learns to map an internal context representation onto a behavioral response via supervised learning. The network shifts its internal context upon receiving supervised signals that are mismatched to its output, thereby changing the "state" to which feedback is associated. A key feature of the model is that such state transitions can either increase learning or decrease learning depending on the duration over which the new state is maintained. Sustained state transitions that occur after changepoints facilitate faster learning and mimic network reset phenomena observed in the brain during rapid learning. In contrast, state transitions after one-off outlier events are short-lived, thereby limiting the impact of outlying observations on future behavior. State transitions in our model provide the first mechanistic interpretation for bidirectional learning signals, such the p300, that relate to learning differentially according to the source of surprising events and may also shed light on discrepant observations regarding the relationship between transient pupil dilations and learning. Taken together, our results demonstrate that dynamic latent state representations can afford normative inference and provide a coherent framework for understanding neural signatures of adaptive learning across different statistical environments. Significance StatementHow humans adjust their sensitivity to new information in a changing world has remained largely an open question. Bridging insights from normative accounts of adaptive learning and theories of latent state representation, here we propose a feed-forward neural network model that adjusts its learning rate online by controlling the speed of transitioning its internal state representations. Our model proposes a mechanistic framework for explaining learning under different statistical contexts, explains previously observed behavior and brain signals, and makes testable predictions for future experimental studies.

Towards A 3D Chromosome Shape Alphabet

Soto, C. J. — 2020-08-05

The study of the 3-dimensional (3D) structure of chromosomes - the largest macromolecules in biology - is one of the most challenging to date in structural biology. Here, we develop a novel representation of chromosomes, as sequences of shape letters from a finite shape alphabet, which provides a compact and efficient way to analyze ensembles of chromosome shape data, akin to the analysis of texts in a language by using letters. We construct a Chromosome Shape Alphabet (CSA) from an ensemble of chromosome 3D structures inferred from Hi-C data - via SIMBA3D or other methods - by segmenting curves based on topologically associating domains (TADs) boundaries, and by clustering all TADs 3D structures into groups of similar shapes. The median shapes of these groups, with some pruning and processing, form the Chromosome Shape Letters (CSLs) of the alphabet. We provide a proof-of-concept for these CSLs by reconstructing independent test curves using only CSLs (and corresponding transformations) and comparing these reconstructions with the original curves. Finally, we demonstrate how CSLs can be used to summarize the variability of shapes in an ensemble of chromosome 3D structures using generalized sequence logos.

Super-human cell death detection with biomarker-optimized neural networks

Linsley, J. W. — 2020-08-05

Cell death is an essential process in biology that must be accounted for in live microscopy experiments. Nevertheless, cell death is difficult to detect without perturbing experiments with stains, dyes or biosensors that can bias experimental outcomes, lead to inconsistent results, and reduce the number of processes that can be simultaneously labelled. These additional steps also make live microscopy difficult to scale for high-throughput screening because of the cost, labor, and analysis they entail. We address this fundamental limitation of live microscopy with biomarker-optimized convolutional neural networks (BO-CNN): computer vision models trained with a ground truth biosensor that detect live cells with superhuman, 96% accuracy more than 100 times faster than previous methods. Our models learn to identify important morphological characteristics associated with cell vitality without human input or additional perturbations, and to generalize to other imaging modalities and cell types for which they have no specialized training. We demonstrate that we can interpret decisions from BO-CNN models to gain biological insight into the patterns they use to achieve superhuman accuracy. The BO-CNN approach is broadly useful for live microscopy, and affords a powerful new paradigm for advancing the state of high-throughput imaging in a variety of contexts.

CIVET-Macaque: an automated pipeline for MRI-based cortical surface generation and cortical thickness in macaques

Lepage, C. — 2020-08-05

The MNI CIVET pipeline for automated extraction of cortical surfaces and evaluation of cortical thickness from in-vivo human MRI has been extended for processing macaque brains. Processing is performed based on the NIMH Macaque Template (NMT), as the reference template, with the anatomical parcellation of the surface following the D99 and CHARM atlases. The modifications needed to adapt CIVET to the macaque brain are detailed. Results have been obtained using CIVET-macaque to process the anatomical scans of the 31 macaques used to generate the NMT and another 95 macaques from the PRIME-DE initiative. It is anticipated that the open usage of CIVET-macaque will promote collaborative efforts in data collection and processing, sharing, and automated analyses from which the non-human primate brain imaging field will advance.

Molecular details of protein condensates probed by microsecond-long atomistic simulations

Zheng, W. — 2020-08-06

The formation of membraneless organelles in cells commonly occurs via liquid-liquid phase separation (LLPS), and is in many cases driven by multivalent interactions between intrinsically disordered proteins (IDPs). Molecular simulations can reveal the specific amino acid interactions driving LLPS, which is hard to obtain from experiment. Coarse-grained simulations have been used to directly observe the sequence determinants of phase separation but have limited spatial resolution, while all-atom simulations have yet to be applied to LLPS due to the challenges of large system sizes and long time scales relevant to phase separation. We present a novel multiscale computational framework by obtaining initial molecular configurations of a condensed protein-rich phase from equilibrium coarse-grained simulations, and back mapping to an all-atom representation. Using the specialized Anton 2 supercomputer, we resolve microscopic structural and dynamical details of protein condensates through microsecond-scale all-atom explicit-solvent simulations. We have studied two IDPs which phase separate in vitro: the low complexity domain of FUS and the N-terminal disordered domain of LAF-1. Using this approach, we explain the partitioning of ions between phases with low and high protein density, demonstrate that the proteins are remarkably dynamic within the condensed phase, identify the key residue-residue interaction modes stabilizing the dense phase, all while showing good agreement with experimental observations. Our approach is generally applicable to all-atom studies of other single and multi-component systems of proteins and nucleic acids involved in the formation of membraneless organelles.

A comprehensive macaque fMRI pipeline and hierarchical atlas

Jung, B. — 2020-08-06

Functional neuroimaging research in the non-human primate (NHP) has been advancing at a remarkable rate. The increase in available data establishes a need for robust analysis pipelines designed for NHP neuroimaging and accompanying template spaces to standardize the localization of neuroimaging results. Our group recently developed the NIMH Macaque Template (NMT), a high-resolution population average anatomical template and associated neuroimaging resources, providing researchers with a standard space for macaque neuroimaging (Seidlitz, Sponheim et al., 2018). Here, we release NMT v2, which includes both symmetric and asymmetric templates in stereotaxic orientation, with improvements in spatial contrast, processing efficiency, and segmentation. We also introduce the Cortical Hierarchy Atlas of the Rhesus Macaque (CHARM), a hierarchical parcellation of the macaque cerebral cortex with varying degrees of detail. These tools have been integrated into the neuroimaging analysis software AFNI (Cox, 1996) to provide a comprehensive and robust pipeline for fMRI processing, visualization and analysis of NHP data. AFNIs new @animal_warper program can be used to efficiently align anatomical scans to the NMT v2 space, and afni_proc.py integrates these results with full fMRI processing using macaque-specific parameters: from motion correction through regression modeling. Taken together, the NMT v2 and AFNI represent an all-in-one package for macaque functional neuroimaging analysis, as demonstrated with available demos for both task and resting state fMRI. HighlightsO_LIThe NMT v2, a stereotaxically aligned symmetric macaque template, is introduced. C_LIO_LIA new atlas (CHARM), defined on NMT v2, parcellates the cortex at six spatial scales. C_LIO_LIAFNIs @animal_warper aligns and maps data between monkey anatomicals and templates. C_LIO_LIAFNIs afni_proc.py facilitates monkey fMRI analysis with automated scripting and QC. C_LIO_LIDemos of macaque task and resting state fMRI analysis with these tools are provided. C_LI

Targeting Vitamin-D receptor (VDR) by a small molecule antagonist MeTC7 inhibits PD-L1 but controls THMYCN neuroblastoma growth PD-L1 independently

Singh, R. K. — 2020-08-17

Vitamin-D receptor (VDR) mRNA is enriched in malignant lung, ovarian and pancreatic tissues and showed poor prognoses. Calcitriol and stable or CRISPR-directed VDR upregulation increased PD-L1mRNA and protein expression in cancer cells in-vitro. A ChIP assay showed the binding of VDR with VDREPD-L1. Stattic, a STAT3 phosphorylation inhibitor blocked calcitriol or VDR overexpression induced PD-L1 upregulation. MeTC7, a VDR antagonist developed by us, reduced PD-L1 expression on macrophages, ovarian, lung, breast, and pancreatic cancer cells in-vitro. In radiotherapy inducible PD-L1 model of orthotopic MC38 murine colon cancer, MeTC7 decreased PD-L1 surface expression, suppressed inflammatory monocytes (IMs) population and increased intra-tumoral CD69+PD1+CD8+T-cells. Intriguingly, MeTC7 reduced TH-MYCN transgenic neuroblastoma tumor growth without affecting PD-L1 and tumor immune milieu. In summary, Vitamin-D/VDR drives PD-L1 expression on cancer cells via STAT-3. Inhibiting VDR exhibited anti-checkpoint effects in orthotopic colon tumors, whereas PDL1-independent and anti-VDR/MYCN effects controlled growth of transgenic neuroblastoma and xenografted tumors. SummaryVitamin-D/VDR induces PD-L1 expression on cancer cells via STAT-3; and targeting VDR by a novel small molecule antagonist MeTC7 exhibits both anti-PD-L1 and anti-VDR/MYCN effects in tumor models.

Sleep-dependent offline performance gain in visual perceptual learning is consistent with a learning-dependent model

Tamaki, M. — 2020-08-17

Are the sleep-dependent offline performance gains of visual perceptual learning (VPL) consistent with a use-dependent or learning-dependent model? Here, we found that a use-dependent model is inconsistent with the offline performance gains in VPL. In two training conditions with matched visual usages, one generated VPL (learning condition), while the other did not (interference condition). The use-dependent model predicts that slow-wave activity (SWA) during posttraining NREM sleep in the trained region increases in both conditions, in correlation with offline performance gains. However, compared with those in the interference condition, sigma activity, not SWA, during NREM sleep and theta activity during REM sleep, source-localized to the trained early visual areas, increased in the learning condition. Sigma activity correlated with offline performance gain. These significant differences in spontaneous activity between the conditions suggest that there is a learning-dependent process during posttraining sleep for the offline performance gains in VPL.

Effects of Peripheral Haptic Feedback on Intracortical Brain-Computer Interface Control and Associated Sensory Responses in Motor Cortex

Deo, D. R. — 2020-08-18

Intracortical brain-computer interfaces (iBCIs) provide people with paralysis a means to control devices with signals decoded from brain activity. Despite recent impressive advances, these devices still cannot approach able-bodied levels of control. To achieve naturalistic control and improved performance of neural prostheses, iBCIs will likely need to include proprioceptive feedback. With the goal of providing proprioceptive feedback via mechanical haptic stimulation, we aim to understand how haptic stimulation affects motor cortical neurons and ultimately, iBCI control. We provided skin shear haptic stimulation as a substitute for proprioception to the back of the neck of a person with tetraplegia. The neck location was determined via assessment of touch sensitivity using a monofilament test kit. The participant was able to correctly report skin shear at the back of the neck in 8 unique directions with 65% accuracy. We found motor cortical units that exhibited sensory responses to shear stimuli, some of which were strongly tuned to the stimuli and well modeled by cosine-shaped functions. We also demonstrated online iBCI cursor control with continuous skin-shear feedback driven by decoded command signals. Cursor control performance increased slightly but significantly when the participant was given haptic feedback, compared to the purely visual feedback condition.

CD300lf conditional knockout mouse reveals strain-specific cellular tropism for murine norovirus

Graziano, V. R. — 2020-08-21

Noroviruses are a leading cause of gastrointestinal infection in humans and mice. Understanding human norovirus (HuNoV) cell tropism has important implications for our understanding of viral pathogenesis. Murine norovirus (MNoV) is extensively used as a surrogate model for HuNoV. We previously identified CD300lf as the receptor for MNoV. Here, we generated a Cd300lf conditional knockout (CD300lfF/F) mouse to elucidate the cell tropism of persistent and non-persistent strains of murine norovirus. Using this mouse model, we demonstrate that CD300lf expression on intestinal epithelial cells (IECs), and on tuft cells in particular, is essential for transmission of the persistent MNoV strain CR6 (MNoVCR6) in vivo. In contrast, the nonpersistent MNoV strain CW3 (MNoVCW3) does not require CD300lf expression on IECs for infection. However, deletion of CD300lf in myelomonocytic cells (LysM Cre+) partially reduces CW3 viral load in lymphoid and intestinal tissues. Disruption of CD300lf expression on B cells (CD19 Cre), neutrophils (Mrp8 Cre), and dendritic cells (CD11c Cre) did not affect CW3 viral RNA levels. Finally, we show that the transcription factor STAT1, which is critical for the innate immune response, partially restricts the cell tropism of MNoVCW3 to LysM+ cells. Taken together, these data demonstrate that CD300lf expression on tuft cells is essential for MNoVCR6, that myelomonocytic cells are a major, but not exclusive, target cell of MNoVCW3, and that STAT1 signaling restricts the cellular tropism of MNoVCW3. This provides the first genetic system to study the cell type-specific role of CD300lf in norovirus pathogenesis. IMPORTANCEHuman noroviruses (HuNoVs) are a leading cause of gastroenteritis resulting in up to 200,000 deaths each year. The receptor and cell tropism of HuNoV in immunocompetent humans are unclear. We use murine norovirus (MNoV) as a model for HuNoV. We recently identified CD300lf as the sole physiologic receptor for MNoV. Here, we leverage this finding to generate a Cd300lf conditional knockout mouse to decipher the contributions of specific cell types to MNoV infection. We demonstrate that persistent MNoVCR6 requires CD300lf expression on tuft cells. In contrast, multiple CD300lf+ cell types, dominated by myelomonocytic cells, are sufficient for non-persistent MNoVCW3 infection. CD300lf expression on epithelial cells, B cells, neutrophils, and dendritic cells is not critical for MNoVCW3 infection. Mortality associated with MNoVCW3 strain in Stat1-/- mice does not require CD300lf expression on LysM+ cells, highlighting that both CD300lf receptor expression and innate immunity regulate MNoV cell tropism in vivo.

Behavioral assays to study neural development in Xenopus laevis tadpoles

Khakhalin, A. S. — 2020-08-22

Escape responses, orienting reflexes, and social behaviors in Xenopus laevis tadpoles have been well documented in the literature (Lee et al. 2010; Roberts et al. 2000; Simmons et al. 2004; Katz et al. 1981; Villinger and Waldman 2012). In this article, we describe several behavioral protocols that together allow researchers efficiently (in terms of financial cost and time investment) and effectively assess developmental abnormalities in pre-metamorphic Xenopus tadpoles.

Emergence of Non-Canonical Parvalbumin-Containing Interneurons in Hippocampus of a Murine Model of Type I Lissencephaly

Ekins, T. G. — 2020-08-21

Type I lissencephaly is a neuronal migration disorder caused by haploinsuffiency of the LIS1 gene and is characterized in humans by agyria, mislamination of brain structures, developmental delays, and epilepsy. Here, we investigate the impact of LIS1 mutation on the cellular migration, morphophysiology, microcircuitry and genomics of mouse hippocampal CA1 parvalbumin-containing inhibitory interneurons (PV+INTs). We find that WT PV+INTs consist of two physiological subtypes (80% fast-spiking (FS), 20% non-fast-spiking (NFS)) and four morphological subtypes (basket, axo-axonic, bistratified, radiatum-targeting). We also discover that cell-autonomous mutations within interneurons disrupts morphological development of PV+INTs and results in the emergence of a non-canonical "intermediate spiking (IS)" subset of PV+INTs. In the GlobalLis mutant, IS/NFS cells become the dominant PV+INT subtypes (56%) and the percentage of FS cells shrinks to 44%. We also find that IS/NFS cells are prone to entering depolarizing block, causing them to temporarily lose the ability to initiate action potentials and control network excitation, potentially promoting seizures. Finally, single-cell nuclear RNAsequencing of PV+INTs revealed several misregulated genes related to morphogenesis, cellular excitability, and synapse formation.

Transcriptional subtype-specific microenvironmental crosstalk and tumor cell plasticity in metastatic pancreatic cancer

Raghavan, S. — 2020-08-25

Bulk transcriptomic studies have defined classical and basal-like gene expression subtypes in pancreatic ductal adenocarcinoma (PDAC) that correlate with survival and response to chemotherapy; however, the underlying mechanisms that govern these subtypes and their heterogeneity remain elusive. Here, we performed single-cell RNA-sequencing of 23 metastatic PDAC needle biopsies and matched organoid models to understand how tumor cell-intrinsic features and extrinsic factors in the tumor microenvironment (TME) shape PDAC cancer cell phenotypes. We identify a novel cancer cell state that co-expresses basal-like and classical signatures, demonstrates upregulation of developmental and KRAS-driven gene expression programs, and represents a transitional intermediate between the basal-like and classical poles. Further, we observe structure to the metastatic TME supporting a model whereby reciprocal intercellular signaling shapes the local microenvironment and influences cancer cell transcriptional subtypes. In organoid culture, we find that transcriptional phenotypes are plastic and strongly skew toward the classical expression state, irrespective of genotype. Moreover, we show that patient-relevant transcriptional heterogeneity can be rescued by supplementing organoid media with factors found in the TME in a subtype-specific manner. Collectively, our study demonstrates that distinct microenvironmental signals are critical regulators of clinically relevant PDAC transcriptional states and their plasticity, identifies the necessity for considering the TME in cancer modeling efforts, and provides a generalizable approach for delineating the cell-intrinsic versus -extrinsic factors that govern tumor cell phenotypes.

miRNA-mediated loss of m6A increases nascent translation in glioblastoma

Zepecki, J. P. — 2020-08-29

Within the glioblastoma cellular niche, glioma stem cells (GSCs) can give rise to differentiated glioma cells (DGCs) and, when necessary, DGCs can reciprocally give rise to GSCs to maintain the cellular equilibrium necessary for optimal tumor growth. Here, using ribosome profiling, transcriptome and m6A RNA sequencing, we show that GSCs from patients with different subtypes of glioblastoma share a set of transcripts, which exhibit a pattern of m6A loss and increased protein translation during differentiation. The target sequences of a group of miRNAs overlap the canonical RRACH m6A motifs of these transcripts, many of which confer a survival advantage in glioblastoma. Ectopic expression of the RRACH-binding miR-145 induces loss of m6A, formation of FTO/AGO1/ILF3/miR-145 complexes on a clinically relevant tumor suppressor gene (CLIP3) and significant increase in its nascent translation. Inhibition of miR-145 maintains RRACH m6A levels of CLIP3 and inhibits its nascent translation. This study highlights a critical role of miRNAs in assembling complexes for m6A demethylation and induction of protein translation during GSC state transition. Author SummaryCellular plasticity and epigenetic adaptation of human glioblastoma stem cells to the tumor microenvironment is a hallmark of this devastating disease. With our present work, we discover the relationship between miRNAs and the RNA methylation machinery in human glioblastoma and show how miRNA-induced loss of m6A results in increase in protein translation of clinically important transcripts during glioblastoma stem cell differentiation. Leveraging the dynamic functions of these miRNAs can be important in the design of optimal therapeutics targeted at cancer cell plasticity.

Homeotic Regulation of Olfactory Receptor Choice via NFI-dependent Heterochromatic Silencing and Genomic Compartmentalization

Bashkirova, E. — 2020-08-30

Expression of one out of >1000 olfactory receptor (OR) genes is stochastic but, yet, spatially organized in stereotypic anatomical segments, or "zones", along the dorsoventral axis of the mouse olfactory epithelium. We discovered that zonal OR expression is specified by OR chromatin structure and genome architecture during olfactory neuron differentiation. Specifically, across every zone dorsally expressed ORs have higher levels of heterochromatic marks and long-range contacts than ORs expressed ventrally. However, OR heterochromatin levels and frequency of genomic contacts between ORs gradually increase towards ventral zones. Consequently, ORs from dorsal indexes accumulate high H3K9me3/H3K79me3 enrichment and become silenced in ventral zones, while ORs from ventral indexes lack activating long-range genomic interactions and, thus, cannot be chosen in dorsal segments. This process is regulated by NFIA, B, and X gradients along the dorsoventral axis, triple deletion of which causes homeotic transformations on zonal OR expression, heterochromatin formation, and genomic compartmentalization.

Confinement Discerns Swarmers from Planktonic Bacteria

Chen, W. — 2020-08-30

Powered by flagella, many bacterial species exhibit collective motion on a solid surface commonly known as swarming. As a natural example of active matter, swarming is also an essential biological phenotype associated with virulence, chemotaxis, and host pathogenesis. Physical changes like cell elongation and hyper flagellation have been shown to accompany the swarming phenotype. However, less noticeable, are the contrasts of collective motion between the swarming cells and the planktonic cells of comparable cell density. Here, we show that confining bacterial movement in designed dimensions allows distinguishing bacterial swarming from collective swimming. We found that on a soft agar plate, a novel bacterial strain Enterobacter sp. SM3 exhibited different motion patterns in swarming and planktonic states when confined to circular microwells of a specific range of sizes. When the confinement diameter was between 40 m and 90 m, swarming SM3 formed a single swirl motion pattern in the microwells whereas planktonic SM3 showed multiple swirls. Similar differential behavior is observed across a range of randomly selected gram-negative bacteria. We hypothesize that the "rafting behavior" of the swarming bacteria upon dilution might account for the motion pattern difference. We verified our conjectures via numerical simulations where swarming cells are modeled with lower repulsion and more substantial alignment force. The novel technical approach enabled us to observe swarming on a non-agar tissue surface for the first time. Our work provides the basis for characterizing bacterial swarming under more sophisticated environments, such as polymicrobial swarmer detection, and in vivo swarming exploration.

Newly regenerated axons through a cell-containing biomaterial scaffold promote reorganization of spinal circuitry and restoration of motor functions with epidural electrical stimulation.

Siddiqui, A. M. — 2020-09-09

We report the effect of newly regenerated neural fibers via bioengineered scaffold on reorganization of spinal circuitry and restoration of motor functions with electrical epidural stimulation (EES) after spinal transection (ST). Restoration across multiple modalities was evaluated for 7 weeks after ST with implanted scaffold seeded with Schwann cells, producing neurotrophic factors and with rapamycin microspheres. Gradual improvement in EES-facilitated stepping was observed in animals with scaffolds, although, no significant difference in stepping ability was found between groups without EES. Similar number of regenerated axons through the scaffolds was found in rats with and without EES-enabled training. Re-transection through the scaffold at week 6, reduced EES-enabled motor function, remaining higher compared to rats without scaffolds. The combination of scaffolds and EES-enabled training demonstrated synaptic changes below the injury. These findings indicate that sub-functional connectivity with regenerated across injury fibers can reorganize of sub-lesional circuitry, facilitating motor functions recovery with EES.

Coordination of two enhancers drives expression of olfactory trace amine-associated receptors

Fei, A. — 2020-09-11

Olfactory sensory neurons (OSNs) are functionally defined by their expression of a unique odorant receptor (OR). Mechanisms underlying singular OR expression are well studied, and involve a massive cross-chromosomal enhancer interaction network. Trace amine-associated receptors (TAARs) form a distinct family of olfactory receptors, and here we find that mechanisms regulating Taar gene choice display many unique features. The epigenetic signature of Taar genes in TAAR OSNs is different from that in OR OSNs. We further identify that two TAAR enhancers conserved across placental mammals are absolutely required for expression of the entire Taar gene repertoire. Deletion of either enhancer dramatically decreases the expression probabilities of different Taar genes, while deletion of both enhancers completely eliminates the TAAR OSN populations. In addition, both of the enhancers are sufficient to drive transgene expression in the partially overlapped TAAR OSNs. We also show that the TAAR enhancers operate in cis to regulate Taar gene expression. Our findings reveal a coordinated control of Taar gene choice in OSNs by two remote enhancers, and provide an excellent model to study molecular mechanisms underlying formation of an olfactory subsystem.

Wireless Ensembles of Sub-mm Microimplants Communicating as a Network near 1 GHz in a Neural Application

Lee, J. — 2020-09-13

Multichannel electrophysiological sensors and stimulators, especially those used for studying the nervous system, are most commonly based on monolithic microelectrode arrays. Such architecture limits the spatial flexibility of individual electrode placement, posing constraints for scaling to a large number of nodes, particularly across non-contiguous locations. We describe the design and fabrication of sub-millimeter size electronic microchips ("Neurograins") which autonomously perform neural sensing or electrical microstimulation, with emphasis on their wireless networking and powering. An [~]1 GHz electromagnetic transcutaneous link to an external telecom hub enables bidirectional communication and control at the individual neurograin level. The link operates on a customized time division multiple access (TDMA) protocol designed to scale up to 1000 neurograins. The system is demonstrated as a cortical implant in a small animal (rat) model with anatomical limitations restricting the implant to 48 neurograins. We suggest that the neurograin approach can be generalized to overcome many scalability issues for wireless sensors and actuators as implantable microsystems.

Dissociable influences of reward and punishment on adaptive cognitive control

Leng, X. — 2020-09-14

To invest effort into any cognitive task, people must be sufficiently motivated. Whereas prior research has focused primarily on how the cognitive control required to complete these tasks is motivated by the potential rewards for success, it is also known that control investment can be equally motivated by the potential negative consequence for failure. Previous theoretical and experimental work has yet to examine how positive and negative incentives differentially influence the manner and intensity with which people allocate control. Here, we develop and test a normative model of control allocation under conditions of varying positive and negative performance incentives. Our model predicts, and our empirical findings confirm, that rewards for success and punishment for failure should differentially influence adjustments to the evidence accumulation rate versus response threshold, respectively. This dissociation further enabled us to infer how motivated a given person was by the consequences of success versus failure. Author SummaryFrom the school to the workplace, whether someone achieves their goals is determined largely by the mental effort they invest in their tasks. Recent work has demonstrated both why and how people adjust the amount of effort they invest in response to variability in the rewards expected for achieving that goal. However, in the real world, we are motivated both by the positive outcomes our efforts can achieve (e.g., praise) and the negative outcomes they can avoid (e.g., rejection), and these two types of incentives can motivate adjustments not only in the amount of effort we invest but also the types of effort we invest (e.g., whether to prioritize performing the task efficiently or cautiously). Using a combination of computational modeling and a novel task that measures voluntary effort allocation under varying incentive conditions, we show that people should and do engage dissociable forms of mental effort in response to positive versus negative incentives. With increasing rewards for achieving their goal, they prioritize efficient performance, whereas with increasing penalties for failure they prioritize performing cautious performance. We further show that these dissociable strategies enable us to infer how motivated a given person was based on the positive consequences of success relative to the negative consequences of failure.

TIMEOR: a web-based tool to uncover temporal regulatory mechanisms from multi-omics data

Conard, A. M. — 2020-09-15

Uncovering how transcription factors (TFs) regulate their targets at the DNA, RNA and protein levels over time is critical to define gene regulatory networks (GRNs) in normal and diseased states. RNA-seq has become a standard method to measure gene regulation using an established set of analysis steps. However, none of the currently available pipeline methods for interpreting ordered genomic data (in time or space) use time series models to assign cause and effect relationships within GRNs, are adaptive to diverse experimental designs, or enable user interpretation through a web-based platform. Furthermore, methods which integrate ordered RNA-seq data with transcription factor binding data are urgently needed. Here, we present TIMEOR (Trajectory Inference and Mechanism Exploration with Omics data in R), the first web-based and adaptive time series multi-omics pipeline method which infers the relationship between gene regulatory events across time. TIMEOR addresses the critical need for methods to predict causal regulatory mechanism networks between TFs from time series multi-omics data. We used TIMEOR to identify a new link between insulin stimulation and the circadian rhythm cycle. TIMEOR is available at https://github.com/ashleymaeconard/TIMEOR.git.

Subcortical Atlas of the Rhesus Macaque (SARM) for Magnetic Resonance Imaging

Hartig, R. — 2020-09-18

Digitized neuroanatomical atlases are crucial for localizing brain structures and analyzing functional networks identified by magnetic resonance imaging (MRI). To aid in MRI data analysis, we have created a comprehensive parcellation of the rhesus macaque subcortex using a high-resolution ex vivo structural imaging scan. The structural scan and its parcellation were warped to the updated NIMH Macaque Template (NMT v2), an in vivo population template, where the parcellation was refined to produce the Subcortical Atlas of the Rhesus Macaque (SARM). The subcortical parcellation and nomenclature reflect those of the 4th edition of the Rhesus Monkey Brain in Stereotaxic Coordinates (RMBSC4; Paxinos et al., in preparation). The SARM features six parcellation levels, arranged hierarchically from fine regions-of-interest (ROIs) to broader composite regions, suited for fMRI studies. As a test, we ran a functional localizer for the dorsal lateral geniculate (DLG) nucleus in three macaques and found significant fMRI activation in this atlas region. The SARM has been made openly available to the neuroimaging community and can easily be used with common MR data processing software, such as AFNI, where the atlas can be embedded into the software alongside cortical macaque atlases. HighlightsO_LIWe present the Subcortical Atlas of the Rhesus Macaque (SARM). C_LIO_LISARM provides a neuroanatomical reference frame for neuroimaging analysis. C_LIO_LIThe entire subcortex is mapped, including the thalamus, basal ganglia, and brainstem. C_LIO_LIROIs are grouped hierarchically, making SARM useful at multiple spatial resolutions. C_LIO_LISARM is in the NMT v2 template space and complements the CHARM atlas for the cortex. C_LI

Inhibition of Streptococcus pneumoniae autolysins highlight distinct differences between chemical and genetic inactivation

Haubrich, B. A. — 2020-09-16

Despite renewed interest, development of chemical biology methods to study peptidoglycan metabolism has lagged in comparison to the glycobiology field in general. To address this, a panel of diamides were screened against the Gram-positive pathogen Streptococcus pneumoniae to identify inhibitors of bacterial growth. The screen identified the diamide fgkc as a narrow spectrum bacteriostatic inhibitor of S. pneumoniae growth with an MIC of 7.8 M. The diamide inhibited detergent-induced autolysis in a concentration dependent manner indicating peptidoglycan degradation as the mode-of-action. Genetic screening of autolysin mutants suggested LytB, an endo-N-acetylglucosaminidase, involved in cell division as the potential target. Surprisingly, biochemical, and phenotypic analysis contradicted the genetic screen results. Phenotypic studies with the{Delta} lytb strain illustrate the difference between genetic and chemical inactivation of autolysins. These findings suggest that meta-phenotypes including autolytic activity, cell morphology, and genetic screening can be the result of the complex interaction of one or more possible pathways that are connected to cell wall metabolism.

Dorsal Anterior Cingulate Cortex Encodes the Subjective Motivational Value of Cognitive Task Performance

Yee, D. M. — 2020-09-20

Humans can seamlessly combine value signals from diverse motivational incentives, yet it is not well-understood how these signals are "bundled" in the brain to modulate cognitive control. The dorsal anterior cingulate cortex (dACC) is theorized to integrate motivational value dimensions in the service of goal-directed action, though this hypothesis has yet to receive rigorous confirmation. In the present study, we examined the role of human dACC in motivational incentive integration. Healthy young adult men and women were scanned with fMRI while engaged in an experimental paradigm that quantifies the combined effects of liquid (e.g., juice, neutral, saltwater) and monetary incentives on cognitive task performance. Monetary incentives modulated trial-by-trial dACC activation, whereas block-related effects of liquid incentives on dACC activity were observed. When bundled together, incentive-related dACC modulation predicted fluctuations in both cognitive performance and self-report motivation ratings. Statistical mediation analyses suggest that dACC encoded the incentives in terms of their integrated subjective motivational value, and that this value signal was most proximally associated with task performance. Finally, we confirmed that these incentive integration effects were selectively present in dACC. Together, the results support an account in which dACC integrates motivational signals to compute the expected value of goal-directed cognitive control. Significance StatementHow are primary and secondary incentives integrated in the brain to influence goal-directed behavior? Using an innovative experimental fMRI paradigm that combines motivational incentives that have historically been studied independently between species (e.g., monetary rewards for humans, food rewards for animals), we examine the relationship between incentive motivational value and cognitive control allocation. We find evidence that the integrated incentive motivational value of combined incentives is encoded in human dorsal anterior cingulate cortex (dACC). Further, self-reported motivational shifts mediated the effects of incentive-modulated dACC activity on task performance, revealing convergence in how self-reported and experimentally-induced motivation are encoded in the human brain. Our findings may inform future translational studies examining affective/motivational and cognitive impairments in psychopathology (e.g., anxiety, depression, addiction).

Transsynaptic mapping of Drosophila mushroom body output neurons

Scaplen, K. M. — 2020-09-23

The Mushroom Body (MB) is a well-characterized associative memory structure within the Drosophila brain. Although previous studies have analyzed MB connectivity and provided a map of inputs and outputs, a detailed map of the downstream targets is missing. Using the genetic anterograde transsynaptic tracing tool, trans-Tango, we identified divergent projections across the brain and convergent downstream targets of the MB output neurons (MBONs). Our analysis revealed at least three separate targets that receive convergent input from MBONs: other MBONs, the fan shaped body (FSB), and the lateral accessory lobe (LAL). We describe, both anatomically and functionally, a multilayer circuit in which inhibitory and excitatory MBONs converge on the same genetic subset of FSB and LAL neurons. This circuit architecture provides an opportunity for the brain to update information and integrate it with previous experience before executing appropriate behavioral responses. Highlights-The postsynaptic connections of the output neurons of the mushroom body, a structure that integrates environmental cues with associated valence, are mapped using trans-Tango. -Mushroom body circuits are highly interconnected with several points of convergence among mushroom body output neurons (MBONs). -The postsynaptic partners of MBONs have divergent projections across the brain and convergent projections to select target neuropils outside the mushroom body important for multimodal integration. -Functional connectivity suggests the presence of multisynaptic pathways that have several layers of integration prior to initiation of an output response.

Single-cell characterization of transcriptomic heterogeneity in lymphoblastoid cell lines

Sorelle, E. D. — 2020-09-25

Lymphoblastoid Cell Lines (LCLs) are generated by transforming primary B cells with Epstein-Barr Virus (EBV) and are used extensively as model systems in viral oncology, immunology, and human genetics research. In this study, we characterized single-cell transcriptomic profiles of five LCLs and present a simple discrete-time simulation to explore the influence of stochasticity on LCL clonal evolution. Single-cell RNA sequencing revealed substantial phenotypic heterogeneity within and across LCLs with respect to immunoglobulin isotype; virus-modulated host pathways involved in survival, proliferation, and differentiation; viral replication state; and oxidative stress. This heterogeneity is likely attributable to intrinsic variance in primary B cells and host-pathogen dynamics. Stochastic simulations demonstrate that initial primary cell heterogeneity, random sampling, time in culture, and even mild differences in phenotype-specific fitness can contribute substantially to dynamic diversity in populations of nominally clonal cells.

Pathway Analysis within Multiple Human Ancestries Reveals Novel Signals for Epistasis in Complex Traits

Turchin, M. C. — 2020-09-25

Genome-wide association (GWA) studies have identified thousands of significant genetic associations in humans across a number of complex traits. However, the majority of these studies focus on linear additive relationships between genotypic and phenotypic variation. Epistasis, or non-additive genetic interactions, has been identified as a major driver of both complex trait architecture and evolution in multiple model organisms; yet, this same phenomenon is not considered to be a significant factor underlying human complex traits. There are two possible reasons for this assumption. First, most large GWA studies are conducted solely with European cohorts; therefore, our understanding of broad-sense heritability for many complex traits is limited to just one ancestry group. Second, current epistasis mapping methods commonly identify significant genetic interactions by exhaustively searching across all possible pairs of SNPs. In these frameworks, estimated epistatic effects size are often small and power can be low due to the multiple testing burden. Here, we present a case study that uses a novel region-based mapping approach to analyze sets of variants for the presence of epistatic effects across six diverse subgroups within the UK Biobank. We refer to this method as the "MArginal ePIstasis Test for Regions" or MAPIT-R. Even with limited sample sizes, we find a total of 245 pathways within the KEGG and REACTOME databases that are significantly enriched for epistatic effects in height and body mass index (BMI), with 67% of these pathways being detected within individuals of African ancestry. As a secondary analysis, we introduce a novel region-based "leave-one-out" approach to localize pathway-level epistatic signals to specific interacting genes in BMI. Overall, our results indicate that non-European ancestry populations may be better suited for the discovery of non-additive genetic variation in human complex traits -- further underscoring the need for publicly available, biobank-sized datasets of diverse groups of individuals.

Measuring effects of trainee professional development on research productivity: A cross-institutional meta-analysis

Brandt, P. — 2020-09-28

PhD-trained scientists are essential contributors to the workforce in diverse employment sectors that include academia, industry, government, and non-profit organizations. Hence, best practices for training the future biomedical workforce are of national concern. Complementing coursework and laboratory research training, many institutions now offer professional training that enables career exploration and develops a broad set of skills critical to various career paths. The National Institutes of Health funded academic institutions to design innovative programming to enable this professional development through a mechanism known as Broadening Experiences in Scientific Training (BEST). Programming at the BEST awardee institutions included career panels, skill-building workshops, job-searching workshops, site visits, and internships. An initial concern was since doctoral training is lengthy and requires focused attention on dissertation research, having students participate in additional complementary training activities might lengthen time to degree and hamper student research productivity. To address this concern, using time to degree and publication records as measures of efficiency and productivity, metrics were analyzed from ten BEST awardee institutions. Comparing doctoral students who participated to those who did not, results revealed that across these diverse academic institutions, there were no differences in time to degree or manuscript output. Furthermore, a few institutions even demonstrated a positive correlation between participation in career and professional development activities and productivity. Our findings suggest that doctoral students should be encouraged to participate in career and professional development opportunities to ensure their preparedness for a variety of diverse and important careers in the workforce. Significance StatementOur study is unique in that it compiled doctoral degree durations at ten different universities, recorded individual participation in career and professional development activities in terms of dosage, and tracked individual engagement in real-time rather than relying on surveys sent to trainees after graduation. Participation in career and professional development activities, including internships, did not decrease efficiency or productivity. Our findings suggest that doctoral students should be encouraged to participate in career and professional development opportunities to ensure their preparedness for a variety of diverse and important careers in the workforce.

Luxotonic signals in human prefrontal cortex: A possible substrate for effects of light on mood and cognition

Sabbah, S. — 2020-09-29

Animal studies revealed a mood-regulating neural pathway linking intrinsically photosensitive retinal ganglion cells (ipRGCs) and the prefrontal cortex (PFC), involved in the pathophysiology of mood disorders. As humans too have luminance-encoding ipRGCs, we asked whether a similar pathway exist in humans. Here, fMRI was used to identify PFC regions and other areas exhibiting luminance-dependent signals. We report 29 human brain regions where activation either monotonically decreased or increased with luminance. Luxotonic activity was identified across the cerebral cortex, in diverse subcortical structures, and in the cerebellum, regions that have functions related to visual image formation, motor control, cognition, emotion, and reward processing. Light suppressed PFC activation level, the activation monotonically decreasing with increasing luminance. The sustained time course of light-evoked PFC responses, and their susceptibility to prior light exposure, most closely resembled those of ipRGCs. These findings offer a functional link between light exposure and PFC-mediated cognitive and affective phenomena.

Proteoglycan 4 (PRG4) expression and functionin dry eye associated inflammation

Menon, N. G. — 2020-10-01

PurposeDry eye disease (DED) affects hundreds of millions worldwide. Proteoglycan 4 (PRG4) has been shown to improve signs and symptoms of DED in humans. The objectives of this study were to characterize endogenous PRG4 expression by telomerase-immortalized human corneal epithelial (hTCEpi) cells, examine how exogenous recombinant human PRG4 (rhPRG4) modulates cytokine and chemokine secretion in response to TNF and IL-1{beta}, explore rhPRG4 as a potential substrate and/or inhibitor of MMP-9, and to understand how experimental dry (EDE) in mice affects PRG4 expression. MethodsPRG4 secretion was quantified by Western blotting and PRG4 expression by immunocytochemistry. Cytokine/chemokine release was measured by ELISA, and MMP-9 inhibition was quantified using an MMP-9 inhibitor kit. EDE was induced in mice, and PRG4 was visualized by immunohistochemistry in the cornea and Western blotting in lacrimal gland lysate. ResultshTCEpi cells synthesize and secrete PRG4 in vitro, which is inhibited by TNF and IL-1{beta}. TNF and IL-1{beta} significantly increased secretion of cytokine IL-6 and chemokines IL-8, IP-10, RANTES, and ENA-78, and several of these chemokines were downregulated after cotreatment with rhPRG4. Fluorescently-labelled rhPRG4 was internalized by hTCEpi cells. rhPRG4 was not digested by MMP-9 and inhibited in vitro activity of exogenous MMP-9 both in solution and in the presence of human tears. Finally, EDE decreased corneal and lacrimal gland expression of PRG4. ConclusionsThese results demonstrate rhPRG4s anti-inflammatory properties in the corneal epithelium and its contribution to ocular surface homeostasis, furthering our understanding of PRG4s immunomodulatory properties in the context of DED inflammation.

The history and evolution of the Denisovan-EPAS1 haplotype in Tibetans

Zhang, X. — 2020-10-02

Recent studies suggest that admixture with archaic hominins played an important role in facilitating biological adaptations to new environments. For example, interbreeding with Denisovans facilitated the adaptation to high altitude environments on the Tibetan Plateau. Specifically, the EPAS1 gene, a transcription factor that regulates the response to hypoxia, exhibits strong signatures of both positive selection and introgression from Denisovans in Tibetan individuals. Interestingly, despite being geographically closer to the Denisova cave, East Asian populations do not harbor as much Denisovan ancestry as populations from Melanesia. Recently, two studies have suggested two independent waves of Denisovan admixture into East Asians, one of which is shared with South Asians and Oceanians. Here we leverage data from EPAS1 in 78 Tibetan individuals to interrogate which of these two introgression events introduced the EPAS1 beneficial sequence into the ancestral population of Tibetans, and we use the distribution of introgressed segment lengths at this locus to infer the timing of the introgression and selection event. We find that the introgression event unique to East Asians most likely introduced the beneficial haplotype into the ancestral population of Tibetans around 43,000 (15,700-60,000) years ago, and selection started 12,000 (1,925-50,000) years ago. Our estimates suggest that one of the most convincing examples of adaptive introgression is in fact selection acting on standing archaic variation.

Uncovering biomarkers during therapeutic neuromodulation with PARRM: Period-based Artifact Reconstruction and Removal Method

Dastin-van Rijn, E. M. — 2020-10-09

Advances in device development have enabled concurrent stimulation and recording at adjacent locations in the central nervous system. However, stimulation artifacts obscure the sensed underlying neural activity. Here, we developed a novel method, termed Period-based Artifact Reconstruction and Removal Method (PARRM), to remove stimulation artifacts from neural recordings by leveraging the exact period of stimulation to construct and subtract a high-fidelity template of the artifact. Benchtop saline experiments, computational simulations, five unique in vivo paradigms across animal and human studies, and an obscured movement biomarker were used for validation. Performance was found to exceed that of state-of-the-art filters in recovering complex signals without introducing contamination. PARRM has several advantages: it is 1) superior in signal recovery; 2) easily adaptable to several neurostimulation paradigms; and 3) low-complexity for future on-device implementation. Real-time artifact removal via PARRM will enable unbiased exploration and detection of neural biomarkers to enhance efficacy of closed-loop therapies. SummaryOnline, real-time artifact removal via PARRM will enable unbiased exploration of neural biomarkers previously obscured by stimulation artifact.

Enhancement of synaptic AMPA receptors depends mutually on Src and PSD-95

Huang, X. — 2020-10-02

Synaptic incorporation and removal of AMPA receptors is highly regulated to modulate the strength of synaptic transmission for long-term synaptic plasticity during brain development and associative learning. PSD-932 and PSD-95, two paralogs of the DLG-MAGUK protein family of signaling scaffolds govern the synaptic incorporation and stabilization of AMPA receptors opposingly, with PSD-95 promoting and PSD-932 inhibiting it. The associated signaling mechanisms that control the synaptic incorporation and stabilization remain elusive. Here, we used domain swapping between the antagonizing signaling scaffolds to identify the protein motifs responsible for enhancing synaptic AMPA receptors and the associated signaling protein. We narrowed down multiple motifs in the N-terminal domain that are principally responsible for governing the enhancement by Src. Specific activation and inhibiting peptides revealed continuous activity of Src. Together, the results depict a mutual dependence of Src and PSD-95 in enhancing and maintaining synaptic AMPA receptors.

Parametric cognitive load reveals hidden costs in the neural processing of perfectly intelligible degraded speech

Ritz, H. — 2020-10-03

Speech is often degraded by environmental noise or hearing impairment. People can compensate for degradation, but this requires cognitive effort. Previous research has identified frontotemporal networks involved in effortful perception, but materials in these works were also less intelligible, and so it is not clear whether activity reflected effort or intelligibility differences. We used functional magnetic resonance imaging to assess the degree to which spoken sentences were processed under distraction, and whether this depended on speech quality even when intelligibility of degraded speech was matched to that of clear speech (i.e., 100%). On each trial, participants either attended to a sentence, or to a concurrent multiple object tracking (MOT) task that imposed parametric cognitive load. Activity in bilateral anterior insula reflected task demands: during the MOT task, activity increased as cognitive load increased, and during speech listening, activity increased as speech became more degraded. In marked contrast, activity in bilateral anterior temporal cortex was speech-selective, and gated by attention when speech was degraded. In this region, performance of the MOT task with a trivial load blocked processing of degraded speech whereas processing of clear speech was unaffected. As load increased, responses to clear speech in these areas declined, consistent with reduced capacity to process it. This result dissociates cognitive control from speech processing: substantially less cognitive control is required to process clear speech than is required to understand even very mildly degraded, 100% intelligible, speech. Perceptual and control systems clearly interact dynamically during real-world speech comprehension. Significance StatementSpeech is often perfectly intelligible even when degraded, e.g., by background sound, phone transmission, or hearing loss. How does degradation alter cognitive demands? Here, we use fMRI to demonstrate a novel and critical role for cognitive control in the processing of mildly degraded but perfectly intelligible speech. We compare speech that is matched for intelligibility but differs in putative control demands, dissociating cognitive control from speech processing. We also impose a parametric cognitive load during perception, dissociating processes that depend on tasks from those that depend on available capacity. Our findings distinguish between frontal and temporal contributions to speech perception and reveal a hidden cost to processing mildly degraded speech, underscoring the importance of cognitive control for everyday speech comprehension.

Applying Machine Learning to Increase Efficiency and Accuracy of Meta-Analytic Review

Gorelik, A. J. — 2020-10-08

The rapidly burgeoning quantity and complexity of publications makes curating and synthesizing information for meta-analyses ever more challenging. Meta-analyses require manual review of abstracts for study inclusion, which is time consuming, and variation among reviewer interpretation of inclusion/exclusion criteria for selecting a paper to be included in a review can impact a studys outcome. To address these challenges in efficiency and accuracy, we propose and evaluate a machine learning approach to capture the definition of inclusion/exclusion criteria using a machine learning model to automate the selection process. We trained machine learning models on a manually reviewed dataset from a meta-analysis of resilience factors influencing psychopathology development. Then, the trained models were applied to an oncology dataset and evaluated for efficiency and accuracy against trained human reviewers. The results suggest that machine learning models can be used to automate the paper selection process and reduce the abstract review time while maintaining accuracy comparable to trained human reviewers. We propose a novel approach which uses model confidence to propose a subset of abstracts for manual review, thereby increasing the accuracy of the automated review while reducing the total number of abstracts requiring manual review. Furthermore, we delineate how leveraging these models more broadly may facilitate the sharing and synthesis of research expertise across disciplines.

Permeabilization-free en bloc immunohistochemistry for correlative microscopy

Fulton, K. A. — 2020-10-08

A dense reconstruction of neuronal synaptic connectivity typically requires high-resolution 3D electron microscopy (EM) data, but EM data alone lacks functional information about neurons and synapses. One approach to augment structural EM datasets is with the fluorescent immunohistochemical (IHC) localization of functionally relevant proteins. We describe a protocol that obviates the requirement of tissue permeabilization in thick tissue sections, a major impediment for correlative pre-embedding IHC and EM. We demonstrate the permeabilization-free labeling of neuronal cell types, intracellular enzymes, and synaptic proteins in tissue sections hundreds of microns thick in multiple brain regions while simultaneously retaining the ultrastructural integrity of the tissue. Finally, we explore the utility of this protocol by performing proof-of-principle correlative experiments combining two-photon imaging of protein distributions and 3D electron microscopy.

Learning when effort matters: Neural dynamics underlying updating and adaptation to changes in performance efficacy

Grahek, I. — 2020-10-10

To determine how much cognitive control to invest in a task, people need to consider whether exerting control matters for obtaining rewards. In particular, they need to account for the efficacy of their performance - the degree to which rewards are determined by performance or by independent factors. Yet it remains unclear how people learn about their performance efficacy in an environment. Here we combined computational modeling with measures of task performance and EEG, to provide a mechanistic account of how people (a) learn and update efficacy expectations in a changing environment, and (b) proactively adjust control allocation based on current efficacy expectations. Across two studies subjects performed an incentivized cognitive control task while their performance efficacy (the likelihood that rewards are performance- contingent or random) varied over time. We show that people update their efficacy beliefs based on prediction errors - leveraging similar neural and computational substrates as those that underpin reward learning - and adjust how much control they allocate according to these beliefs. Using computational modeling, we show that these control adjustments reflect changes in information processing, rather than the speed-accuracy tradeoff. These findings demonstrate the neurocomputational mechanism through which people learn how worthwhile their cognitive control is.

The transcription factor CLAMP is required for neurogenesis in Drosophila melanogaster.

Tsiarli, M. A. — 2020-10-09

AbstractNeural stem cell (NSC) differentiation is controlled by cell-intrinsic and external signals from the stem cell niche including niche surface glia (SG). However, the mechanisms by which transcription factors drive NSC differentiation within the niche remain largely unknown. Here, we show that the Drosophila melanogaster transcription factor, Chromatin-linked adaptor for MSL proteins (CLAMP) is required for regulation of stemness and proliferation of NSCs, especially of the optic lobe (OL). CLAMP promotes transcription of genes involved in stemness, proliferation, and glial development and represses transcription of genes involved in neurogenesis and niche survival. Consistent with transcriptional changes, CLAMP promotes NSC proliferation and niche SG production, while lack of CLAMP severely and specifically impacts OL development. To identify potential mechanisms by which CLAMP may regulate brain development, we examined CLAMP motifs and available CLAMP ChIP-seq data to determine which genes may be direct versus indirect targets. CLAMP motifs are present at many target genes including the glial-determining gene, glial cells missing, while Tailless, the master regulator of OL-development is directly bound by CLAMP. In accordance to these results, in larval OL NSCs lacking CLAMP, Tailless levels are decreased dramatically, suggesting that CLAMP controls OL neurogenesis via Tailless. Overall, our results suggest that CLAMP regulates a transcriptional program which drives NSC proliferation and differentiation via cell-intrinsic and niche-dependent mechanisms that involve transcriptional regulation of Tailless and niche glia.

Elucidating the viral and host factors enabling the cross-species transmission of primate lentiviruses from simians to humans

Tebit, D. — 2020-10-13

The HIV-1 epidemic originated from a cross-species transmission of a primate lentivirus from chimpanzees to humans near the turn of the 18th century. Simian immunodeficiency viruses have been jumping between old world monkeys in West/Central Africa for thousands of years. So why did HIV-1 only emerge in the past century? This study examined the replicative fitness, transmission, restriction, and cytopathogenicity of 26 primate lentiviruses. Pairwise competitions of these primate lentiviruses revealed that SIVcpz had the highest replicative fitness in human or chimpanzee peripheral blood mononuclear cells, even higher fitness than HIV-1 group M strains responsible for 37 million infections worldwide. In contrast the "HIV-2 lineage" (SIVsmm, SIVmac, SIVagm, and HIV-2) had the lowest replicative fitness. SIVcpz strains were less inhibited by human restriction factors than the "HIV-2 lineage" strains, a restriction that was inversely correlated with replicative fitness. SIVcpz from the chimpanzee subspecies Pan troglodytes troglodytes (Ptt) was slightly more fit in human cells than the strains from Pt schweinfurthii (Pts). However, unlike all other primate lentiviruses (including the HIV-2 lineage), SIVcpz was nonpathogenic in human tonsillar tissue and did not deplete CD4+ T-cells, consistent with the slow or nonpathogenic disease observed in chimpanzees. Despite the close phylogenetic relationship between SIVcpz_Ptt and HIV-1, this epidemic was either caused by cross species transmission of a rare, undiscovered SIVcpz strain of higher virulence or higher virulence differentially evolved among HIV-1 subtypes during the human epidemic. Author summaryInvasion of wild animal habitats by humans can have devastating consequences for the human population as evident by the HIV-1 and SARS-CoV-2 epidemics. With SARS-CoV-2, a recent zoonotic jump, likely from bats, will help to identify a coronavirus progenitor. In contrast, simian immunodeficiency virus (SIV) jumped into humans over 100 years ago from a possibly extinct sub-species of chimpanzees and/or extinct lineage of SIV. We examined replicative fitness and pathogenesis of 26 different primate lentiviruses in human and chimpanzee primary lymphoid cells from blood and within tonsils. SIV from a specific chimpanzee species and lowland gorillas were the most capable of infecting and replicating in human and chimp lymphoid cells but they did not result in the pathogenesis related to disease in humans. In contrast, SIV from other old world monkeys were pathogenic but could not replicate efficiently in human cells. We propose the main HIV-1 is derived from a distinct jump of a very rare SIV strain in chimps leading to AIDS pandemic.

Noise correlations for faster and more robust learning.

Nassar, M. R. — 2020-10-15

Distributed population codes are ubiquitous in the brain and pose a challenge to downstream neurons that must learn an appropriate readout. Here we explore the possibility that this learning problem is simplified through inductive biases implemented by stimulus-independent noise correlations that constrain learning to task-relevant dimensions. We test this idea in a set of neural networks that learn to perform a perceptual discrimination task. Correlations among similarly tuned units were manipulated independently of overall population signal-to-noise ratio in order to test how the format of stored information affects learning. Higher noise correlations among similarly tuned units led to faster and more robust learning, favoring homogenous weights assigned to neurons within a functionally similar pool, and could emerge through Hebbian learning. When multiple discriminations were learned simultaneously, noise correlations across relevant feature dimensions sped learning whereas those across irrelevant feature dimensions slowed it. Our results complement existing theory on noise correlations by demonstrating that when such correlations are produced without significant degradation of the signal-to-noise ratio, they can improve the speed of readout learning by constraining it to appropriate dimensions. Significance statementPositive noise correlations between similarly tuned neurons theoretically reduce the representational capacity of the brain, yet they are commonly observed, emerge dynamically in complex tasks, and persist even in well-trained animals. Here we show that such correlations, when embedded in a neural population with a fixed signal to noise ratio, can improve the speed and robustness with which an appropriate readout is learned. In a simple discrimination task such correlations can emerge naturally through Hebbian learning. In more complex tasks that require multiple discriminations, correlations between neurons that similarly encode the task-relevant feature improve learning by constraining it to the appropriate task dimension.

Control of archetype BK polyomavirus miRNA expression

Imperiale, M. J. — 2020-10-20

BK polyomavirus (BKPyV) is a ubiquitous human pathogen, with over 80% of adults worldwide persistently infected. BKPyV infection is usually asymptomatic in healthy people; however, it causes polyomavirus-associated nephropathy in renal transplant patients and hemorrhagic cystitis in bone marrow transplant patients. BKPyV has a circular, double-stranded DNA genome that is divided genetically into three parts: an early region, a late region, and a non-coding control region (NCCR). The NCCR contains the viral DNA replication origin and cis-acting elements regulating viral early and late gene expression. It was previously shown that a BKPyV miRNA expressed from the late strand regulates viral large T antigen expression and limits the replication capacity of archetype BKPyV. A major unanswered question in the field is how expression of the viral miRNA is regulated. Typically, miRNA is expressed from introns in cellular genes but there is no intron readily apparent in the BKPyV from which the miRNA could derive. Here we provide evidence for primary RNA transcripts that circle the genome more than once and include the NCCR. We identified splice junctions resulting from splicing of primary transcripts circling the genome more than once, and Sanger sequencing of RT-PCR products indicates that there are viral transcripts that circle the genome up to four times. Our data suggest that the miRNA is expressed from the intron of these greater-than-genome size primary transcripts.

P53-independent restoration of p53 pathway in tumors with mutated p53 through ATF4 transcriptional modulation by ERK1/2 and CDK9

Tian, X. — 2020-10-20

A long-term goal in the cancer-field has been to develop strategies for treating p53-mutated tumors. A novel small-molecule, PG3-Oc, restores p53 pathway-signaling in tumor cells with mutant-p53, independently of p53/p73. PG3-Oc partially upregulates the p53-transcriptome (13.7% of public p53 target-gene dataset; 15.2% of in-house dataset) and p53-proteome (18%, HT29; 16%, HCT116-p53-/-). Bioinformatic analysis indicates critical p53-effectors of growth-arrest (p21), apoptosis (PUMA, DR5, Noxa), autophagy (DRAM1), and metastasis-suppression (NDRG1) are induced by PG3-Oc. ERK1/2- and CDK9-kinases are required to upregulate ATF4 by PG3-Oc which restores p53 transcriptomic-targets in cells without functional-p53. PG3-Oc represses MYC (ATF4-independent), and upregulates PUMA (ATF4-dependent) in mediating cell death. With largely nonoverlapping transcriptomes, induced-ATF4 restores p53 transcriptomic targets in drug-treated cells including functionally important mediators such as PUMA and DR5. Our results demonstrate novel p53-independent drug-induced molecular reprogramming involving ERK1/2, CDK9, and ATF4 to restore upregulation of p53 effector genes required for cell death and tumor suppression.

HIF1α inhibition by dual targeting of CDK4/6 and HSP90 reduces cancer cell viability including Rb-deficient cells

Zhao, S. — 2020-10-20

Most cancers harbor intra-tumoral hypoxia which promotes tumor progression and therapy resistance. Hypoxia-inducible factor 1 (HIF1) mediates an adaptive response to hypoxia and contributes to multiple cancer hallmarks. We describe cancer therapeutic targeting of HIF1 by combination of CDK4/6 inhibitors (CDK4/6i) and heat-shock protein 90 inhibitors (HSP90i). CDK1 contributes to HSP90-mediated HIF1 stabilization whereas CDK1-knockdown enhances HIF1 reduction by HSP90i. Dual CDK1- and HSP90-inhibition increases apoptosis and synergistically inhibits cancer cell viability. To translate our findings, we use FDA-approved CDK4/6i in combination with HSP90i to reduce HIF1 expression and suppress viability of multiple cancer cell types, including Rb-deficient cancer cells. Overexpression of HIF1668E partially rescues the cell viability inhibition by combination CDK4/6i and HSP90i treatment under hypoxia. CDK4/6i and HSP90i suppresses tumor growth in vivo. Thus, combined targeting of CDK4/6 and HSP90, through a drug class effect, inhibits HIF1 and shows preclinical anti-cancer therapeutic efficacy, including with Rb-deficiency.

HiCRep.py: Fast comparison of Hi-C contact matrices in Python

Lin, D. — 2020-10-28

Hi-C is the most widely used assay for investigating genome-wide 3D organization of chromatin. When working with Hi-C data, it is often useful to calculate the similarity between contact matrices in order to asses experimental reproducibility or to quantify relationships among Hi-C data from related samples. The HiCRep algorithm has been widely adopted for this task, but the existing R implementation suffers from run time limitations on high resolution Hi-C data or on large single-cell Hi-C datasets. We introduce a Python implementation of HiCRep and demonstrate that it is much faster than the existing R implementation. Furthermore, we give examples of HiCReps ability to accurately distinguish replicates from non-replicates and to reveal cell type structure among collections of Hi-C data. HiCRep.py and its documentation are available with a GPL license at https://github.com/Noble-Lab/hicrep. The software may be installed automatically using the pip package installer.

Intolerance to uncertainty modulates neural synchrony between political partisans

van Baar, J. M. — 2020-10-29

Political partisans see the world through an ideologically biased lens. What drives political polarization? It has been posited that polarization arises because holding extreme political views satisfies a need for certain and stable beliefs about the world. We examined the relationship between uncertainty tolerance and political polarization using brain-to-brain synchrony analysis, which measured committed liberals and conservatives subjective interpretation of a continuous political narrative. Participants (N=44) watched a political debate while undergoing fMRI. Shared ideology between participants increased neural synchrony in many brain areas including key regions of the valuation and theory-of-mind networks (e.g. temporoparietal junction). The degree of neural synchrony was modulated by uncertainty aversion: Uncertainty-intolerant individuals experienced greater brain-to-brain synchrony with politically like-minded peers and lower synchrony with political opponents. This effect was observed for liberals and conservatives alike. Moreover, increasing neural synchrony between committed partisans predicted subsequent polarized attitude formation about the debate after the scanning session. These results suggest that uncertainty attitudes gate the shared neural processing of political narratives, thereby fueling polarized attitude formation about hot-button issues.

Infant functional connectivity fingerprints predict long-term language and pre-literacy outcomes

Yu, X. — 2020-10-29

Functional brain networks undergo extensive development within the first few years of life. Previous studies have linked infant functional connectivity to cognitive abilities in toddlerhood. However, little is known regarding the long-term relevance of functional connections established in infancy for the protracted development of higher-order abilities of language and literacy. Employing a five-year longitudinal imaging project starting in infancy, this study utilizes resting-state functional MRI to demonstrate prospective associations between infant functional connectivity fingerprints and subsequent language and foundational literacy skills at a mean age of 6.5. These longitudinal associations are preserved when key environmental influences are controlled for and are independent of emergent language abilities in infancy, suggesting early development of functional network characteristics in supporting the acquisition of high-order language and pre-literacy skills. Altogether, the current results highlight the importance of functional organization established in infancy as a neural scaffold underlying the learning process of complex cognitive functions.

Phase separation of the LINE-1 ORF1 protein is mediated by the N-terminus and coiled-coil domain

Jogl, G. — 2020-10-29

Long Interspersed Nuclear Element-1 (LINE-1 or L1) is a retrotransposable element that autonomously replicates in the human genome, resulting in DNA damage and genomic instability. Activation of L1 in senescent cells triggers a type I interferon response and age-associated inflammation. Two open reading frames encode an ORF1 protein functioning as mRNA chaperone and an ORF2 protein providing catalytic activities necessary for retrotransposition. No function has been identified for the conserved, disordered N-terminal region of ORF1. Using microscopy and NMR spectroscopy, we demonstrate that ORF1 forms liquid droplets in vitro in a salt-dependent manner and that interactions between its N-terminal region and coiled-coil domain are necessary for phase separation. Mutations disrupting blocks of charged residues within the N-terminus impair phase separation while some mutations within the coiled-coil domain enhance phase separation. Demixing of the L1 particle from the cytosol may provide a mechanism to protect the L1 transcript from degradation. Statement of significanceOver half of the human genome is comprised of repetitive sequences. The Long Interspersed Nuclear Element-1 (L1) is an autonomous mobile DNA element that can alter its genomic location, resulting in genomic instability and DNA damage. L1 encodes two proteins that are required for this function: the ORF1 RNA chaperone and the enzymatic ORF2. Here, we demonstrate that ORF1 forms liquid-liquid phase separated states in vitro, which is mediated by electrostatic interactions between the conserved, disordered N-terminus and coiled-coil domain. This work provides a framework to explore how L1 phase separation may enhance the ability of the retrotransposable element to colonize the genome by preventing degradation of the L1 transcript and evasion of host immune responses.

Reduced ech-6 Expression Attenuates Fat-induced Premature Aging in C. elegans

Liu, Y. J. — 2020-11-02

Deregulated energy homeostasis represents a hallmark of aging and results from complex gene-by-environment interactions. Here, we discovered that reducing the expression of the gene ech-6 encoding enoyl-CoA hydratase remitted fat diet-induced deleterious effects on lifespan in Caenorhabditis elegans, while a basal expression of ech-6 was important for survival under normal dietary conditions. Lipidomics revealed that supplementation of fat in ech-6-silenced worms had marginal effects on lipid profiles, suggesting an alternative fat utilization for energy production. Transcriptomics further suggest a causal relation between the lysosomal pathway, energy production, and the longevity effect conferred by the interaction between ech-6 and high-fat diets. Indeed, enhancing energy production from endogenous fat by overexpressing lysosomal lipase lipl-4 recapitulated the lifespan effects of high-fat diets on ech-6-silenced worms. Collectively, these results reveal that the gene ech-6 modulates metabolic flexibility and may be a target for promoting metabolic health and longevity.

The zinc finger protein CLAMP promotes long-range chromatin interactions that mediate dosage compensation of the Drosophila male X-chromosome

Jordan, W. — 2020-11-02

Drosophila dosage compensation is an important model system for defining how active chromatin domains are formed. The Male-specific lethal dosage compensation complex (MSLc) increases transcript levels of genes along the length of the single male X-chromosome to equalize with that on the two female X-chromosomes. The strongest binding sites for MSLc cluster together in three-dimensional space independent of MSLc because clustering occurs in both sexes. CLAMP, a non-sex specific, ubiquitous zinc finger protein, binds synergistically with MSLc to enrich the occupancy of both factors on the male X-chromosome. Here, we demonstrate that CLAMP promotes the observed clustering of MSLc bindings sites. Genome-wide, CLAMP promotes interactions between active chromatin regions. Moreover, the X-enriched CLAMP protein more strongly promotes longer-range interactions on the X-chromosome than autosomes. Genome-wide, CLAMP promotes interactions between active chromatin regions together with other insulator proteins. Overall, we define how long-range interactions which are modulated by a locally enriched ubiquitous transcription factor promote hyper-activation of the X-chromosome to mediate dosage compensation.

Cross-Platform Validation of Neurotransmitter Release Impairments in Schizophrenia Patient-Derived NRXN1-Mutant Neurons

Pak, C. — 2020-11-03

Heterozygous NRXN1 deletions constitute the most prevalent currently known single-gene mutation predisposing to schizophrenia. Previous studies showed that engineered heterozygous NRXN1 deletions impaired neurotransmitter release in human neurons, suggesting a synaptic pathophysiological mechanism. Utilizing this observation for drug discovery, however, requires confidence in its robustness and validity. Here, we describe a multi-center effort to test the generality of this pivotal observation, using independent analyses at two laboratories of patient-derived and newly engineered human neurons with heterozygous NRXN1 deletions. We show that in neurons that were trans-differentiated from induced pluripotent stem cells derived from three NRXN1-deletion patients, the same impairment in neurotransmitter release was observed as in engineered NRXN1-deficient neurons. This impairment manifested as a decrease in spontaneous synaptic events and in evoked synaptic responses, and an alteration in synaptic paired-pulse depression. Nrxn1-deficient mouse neurons generated from embryonic stem cells by the same method as human neurons did not exhibit impaired neurotransmitter release, suggesting a human-specific phenotype. NRXN1 deletions produced a reproducible increase in the levels of CASK, an intracellular NRXN1-binding protein, and were associated with characteristic gene expression changes. Thus, heterozygous NRXN1 deletions robustly impair synaptic function in human neurons regardless of genetic background, enabling future drug discovery efforts.

The Trypanosoma brucei subpellicular microtubule array is organized into functionally discrete subdomains defined by microtubule associated proteins

Sinclair, A. N. — 2020-11-09

Microtubules are inherently dynamic cytoskeletal polymers whose length and activity can be altered to perform essential functions in eukaryotic cells, such as providing tracks for intracellular trafficking and forming the mitotic spindle. Microtubules can be bundled to create more stable structures that collectively propagate force, such as in the flagellar axoneme, which provides motility. The subpellicular microtubule array of the protist parasite Trypanosoma brucei, the causative agent of African sleeping sickness, is a remarkable example of a highly specialized microtubule bundle, comprising a single microtubule layer that is crosslinked to each other and the plasma membrane. The array microtubules appear to be highly stable and remain intact throughout the cell cycle, but very little is known about the pathways that tune microtubule properties in trypanosomatids. Here, we show that the subpellicular microtubule array is organized into subdomains that consist of differentially localized array-associated proteins. We characterize the localization and function of the array-associated protein PAVE1, which is a component of the inter-microtubule crosslinking fibrils present within the posterior subdomain. PAVE1 functions to stabilize these microtubules to produce the tapered cell posterior. PAVE1 and the newly identified PAVE2 form a complex that binds directly to the microtubule lattice. TbAIR9, which localizes to the entirety of the subpellicular array, is necessary for retaining PAVE1 within the posterior subdomain, and also maintains array-associated proteins in the middle and anterior subdomains of the array. The arrangement of proteins within the array is likely to tune the local properties of the array microtubules and create the asymmetric shape of the cell, which is essential for parasite viability. Author summaryMany parasitic protists use arrays of microtubules that contact the inner leaflet of the plasma membrane, typically known as subpellicular microtubules, to shape their cells into forms that allow them to efficiently infect their hosts. While subpellicular arrays are found in a wide range of parasites, very little is known about how they are assembled and maintained. Trypanosoma brucei, which is the causative agent of human African trypanosomiasis, has an elaborate subpellicular array that produces the helical shape of the parasite, which is essential for its ability to move within crowded and viscous solutions. We have identified a series of proteins that have a range of localization patterns within the array, which suggests that the array is regulated by subdomains of array-associated proteins that may tune the local properties of the microtubules to suit the stresses found at different parts of the cell body. Among these proteins are the first known components of the inter-microtubule crosslinks that are thought to stabilize array microtubules, as well as a potential regulator of the array subdomains. These results establish a foundation to understand how subpellicular arrays are built, shaped, and maintained, which has not previously been appreciated.

Distributed Representation of Taste Quality by Second-Order Gustatory Neurons in Drosophila

Snell, N. J. — 2020-11-11

Sweet and bitter compounds excite different sensory cells and drive opposing behaviors. It is commonly thought that the neural circuits linking taste sensation to behavior conform to a labeled-line architecture, but in Drosophila, evidence for labeled lines beyond first-order neurons is lacking. To address this, we devised trans-Tango(activity), a strategy for calcium imaging of second-order gustatory projection neurons based on trans-Tango, a genetic transsynaptic tracing technique. We found distinct projection neuron populations that respond to sweet and bitter tastants. However, the bitter-responsive population was also activated by water alone. We further discovered that bitter tastants evoke activity upon both stimulus onset and offset. Bitter offset responses are exhibited by both first- and second-order gustatory neurons, but these responses are distributed among multiple types of projection neurons in the second order. These findings suggest a more complex coding scheme for gustatory information than can be explained by a labeled line model.

Likelihood Approximation Networks (LANs) for Fast Inference of Simulation Models in Cognitive Neuroscience

Fengler, A. — 2020-11-22

In cognitive neuroscience, computational modeling can formally adjudicate between theories and affords quantitative fits to behavioral/brain data. Pragmatically, however, the space of plausible generative models considered is dramatically limited by the set of models with known likelihood functions. For many models, the lack of a closed-form likelihood typically impedes Bayesian inference methods. As a result, standard models are evaluated for convenience, even when other models might be superior. Likelihood-free methods exist but are limited by their computational cost or their restriction to particular inference scenarios. Here, we propose neural networks that learn approximate likelihoods for arbitrary generative models, allowing fast posterior sampling with only a one-off cost for model simulations that is amortized for future inference. We show that these methods can accurately recover posterior parameter distributions for a variety of neurocognitive process models. We provide code allowing users to deploy these methods for arbitrary hierarchical model instantiations without further training.

Improve Concentration Of Frequency And Time (ConceFT) By Novel Complex Spherical Designs

Sourisseau, M. — 2020-11-23

AO_SCPLOWBSTRACTC_SCPLOWConcentration of frequency and time (ConceFT) is a generalized multitaper algorithm introduced to analyze complicated non-stationary time series. To avoid the randomness in the original ConceFT algorithm, we apply the novel complex spherical design technique to standardize ConceFT, which we coin CQU-ConceFT. The proposed CQU-ConceFT is applied to visualize the spindle structure in the electroencephalogram signal during the N2 sleep stage and other physiological time series.

Integrating long-range regulatory interactions to predict gene expression using graph convolutional neural networks

Bigness, J. — 2020-11-24

Long-range spatial interactions among genomic regions are critical for regulating gene expression, and their disruption has been associated with a host of diseases. However, when modeling the effects of regulatory factors, most deep learning models either neglect long-range interactions or fail to capture the inherent 3D structure of the underlying genomic organization. To address these limitations, we present GC-MERGE, a Graph Convolutional Model for Epigenetic Regulation of Gene Expression. Using a graph-based framework, the model incorporates important information about long-range interactions via a natural encoding of spatial interactions into the graph representation. It integrates measurements of both the spatial genomic organization and local regulatory factors, specifically histone modifications, to not only predict the expression of a given gene of interest but also quantify the importance of its regulatory factors. We apply GC-MERGE to datasets for three cell lines - GM12878 (lymphoblastoid), K562 (myelogenous leukemia), and HUVEC (human umbilical vein endothelial) - and demonstrate its state-of-the-art predictive performance. Crucially, we show that our model is interpretable in terms of the observed biological regulatory factors, high-lighting both the histone modifications and the interacting genomic regions contributing to a genes predicted expression. We provide model explanations for multiple exemplar genes and validate them with evidence from the literature. Our model presents a novel setup for predicting gene expression by integrating multimodal datasets in a graph convolutional framework. More importantly, it enables interpretation of the biological mechanisms driving the models predictions. Available at: https://github.com/rsinghlab/GC-MERGE.

A pipeline to characterize local cortical folds by mapping them to human-interpretable shapes

Roy, A. — 2020-11-26

BackgroundVariations in regional cortical folds across individuals have been examined using computationally-derived morphological measures, or by manual characterization procedures that map distinct variants of a regional fold to a set of human-interpretable shapes. Although manual mapping approaches have proven useful for identifying morphological differences of clinical relevance, such procedures are subjective and not amenable to scaling. New MethodWe propose a 3-step pipeline to develop computational models of manual mapping. The steps are: represent regional folds as feature vectors, manually map each feature vector to a shape-variant that the underlying fold represents, and train classifiers to learn the mapping. ResultsFor demonstration, we chose a 2D-problem of detecting within slice discontinuity of medial and lateral sulci of orbitofrontal cortex (OFC); the discontinuity may be visualized as a broken H-shaped pattern, and is fundamental to OFC-type-characterization. The classifiers predicted discontinuities with 86-95% test-accuracy. Comparison with Existing MethodsThere is no existing pipeline that automates a manual characterization process. For the current demonstration problem, we conduct multiple analyses using existing softwares to explain our design decisions, and present guidelines for using the pipeline to examine other regional folds using conventional or non-conventional morphometric measures. ConclusionWe show that this pipeline can be useful for determining axial-slice discontinuity of sulci in the OFC and can learn structural-features that human-raters may rely on during manual-characterization.The pipeline can be used for examining other regional folds and may facilitate discovery of various statistically-reliable 2D or 3D human-interpretable shapes that are embedded throughout the brain.

Visual perceptual learning of a primitive feature in human V1/V2 as a result of unconscious processing, revealed by Decoded fMRI Neurofeedback (DecNef)

Wang, Z. — 2020-12-02

While numerous studies have shown that visual perceptual learning (VPL) occurs as a result of exposure to a visual feature in a task-irrelevant manner, the underlying neural mechanism is poorly understood. In a previous psychophysical study, subjects were repeatedly exposed to a task-irrelevant global motion display that induced the perception of not only the local motions but also a global motion moving in the direction of the spatiotemporal average of the local motion vectors. As a result, subjects enhanced their sensitivity only to the local moving directions, suggesting that early visual areas (V1/V2) that process local motions are involved in task-irrelevant VPL. However, this hypothesis has never been examined by directly examining the involvement of early visual areas (V1/V2). Here, we employed a decoded neurofeedback technique (DecNef) using functional magnetic resonance imaging. During the DecNef training, subjects were trained to induce the activity patterns in V1/V2 that were similar to those evoked by the actual presentation of the global motion display. The DecNef training was conducted with neither the actual presentation of the display nor the subjects awareness of the purpose of the experiment. As a result, subjects increased the sensitivity to the local motion directions but not specifically to the global motion direction. The training effect was strictly confined to V1/V2. Moreover, subjects reported that they neither perceived nor imagined any motion during the DecNef training. These results together suggest that that V1/V2 are sufficient for exposure-based task-irrelevant VPL to occur unconsciously. Significance StatementWhile numerous studies have shown that visual perceptual learning (VPL) occurs as a result of exposure to a visual feature in a task-irrelevant manner, the underlying neural mechanism is poorly understood. Previous psychophysical experiments suggest that early visual areas (V1/V2) are involved in task-irrelevant VPL. However, this hypothesis has never been examined by directly examining the involvement of early visual areas (V1/V2). Here, using decoded fMRI neurofeedback, the activity patterns similar to those evoked by the presentation of a complex motion display were repeatedly induced only in early visual areas. The training sensitized only the local motion directions and not the global motion direction, suggesting that V1/V2 are involved in task-irrelevant VPL.

Molecular characterization of projection neuron subtypes in the mouse olfactory bulb

Zeppilli, S. — 2020-12-02

Projection neurons (PNs) in the mammalian olfactory bulb (OB) receive direct input from the nose and project to diverse cortical and subcortical areas. Morphological and physiological studies have highlighted functional heterogeneity, yet no molecular markers have been described that delineate PN subtypes. Here, we used viral injections into olfactory cortex and fluorescent nucleus sorting to enrich PNs for high-throughput single nucleus and bulk RNA deep sequencing. Transcriptome analysis and RNA in situ hybridization identified three mitral and five tufted cell populations with characteristic transcription factor network topology and cell adhesion and excitability-related gene expression. Finally, by integrating bulk and snRNA-seq data we propose that different mitral cell populations selectively project to different regions of olfactory cortex. Together, we have identified potential molecular and gene regulatory mechanisms underlying PN diversity and provide new molecular entry points into studying the diverse functional roles of mitral and tufted cell subtypes.

The Genetic Architecture of Robustness for Flight Performance in Drosophila

Spierer, A. N. — 2020-12-06

A central challenge of quantitative genetics is partitioning phenotypic variation into genetic and non-genetic components. These non-genetic components are usually interpreted as environmental effects; however, variation between genetically identical individuals in a common environment can still exhibit phenotypic variation. A traits resistance to variation is called robustness, though the genetics underlying it are poorly understood. Accordingly, we performed an association study on a previously studied, whole organism trait: flight performance. Using 197 of the Drosophila Genetic Reference Panel (DGRP) lines, we surveyed variation at the level of single nucleotide polymorphisms and whole genes using additive, marginal, and epistatic analyses that associated with robustness for flight performance. Many genes had developmental and neurodevelopmental annotations, and many more were identified from associations that differed between sexes. Additionally, many genes were pleiotropic, with several annotated for fitness-associated traits (e.g. gametogenesis and courtship). Our results corroborate a previous study for genetic modifiers of micro-environmental variation, and have sizable overlap with studies for modifiers of wing morphology and courtship behavior. These results point to an important and shared role for genetic modifiers of robustness of flight performance affecting development, neurodevelopment, and behavior.

Hippocampal cells multiplex positive and negative engrams

Shpokayte, M. — 2020-12-11

The hippocampus is involved in processing a variety of mnemonic computations specifically the spatiotemporal components and emotional dimensions of contextual memory.1-3 Recent studies have demonstrated vast structural and functional heterogeneity along the dorsal-ventral axis1, 5 of the hippocampus. The ventral hippocampus has been shown to be important in the processing of emotion and valence.6-9 Here, we combine transgenic and all-virus based activity-dependent tagging strategies to visualize multiple valence-specific engrams in the vHPC and demonstrate two partially segregated cell populations and projections that respond to appetitive and aversive experiences. Next, using RNA sequencing and DNA methylation sequencing approaches, we find that vHPC appetitive and aversive engram cells display distinct transcriptional programs and DNA methylation landscapes compared to a neutral engram population. Additionally, while optogenetic manipulation of tagged cell bodies in vHPC is not sufficient to drive appetitive or aversive behavior in real-time place preference, stimulation of tagged vHPC terminals projecting to the amygdala and nucleus accumbens (NAc), but not the prefrontal cortex (PFC), had the capacity drive preference and avoidance. These terminals can also undergo a "switch" or "reset" in their capacity to drive either, thereby demonstrating their adaptable contributions to behavior. We conclude that the vHPC contains genetically, cellularly, and behaviorally distinct populations of cells processing appetitive and aversive memory engrams. Together, our findings provide a novel means by which to visualize multiple engrams within the same brain and point to their unique genetic signatures as reference maps for the future development of new therapeutic strategies. One sentence summaryThe hippocampus contains neurons that correspond to positive and negative engrams, which are segregated by their molecular, cellular, and projection-specific features.

Distinct expression of select and transcriptome-wide isolated 3'UTRs suggests critical roles in development and transition states

Hynes, M. — 2020-12-11

Mature mRNA molecules are typically considered to be comprised of a 5UTR, a 3UTR and a coding region (CDS), all attached until degradation. Unexpectedly, however, there have been multiple recent reports of widespread differential expression of mRNA 3UTRs and their cognate coding regions, resulting in the expression of isolated 3UTRs (i3UTRs); these i3UTRs can be highly expressed, often in reciprocal patterns to their cognate CDS. Similar to the role of other lncRNAs, isolated 3UTRs are likely to play an important role in gene regulation but little is known about the contexts in which they are deployed. To begin to parse the functions of i3UTRs, here we carry out in vitro, in vivo and in silico analyses of differential 3UTR/CDS mRNA ratio usage across tissues, development and cell state changes both for a select list of developmentally important genes as well as through unbiased transcriptome-wide analyses. Across two developmental paradigms we find a distinct switch from high i3UTR expression of stem cell related genes in proliferating cells compared to newly differentiated cells. Our unbiased transcriptome analysis across multiple gene sets shows that regardless of tissue, genes with high 3UTR to CDS ratios belong predominantly to gene ontology categories related to cell-type specific functions while in contrast, the gene ontology categories of genes with low 3UTR to CDS ratios are similar and relate to common cellular functions. In addition to these specific findings our data provide critical information from which detailed hypotheses for individual i3UTRs can be tested-with a common theme that i3UTRs appear poised to regulate cell-specific gene expression and state. Significance StatementThe widespread existence and expression of mRNA 3 untranslated sequences in the absence of their cognate coding regions (called isolated 3UTRs or i3UTRs) opens up considerable avenues for gene regulation not previously envisioned. Each isolated 3UTR may still bind and interact with micro RNAs, RNA binding proteins as well as other nucleic acid sequences, all in the absence or low levels of cognate protein production. Here we document the expression, localization and regulation of i3UTRs both within particular biological systems as well as across the transcriptome. As this is an entirely new area of experimental investigation these early studies are seminal to this burgeoning field.

Analogous computations in working memory input, output and motor gating: Electrophysiological and computational modeling evidence

Rac-Lubashevsky, R. — 2020-12-22

Adaptive cognitive-control involves a hierarchical cortico-striatal gating system that supports selective updating, maintenance, and retrieval of useful cognitive and motor information. Here, we developed a task that independently manipulates selective gating operations into working-memory (input gating), from working-memory (output gating), and of responses (motor gating) and tested the neural dynamics and computational principles that support them. Increases in gating demands, captured by gate switches, were expressed by distinct EEG correlates at each gating level that evolved dynamically in partially overlapping time windows. Further, categorical representations of specific maintained items and of motor responses could be decoded from EEG when the corresponding gate was switching, thereby linking gating operations to prioritization. Finally, gate switching at all levels was related to increases in the motor decision threshold as quantified by the drift diffusion model. Together these results support the notion that cognitive gating operations scaffold on top of mechanisms involved in motor gating. Author summaryHow do humans decide which information is relevant to attend to in memory, which cognitive operation to take, and when? Flexibly updating, maintenance and retrieval of relevant information from working memory (WM) are thought to be managed by gating computations in the frontostriatal network, supporting higher order learning and cognitive flexibility. Using the reference-back-2 task, we tested the key properties of gating. Namely that they are selective ("content-addressable") and that principles of cognitive "actions" (including input gating of WM, output gating from WM) are scaffold on top of the motor gating operations. Using trial-by-trial EEG indexing and quantitative computational modeling (the hierarchical drift-diffusion model) we showed that action selection at all three levels of gating have separable neural signatures but they operate partly in parallel, such that decisions about a response are processed to some degree even while the identity of the cognitive rule were uncertain. Furthermore, we showed analogous computations across levels of gating as selection of WM representation and of motor action lead to increase in the estimated decision threshold and to enhanced neural coding of the selected information thereby providing a novel link between WM gating and WM prioritization.

Rare coding variants in 35 genes associate with circulating lipid levels: a multi-ancestry analysis of 170,000 exomes

Hindy, G. — 2020-12-23

Large-scale gene sequencing studies for complex traits have the potential to identify causal genes with therapeutic implications. We performed gene-based association testing of blood lipid levels with rare (minor allele frequency<1%) predicted damaging coding variation using sequence data from >170,000 individuals from multiple ancestries: 97,493 European, 30,025 South Asian, 16,507 African, 16,440 Hispanic/Latino, 10,420 East Asian, and 1,182 Samoan. We identified 35 genes associated with circulating lipid levels. Ten of these: ALB, SRSF2, JAK2, CREB3L3, TMEM136, VARS, NR1H3, PLA2G12A, PPARG and STAB1 have not been implicated for lipid levels using rare coding variation in population-based samples. We prioritize 32 genes identified in array-based genome-wide association study (GWAS) loci based on gene-based associations, of which three: EVI5, SH2B3, and PLIN1, had no prior evidence of rare coding variant associations. Most of the associated genes showed evidence of association in multiple ancestries. Also, we observed an enrichment of gene-based associations for low-density lipoprotein cholesterol drug target genes, and for genes closest to GWAS index single nucleotide polymorphisms (SNP). Our results demonstrate that gene-based associations can be beneficial for drug target development and provide evidence that the gene closest to the array-based GWAS index SNP is often the functional gene for blood lipid levels.

Narratives: fMRI data for evaluating models of naturalistic language comprehension

Nastase, S. A. — 2020-12-23

The "Narratives" collection aggregates a variety of functional MRI datasets collected while human subjects listened to naturalistic spoken stories. The current release includes 345 subjects, 891 functional scans, and 27 diverse stories of varying duration totaling ~4.6 hours of unique stimuli (~43,000 words). This data collection is well-suited for naturalistic neuroimaging analysis, and is intended to serve as a benchmark for models of language and narrative comprehension. We provide standardized MRI data accompanied by rich metadata, preprocessed versions of the data ready for immediate use, and the spoken story stimuli with time-stamped phoneme- and word-level transcripts. All code and data are publicly available with full provenance in keeping with current best practices in transparent and reproducible neuroimaging.

A comprehensive computational model of animal biosonar signal processing

Ming, C. — 2020-12-29

Computational models of animal biosonar seek to identify critical aspects of echo processing responsible for the superior, real-time performance of echolocating bats and dolphins in target tracking and clutter rejection. The Spectrogram Correlation and Transformation (SCAT) model replicates aspects of biosonar imaging in both species by processing wideband biosonar sounds and echoes with auditory mechanisms identified from experiments with bats. The model acquires broadband biosonar broadcasts and echoes, represents them as time-frequency spectrograms using parallel bandpass filters, translates the filtered signals into ten parallel amplitude threshold levels, and then operates on the resulting time-of-occurrence values at each frequency to estimate overall echo range delay. It uses the structure of the echo spectrum by depicting it as a series of local frequency nulls arranged regularly along the frequency axis of the spectrograms after dechirping them relative to the broadcast. Computations take place entirely on the timing of threshold-crossing events for each echo relative to threshold-events for the broadcast. Threshold-crossing times take into account amplitude-latency trading, a physiological feature absent from conventional digital signal processing. Amplitude-latency trading transposes the profile of amplitudes across frequencies into a profile of time-registrations across frequencies. Target shape is extracted from the spacing of the objects individual acoustic reflecting points, or glints, using the mutual interference pattern of peaks and nulls in the echo spectrum. These are merged with the overall range-delay estimate to produce a delay-based reconstruction of the objects distance as well as its glints. Clutter echoes indiscriminately activate multiple parts in the null-detecting system, which then produces the equivalent glint-delay spacings in images, thus blurring the overall echo-delay estimates by adding spurious glint delays to the image. Blurring acts as an anticorrelation process that rejects clutter intrusion into perceptions. Author summaryBats and dolphins use their biological sonar as a versatile, high-resolution perceptual system that performs at levels desirable in man-made sonar or radar systems. To capture the superior real-time capabilities of biosonar so they can be imported into the design of new man-made systems, we developed a computer model of the sonar receiver used by echolocating bats and dolphins. Our intention was to discover the processing methods responsible for the animals ability to find and identify targets, guide locomotion, and prevent classic types of sonar or radar interference that hamper performance of man-made systems in complex, rapidly-changing surroundings. We have identified several features of the ears, hearing, time-frequency representation, and auditory processing that are critical for organizing echo-processing methods and display manifested in the animals perceptions.

Neuromolecular and behavioral effects of ethanol deprivation in Drosophila

D'Silva, N. M. — 2021-01-04

Alcohol use disorder (AUD) is characterized by loss of control in limiting alcohol intake. This may involve intermittent periods of abstinence followed by alcohol seeking and, consequently, relapse. However, little is understood of the molecular mechanisms underlying the impact of alcohol deprivation on behavior. Using a new Drosophila melanogaster repeated intermittent alcohol exposure model, we sought to identify how ethanol deprivation alters spontaneous behavior, determine the associated neural structures, and reveal correlated changes in brain gene expression. We found that repeated intermittent ethanol-odor exposures followed by ethanol-deprivation dynamically induces behaviors associated with a negative affect state. Although behavioral states broadly mapped to many brain regions, persistent changes in social behaviors mapped to the mushroom body and surrounding neuropil. This occurred concurrently with changes in expression of genes associated with sensory responses, neural plasticity, and immunity. Like social behaviors, immune response genes were upregulated following three-day repeated intermittent ethanol-odor exposures and persisted with one or two days of ethanol-deprivation, suggesting an enduring change in molecular function. Our study provides a framework for identifying how ethanol deprivation alters behavior with correlated underlying circuit and molecular changes.

Chitinase 3-like-1 is a Therapeutic Target That Mediates the Effects of Aging in COVID-19

Kamle, S. — 2021-01-06

COVID-19 is caused by the SARS-CoV-2 (SC2) virus and is more prevalent and severe in the elderly and patients with comorbid diseases (CM). Because chitinase 3-like-1 (CHI3L1) is induced during aging and CM, the relationships between CHI3L1 and SC2 were investigated. Here we demonstrate that CHI3L1 is a potent stimulator of the SC2 receptor ACE2 and viral spike protein priming proteases (SPP), that ACE2 and SPP are induced during aging and that anti-CHI3L1, kasugamycin and inhibitors of phosphorylation, abrogate these ACE2- and SPP-inductive events. Human studies also demonstrated that the levels of circulating CHI3L1 are increased in the elderly and patients with CM where they correlate with COVID-19 severity. These studies demonstrate that CHI3L1 is a potent stimulator of ACE2 and SPP; that this induction is a major mechanism contributing to the effects of aging during SC2 infection and that CHI3L1 coopts the CHI3L1 axis to augment SC2 infection. CHI3L1 plays a critical role in the pathogenesis of and is an attractive therapeutic target in COVID-19.

Generation of Pathogenic TPP1 Mutations in Human Stem Cells as a Model for CLN2 Disease

Ma, L. — 2021-01-06

Neuronal ceroid lipofuscinosis type 2 (CLN2 disease) is an autosomal recessive neurodegenerative disorder generally with onset at 2 to 4 years of age and characterized by seizures, loss of vision, progressive motor and mental decline, and premature death. CLN2 disease is caused by loss-of-function mutations in the tripeptidyl peptidase 1 (TPP1) gene leading to deficiency in TPP1 enzyme activity. Approximately 60% of patients have one of two pathogenic variants (c.509-1G>C or c.622C>T [p.(Arg208*)]). In order to generate a human stem cell model of CLN2 disease, we used CRISPR/Cas9-mediated knock-in technology to introduce these mutations in a homozygous state into H9 human embryonic stem cells. Heterozygous lines of the c.622C>T (p.(Arg208*)) mutation were also generated, which included a heterozygous mutant with a wild-type allele and different compound heterozygous coding mutants resulting from indels on one allele. We describe the methodology that led to the generation of the lines and provide data on the initial validation and characterization of these CLN2 disease models. Notably, both mutant lines (c.509-1G>C and c.622C>T [p.(Arg208*)]) in the homozygous state were shown to have reduced or absent protein, respectively, and deficiency of TPP1 enzyme activity. These models, which we are making available for wide-spread sharing, will be useful for future studies of molecular and cellular mechanisms underlying CLN2 disease and for therapeutic development.

Heparin-modified alginate microspheres enhance neovessel formation in hiPSC-derived endothelial cells and heterocellular in vitro models by controlled release of VEGF

Munarin, F. — 2021-01-09

A formidable challenge in regenerative medicine is the development of stable microvascular networks to restore adequate blood flow or to sustain graft viability and long-term function in implanted or ischemic tissues. In this work, we develop a biomimetic approach to increase the binding affinity of the extracellular matrix for the class of heparin-binding growth factors to localize and control the release of proangiogenic cues while maintaining their bioactivity. Sulfate and heparin moieties are covalently coupled to alginate, and alginate microspheres are produced and used as local delivery depots for vascular endothelial growth factor (VEGF). Release of VEGF from sulfate-alginate and heparin-alginate bulk hydrogels and microspheres was sustained over 14 days. In vitro evaluation with human induced pluripotent stem cell (hiPSC)-derived endothelial cells and aortic ring assay in a chemically-defined hydrogel demonstrates development of primitive three-dimensional vessel-like networks in the presence of VEGF released from the chemically modified alginate microspheres. Furthermore, our results suggest that the sulfate groups available on the chemically modified alginate microspheres promote some new vessel formation even in VEGF-free samples. Based on this evidence, we conclude that sulfate- and heparin-alginate hydrogels are adaptive and bioactive delivery systems for revascularization therapy and translational vascular tissue engineering.

Phospho-dependent Signaling during the General Stress Response by the Atypical Response Regulator and ClpXP Adaptor RssB

Schwartz, J. — 2021-01-11

In the model organism Escherichia coli and related species, the general stress response relies on tight regulation of the intracellular levels of the promoter specificity subunit RpoS. RpoS turnover is exclusively dependent on RssB, a two-domain response regulator that functions as an adaptor that delivers RpoS to ClpXP for proteolysis. Here we report crystal structures of the receiver domain of RssB both in its unphosphorylated form and bound to the phosphomimic BeF3-. Surprisingly, we find only modest differences between these two structures, suggesting that truncating RssB may partially activate the receiver domain to a "meta-active" state. Our structural and sequence analysis points to RssB proteins not conforming to either the Y-T coupling scheme for signaling seen in prototypical response regulators, such as CheY, or to the signaling model of the less understood FATGUY proteins.

An enhancer sequence in the intrinsically disordered region of the essential cell division protein FtsZ promotes conformation-guided substrate processing by ClpXP in Escherichia coli

Viola, M. G. — 2021-01-14

The essential bacterial division protein in Escherichia coli, FtsZ, assembles into the FtsZ-ring at midcell and recruits other proteins to the division site to promote septation. A region of the FtsZ amino acid sequence that links the conserved polymerization domain to a C-terminal protein interaction site was predicted to be intrinsically disordered and has been implicated in modulating spacing and architectural arrangements of FtsZ filaments. While the majority of cell division proteins that directly bind to FtsZ engage either the polymerization domain or the C-terminal interaction site, ClpX, the recognition and unfolding component of the bacterial ClpXP proteasome, has a secondary interaction with the predicted intrinsically disordered region (IDR) of FtsZ when FtsZ is polymerized. Here, we use NMR spectroscopy and reconstituted degradation reactions in vitro to demonstrate that this linker region is indeed disordered in solution and, further, that amino acids in the IDR of FtsZ enhance the degradation by conformationally-guided interactions.

Chitinase 3-like-1 Stimulates PD-L1 and Other Immune Checkpoint Inhibitors

Ma, B. — 2021-01-17

PD-1 and its ligand PD-L1 are major mediators of tumor-induced immunosuppression. Chitinase 3-like-1 (Chi3l1) is induced in many cancers where it portends a poor prognosis and contributes to tumor metastasis. Here we demonstrate that Chi3l1 regulates the expression of PD-L1, PD-L2, PD-1 and LAG3 in melanoma lung metastasis. Chi3l1 stimulates macrophage PD-L1 expression and mediates optimal IFN-{gamma}-stimulated PD-L1 expression via IL-13R2. We also demonstrate that RIG-like helicase innate immune activation suppresses Chi3l1, PD-L1, LAG3 and pulmonary metastasis. At least additive antitumor responses were seen in metastasis models treated simultaneously with individual antibodies against PD-1 and Chi3l1. At least additive cytotoxic T cell-induced tumor cell death was also seen in co-cultures of T and tumor cells treated with antibodies that target Chi3l1 and PD-1. Thus, Chi3l1 contributes to pulmonary metastasis by stimulating the PD1-PD-L1 axis and other checkpoint molecules. The simultaneous targeting of Chi3l1 and the PD-1-PD-L1 axis, represents a promising therapeutic strategy for pulmonary metastasis.

Spatiotemporal dynamics of maximal and minimal EEG spectral power

Menceloglu, M. — 2021-01-17

Oscillatory neural activities are prevalent in the brain with their phase realignment contributing to the coordination of neural communication. Phase realignments may have especially strong (or weak) impact when neural activities are strongly synchronized (or desynchronized) within the interacting populations. We report that the spatiotemporal dynamics of strong regional synchronization measured as maximal EEG spectral power--referred to as activation--and strong regional desynchronization measured as minimal EEG spectral power--referred to as suppression--are characterized by the spatial segregation of small-scale and large-scale networks. Specifically, small-scale spectral-power activations and suppressions involving only 2-7% (1-4 of 60) of EEG scalp sites were prolonged (relative to stochastic dynamics) and consistently co-localized in a frequency specific manner. For example, the small-scale networks for{theta} , , {beta}1, and {beta}2 bands (4-30 Hz) consistently included frontal sites when the eyes were closed, whereas the small-scale network for{gamma} band (31-55 Hz) consistently clustered in medial-central-posterior sites whether the eyes were open or closed. Large-scale activations and suppressions involving over 17-30% (10-18 of 60) of EEG sites were also prolonged and generally clustered in regions complementary to where small-scale activations and suppressions clustered. In contrast, intermediate-scale activations and suppressions (involving 7-17% of EEG sites) tended to follow stochastic dynamics and were less consistently localized. These results suggest that strong synchronizations and desynchronizations tend to occur in small-scale and large-scale networks that are spatially segregated and frequency specific. These synchronization networks may broadly segregate the relatively independent and highly cooperative oscillatory processes while phase realignments fine-tune the network configurations based on behavioral demands.

Somatic Mutations in Collagens are Associated with a Distinct Tumor Environment and Overall Survival in Gastric Cancer

Brodsky, A. S. — 2021-01-19

BackgroundGastric cancer is a heterogeneous disease with poorly understood genetic and microenvironmental factors. Mutations in collagen genes are associated with genetic diseases that compromise tissue integrity, but their role in tumor progression has not been extensively reported. Aberrant collagen expression has been long associated with malignant tumor growth, invasion, chemoresistance, and patient outcomes. We hypothesized that somatic mutations in collagens could functionally alter the tumor extracellular matrix. MethodsWe used publicly available datasets including The Tumor Cancer Genome Atlas (TCGA) to interrogate somatic mutations in collagens in stomach adenocarcinomas. To demonstrate that collagens were significantly mutated above background mutation rates, we used a moderated Kolmogorov-Smirnov test along with combination analysis with a bootstrap approach to define the background accounting for mutation rates. Association between mutations and clinicopathological features was evaluated by Fisher or chi-squared tests. Association with overall survival was assessed by Kaplan-Meier and the Cox-Proportional Hazards Model. Gene Set Enrichment Analysis was used to interrogate pathways. Immunohistochemistry and in situ hybridization tested expression of COL7A1 in stomach tumors. ResultsIn stomach adenocarcinomas, we identified individual collagen genes and sets of collagen genes harboring somatic mutations at a high frequency compared to background in both microsatellite stable, and microsatellite instable tumors in TCGA. Many of the missense mutations resemble the same types of loss of function mutations in collagenopathies that disrupt tissue formation and destabilize cells providing guidance to interpret the somatic mutations. We identified combinations of somatic mutations in collagens associated with overall survival, with a distinctive tumor microenvironment marked by lower matrisome expression and immune cell signatures. Truncation mutations were strongly associated with improved outcomes suggesting that loss of expression of secreted collagens impact tumor progression and treatment response. Germline collagenopathy variants guided interpretation of impactful somatic mutations on tumors. ConclusionsThese observations highlight that many collagens, expressed in non-physiologically relevant conditions in tumors, harbor impactful somatic mutations in tumors, suggesting new approaches for classification and therapy development in stomach cancer. In sum, these findings demonstrate how classification of tumors by collagen mutations identified strong links between specific genotypes and the tumor environment. Research HighlightsCollagen mutations are prevalent in stomach cancer Collagen somatic missense mutations resemble collagenopathy mutations Collagen mutations associate with overall survival in stomach cancer Tumors with collagen mutations have distinct molecular pathways and tumor microenvironments

LINE-1 Retrotransposon expression in cancerous, epithelial and neuronal cells revealed by 5'-single cell RNA-Seq

McKerrow, W. — 2021-01-20

LINE-1 retrotransposons are sequences capable of copying themselves to new genomic loci via an RNA intermediate. New studies implicate LINE-1 in a range of diseases, especially in the context of aging, but without an accurate understanding of where and when LINE-1 is expressed, a full accounting of its role in health and disease is not possible. We therefore developed a method - 5 scL1seq - that makes use of a widely available library preparation method (10x Genomics 5 single cell RNA-seq) to measure LINE-1 expression in tens of thousands of single cells. We recapitulated the known pattern of LINE-1 expression in tumors - present in cancer cells, absent from immune cells - and identified hitherto undescribed LINE-1 expression in human epithelial cells and mouse hippocampal neurons. In both cases, we saw a modest increase with age, supporting recent research connecting LINE-1 to age related diseases.

Fiber photometry does not reflect spiking activity in the striatum

Legaria, A. A. — 2021-01-21

Calcium recording via fiber photometry is commonly used as a proxy for recording population neuronal activity in vivo, yet the biological source of the photometry signal remains unclear. Here, using simultaneous in vivo extracellular electrophysiology and fiber photometry in the striatum, along with endoscopic 1-photon and 2-photon calcium imaging, we determined that the striatal fiber photometry signal reflects primarily non-somatic, and not somatic, changes in calcium.

Succinate Dehydrogenase (SDH)-subunit C regulates muscle oxygen consumption and fatigability in an animal model of pulmonary emphysema.

Balnis, J. — 2021-01-22

Patients with pulmonary emphysema often develop locomotor muscle dysfunction, which is independently associated with disability and higher mortality in that population. Muscle dysfunction entails reduced force-generation capacity which partially depends on fibers oxidative potential, yet very little mechanistic research has focused on muscle respiration in pulmonary emphysema. Using a recently established animal model of pulmonary emphysema-driven skeletal muscle dysfunction, we found downregulation of succinate dehydrogenase (SDH) subunit C in association with lower oxygen consumption and fatigue-tolerance in locomotor muscles. Reduced SDH activity has been previously observed in muscles from patients with pulmonary emphysema and we found that SDHC is required to support respiration in cultured muscle cells. Moreover, in-vivo gain of SDH function in emphysema animals muscles resulted in better oxygen consumption rate (OCR) and fatigue tolerance. These changes correlated with a larger number of relatively more oxidative type 2-A and 2X fibers, and a reduced amount of 2B fibers. Our data suggests that SDHC is a key regulator of respiration and fatigability in pulmonary emphysema-driven skeletal muscles, which could be impactful to develop strategies aimed at attenuating this comorbidity.

Pre-mRNA processing entropy in a mouse model of trauma

Jentzsch, M. S. — 2021-01-26

PurposeNext generation sequencing has expanded our understanding of many disease processes, including trauma and critical illness. Many studies focus identifying a small set of genes or proteins that are aberrantly expressed. Our objective was to determine whether global differences in pre-mRNA processing entropy, or disorder, could offer novel insights in the setting of critical illness. MethodsWe used an established murine model of trauma that consisted of hemorrhagic shock and cecal ligation and puncture. In our first experiment mice exposed to trauma were compared to controls. In our second experiment, survival 14 days after exposure to trauma was studied. Using deep RNA sequencing we determined entropy values for every pre-mRNA processing event identified. We then used principal component analysis (PCA) to conduct unsupervised classification of the data. ResultsMice exposed to trauma separated from controls using PCA. Similarly, mice that did not survive 14 days post exposure clustered closely together on PCA. ConclusionOur results suggest that there is a substantial difference in global pre-mRNA processing entropy in mice exposed to trauma vs. controls, and that pre-mRNA processing entropy may be helpful in predicting mortality. The method introduced here is easily transferrable to other disease processes and samples.

The lysosomal chloride-proton exchanger CLC7 functions in melanosomes as a negative regulator of human pigmentation

Koroma, D. C. — 2021-02-06

Mutations in the Cl-/H+ exchanger CLC7 and its subunit OSTM1 result in osteopetrosis, lysosomal disorders, and pigmentation defects in mice and humans. How CLC7/OSTM1 regulates pigmentation in skin and hair melanocytes remains unexplored. In human epidermal melanocytes, we found CLC7/OSTM1 localized to melanosomes, the organelles in which melanin is synthesized, where it negatively regulates melanin production. Using a novel ratiometric melanosomal pH indicator, we showed that CLC7 acidifies melanosomes, opposing the function of the oculocutaneous albinism II (OCA2) Cl- ion channel. The de novo CLC7 variant (CLC7-Y715C) that causes albinism in humans and mice, decreased melanocytes pigmentation, which was restored by coexpression of OCA2. Remarkably, the enlarged hyperacidic vacuoles caused by CLC7-Y715C were also rescued by OCA2 coexpression in both melanocytes and non-melanocytic cells. Our data uncover a novel mechanism by which CLC7 regulates melanocyte pigmentation and identifies OCA2 as a tool to counteract the effects of CLC7 activating mutations.

Evolution of gene-rich germline restricted chromosomes in black-winged fungus gnats through introgression (Diptera: Sciaridae)

Hodson, C. N. — 2021-02-09

Germline restricted DNA has evolved in diverse animal taxa, and is found in several vertebrate clades, nematodes, and flies. In these lineages, either portions of chromosomes or entire chromosomes are eliminated from somatic cells early in development, restricting portions of the genome to the germline. Little is known about why germline restricted DNA has evolved, especially in flies, in which three diverse families, Chironomidae, Cecidomyiidae, and Sciaridae exhibit germline restricted chromosomes (GRCs). We conducted a genomic analysis of germline restricted chromosomes in the fungus gnat Bradysia (Sciara) coprophila (Diptera: Sciaridae), which carries two large germline restricted "L" chromosomes. We sequenced and assembled the genome of B. coprophila, and used differences in sequence coverage and k-mer frequency between somatic and germ tissues to identify GRC sequence and compare it to the other chromosomes in the genome. We found that the GRCs in B. coprophila are large, gene-rich, and have many genes with paralogs on other chromosomes in the genome. We also found that the GRC genes are extraordinarily divergent from their paralogs, and have sequence similarity to another Dipteran family (Cecidomyiidae) in phylogenetic analyses, suggesting that these chromosomes have arisen in Sciaridae through introgression from a related lineage. These results suggest that the GRCs may have evolved through an ancient hybridization event, raising questions about how this may have occurred, how these chromosomes became restricted to the germline after introgression, and why they were retained over time.

The foraging gene affects alcohol sensitivity, metabolism and memory in Drosophila

Oepen, A. S. — 2021-02-10

The genetic basis of alcohol use disorder (AUD) is complex. Understanding how natural genetic variation contributes to alcohol phenotypes can help identify mechanisms underlying the genetic contribution of AUD. Recently, a single nucleotide polymorphism in the human foraging (for) gene ortholog, Protein Kinase cGMP-Dependent 1 (PRKG1), was found to be associated with stress-induced risk for alcohol abuse. However, the mechanistic role that PRKG1 plays in AUD is not well understood. We use natural variation in the Drosophila for gene to describe how variation of cGMP-dependent protein kinase (PKG) activity modifies ethanol-induced phenotypes. We found that variation in for affects ethanol-induced increases in locomotion and memory of the appetitive properties of ethanol intoxication. Further, these differences may stem from the ability to metabolize ethanol. Together, this data suggests that natural variation in PKG modulates cue reactivity for alcohol, and thus could influence alcohol cravings by differentially modulating metabolic and behavioral sensitivities to alcohol.

Dynamic Representation of the Subjective Value of Information

Kobayashi, K. — 2021-02-16

To improve future decisions, people should seek information based on the value of information (VOI), which depends on the current evidence and the reward structure of the upcoming decision. When additional evidence is supplied, people should update VOI to adjust subsequent information seeking, but the neurocognitive mechanisms of this updating process remain unknown. We used a modified beads task to examine how the VOI is represented and updated in the human brain. We theoretically derived, and empirically verified, a normative prediction that the VOI depends on decision evidence and is biased by reward asymmetry. Using fMRI, we found that the subjective VOI is represented in right dorsolateral prefrontal cortex (DLPFC). Critically, this VOI representation was updated when additional evidence was supplied, showing that DLPFC dynamically tracks the up-to-date VOI over time. These results provide new insights into how humans adaptively seek information in the service of decision making.

Laminar dynamics of beta bursts in human motor cortex

Bonaiuto, J. J. — 2021-02-17

Modulation of motor cortical activity in the beta frequency range is one of the strongest and most studied movement-related neural signals. At the single trial level, beta band activity is often characterized by transient bursting events rather than slowly modulating oscillations, suggesting a more rapid, information-encoding functional role than previously believed. Insight into how beta bursts are generated in sensorimotor circuits can provide important constraints to theories about their functional role for movement control. To this end, we leverage and extend recent developments in high precision MEG for temporally resolved laminar analysis of burst activity, combined with a neocortical circuit model that simulates the biophysical generators of the electrical currents which drive beta bursts. This approach pinpoints the generation of beta bursts in human motor cortex to distinct excitatory synaptic inputs to deep and superficial cortical layers, which drive current flow in opposite directions. These laminar dynamics of beta bursts in motor cortex align with prior invasive animal recordings within the somatosensory cortex, and suggest a conserved mechanism for somatosensory and motor cortical beta bursts. More generally, we demonstrate the ability for uncovering the laminar dynamics of event-related neural signals in human non-invasive recordings.

Aging predisposes B cells to malignancy by activating c-Myc and perturbing the genome and epigenome

Shindyapina, A. V. — 2021-02-23

While cancer is an age-related disease, many cancer studies utilize younger animal models. Here, we uncover how a cancer, B-cell lymphoma, develops as a consequence of a naturally aged system. We show that this malignancy is associated with increased cell size, splenomegaly, and a newly discovered age-associated clonal B-cell (ACBC) population. Driven by exogenous c-Myc activation, hypermethylated promoters and somatic mutations, ACBC cells clonally expand independent of germinal centers (IgM+) and show increased biological age and hypomethylation in partially methylated domains related to mitotic solo-CpGs. Epigenetic changes in transformed mouse B cells are enriched for changes observed in human B-cell lymphomas. Mechanistically, the data suggest that cancerous ACBC cells originate from age-associated B cells, in part involving CD22 protein signaling fostered by the aging microenvironment. Transplantation assays demonstrate that ACBC evolve to become self-sufficient and support malignancy when transferred into young recipients. Inhibition of mTOR or c-Myc in old mice attenuates premalignant changes in B cells during aging and emerges as a therapeutic strategy to delay the onset of age-related lymphoma. Together, we show how aging contributes to B-cell lymphoma through a previously unrecognized mechanism involving cell-intrinsic changes and the aged microenvironment, characterize a model that captures the origin and progression of spontaneous cancer during aging and identify candidate interventions against age-associated lymphoma.

Glia-neuron signaling mediated by two different BMP ligands impacts synaptic growth

Bartoletti, M. — 2021-02-24

The nervous system is a complex network of cells whose interactions provide circuitry necessary for an organism to perceive and move through its environment. Revealing the molecular basis of how neurons and non-neuronal glia communicate is essential for understanding neural development, behavior, and abnormalities of the nervous system. BMP signaling in motor neurons, activated in part by retrograde signals from muscle expressed Gbb (BMP5/6/7) has been implicated in synaptic growth, function and plasticity in Drosophila melanogaster. Through loss-of-function studies, we establish Gbb as a critical mediator of glia to neuron signaling important for proper synaptic growth. Furthermore, the BMP2/4 ortholog, Dpp, expressed in a subset of motor neurons, acts by autocrine signaling to also facilitate neuromuscular junction (NMJ) growth at specific muscle innervation sites. In addition to signaling from glia to motor neurons, autocrine Gbb induces signaling in larval VNC glia which strongly express the BMP type II receptor, Wit. In addition to Dpps autocrine motor neuron signaling, Dpp also engages in paracrine signaling to adjacent glia but not to neighboring motor neurons. In one type of dorsal midline motor neuron, RP2, dpp transcription is under tight regulation, as its expression is under autoregulatory control in RP2 but not aCC neurons. Taken together our findings indicate that bi-directional BMP signaling, mediated by two different ligands, facilitates communication between glia and neurons. Gbb, prominently expressed in glia, and Dpp acting from a discrete set of neurons induce active Smad-dependent BMP signaling to influence bouton number during neuromuscular junction growth.

The oncogenic transcription factor FUS-CHOP can undergo nuclear liquid-liquid phase separation

Owen, I. — 2021-02-24

Myxoid liposarcoma is caused by a chromosomal translocation resulting in a fusion protein comprised of the N-terminus of FUS (fused in sarcoma) and the full-length transcription factor CHOP (CCAAT/Enhancer Binding Protein Homologous Protein). FUS functions in RNA metabolism and CHOP is a stress-induced transcription factor. The FUS-CHOP fusion protein causes unique gene expression and oncogenic transformation. The FUS segment is required for oncogenic transformation, but the mechanism of FUS-CHOP-induced transcriptional activation is unknown. Recently, some transcription factors and super enhancers were proposed to undergo liquid-liquid phase separation and form membraneless compartments that recruit transcription machinery to gene promoters. Since phase separation of FUS depends on its N-terminus, transcriptional activation by FUS-CHOP could result from the N-terminus driving nuclear phase transitions. Here, we characterized FUS-CHOP in cells and in vitro, and observed novel phase-separating properties relative to unmodified CHOP. Our data indicate FUS-CHOP forms phase-separated condensates at super enhancer transcriptional sites. We provide strong evidence that the FUS-CHOP phase transition is a novel oncogenic mechanism and potential therapeutic target for treatment of myxoid liposarcoma.

Kasugamycin is a novel chitinase 1 inhibitor with strong antifibrotic effects on pulmonary fibrosis

Lee, J.-H. — 2021-02-26

RationalePulmonary fibrosis is a devastating lung disease with few therapeutic options. Chitinase 1 (CHIT1), an 18 glycosyl hydrolase family member, contributes to the pathogenesis of pulmonary fibrosis through regulation of Transforming Growth Factor (TGF)-{beta} signaling and effector function. Therefore, CHIT1 is a potential therapeutic target of pulmonary fibrosis. ObjectivesThis study aimed to identify and characterize a druggable CHIT1 inhibitor with strong antifibrotic activity and minimal toxicity for therapeutic application to pulmonary fibrosis. MethodsExtensive screening of small molecule libraries identified the aminoglycoside antibiotic Kasugamycin as a potent CHIT1 inhibitor. Measurements and Main ResultsElevated levels of CHIT1 were detected in the lungs of patients with pulmonary fibrosis. In vivo bleomycin- and TGF-{beta}-stimulated murine models of pulmonary fibrosis, Kasugamycin showed impressive anti-fibrotic effects in both preventive and therapeutic conditions. In vitro studies also demonstrated that Kasugamycin inhibits fibrotic macrophage activation, fibroblast proliferation and myofibroblast transformation. Null mutation of transforming growth factor beta associated protein 1 (TGFBRAP1), a recently identified CHIT1 interacting signaling molecule, phenocopied antifibrotic effects of Kasugamycin in in vivo lungs and in vitro fibroblasts responses. Kasugamycin inhibits physical association between CHIT1 and TGFBRAP1, suggesting that antifibrotic effect of Kasugamycin is mediated through regulation of TGFBRAP1, at least in part. ConclusionsThese studies demonstrate that Kasugamycin is a novel CHIT1 inhibitor with strong antifibrotic effect that can be further developed as an effective and safe therapeutic drug for pulmonary fibrosis.

Graphene oxide/silver nanoparticle ink formulations rapidly inhibit influenza A virus and OC43 coronavirus infection in vitro

Crane, M. J. — 2021-02-26

Respiratory tract infections present a significant risk to the human population, both through seasonal circulation and novel introductions with pandemic potential. There is a strong need for antiviral compounds with broad antimicrobial activity that can be coated onto filtration systems and personal protective equipment to augment their ability to remove infectious particles from the environment. Graphene oxide and silver nanoparticles are both materials with documented antimicrobial properties. Here, we tested the in vitro antiviral properties of several graphene oxide-silver nanoparticle composite materials, which were prepared through three different methods: reduction with silver salt, direct addition of silver nanospheres, and direct addition of silver nanospheres to thiolized graphene. These materials were tested over short time scales for their antiviral activity against two enveloped RNA viruses, influenza A virus and OC43 coronavirus, by performing viral plaque assays after exposure of the viruses to each material. It was found that the graphene oxide - silver nanoparticle materials generated by direct addition of the silver nanospheres were able to completely inhibit plaque formation by both viruses within one minute of exposure. Materials generated by the other two methods had varying levels of efficacy against influenza A virus. These studies indicate that graphene oxide-silver nanoparticle composite materials can rapidly neutralize RNA viruses and demonstrate their potential for use in a wide range of applications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/432893v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@7940d4org.highwire.dtl.DTLVardef@8f3207org.highwire.dtl.DTLVardef@11d9c1eorg.highwire.dtl.DTLVardef@f489a2_HPS_FORMAT_FIGEXP M_FIG C_FIG

Identification of Human CD4+ Sub-population of Resident Cardiac Fibroblasts Linked to Inflammation-Mediated Cardiac Fibrosis

Siamwala, J. H. — 2021-02-26

Infiltration with inflammatory T-cells and accumulation of cardiac myofibroblasts are hallmarks of cardiac fibrosis and maladaptive remodeling. The origin, identity, and functions of the resident cardiac cells involved in this process are, however, unclear. To determine the identity of cells contained in regions exhibiting fibrosis, mass cytometry profiling was performed using resident human ventricular cardiac fibroblasts and right ventricle autopsy tissues from individuals diagnosed with pulmonary hypertension and SUGEN/hypoxia rats. Results showed that a subpopulation of resident myocardial fibroblasts expresses increased levels of CD4+, a helper T-cell surface marker, in addition to mesenchymal markers in humans and rats. Characterization of the resident cardiac fibroblast subpopulation, both structurally and functionally, using transcriptome and secretome analysis of the secreted cytokines, chemokines, proteins, and metabolites, evidenced that IL-1{beta} induces a phenotypic switch of human cardiac fibroblasts from mesenchymal to CD4+ lymphoidal lineage in vitro. RNA sequencing (RNA-seq) analysis of FACS-sorted CD4-expressing cardiac fibroblasts further revealed that the transcriptome of such IL-1{beta}-induced CD4+ fibroblast population exhibited classical lymphoidal and stem cell-like signatures. Lastly, reversal of cell clustering, phosphorylation of MAPK p38 and NF-{kappa}B p65, and phenotypic switching was achieved with the administration of an IL-1R antagonist. In conclusion, we have identified a subpopulation of cardiac fibroblasts which exhibits structural and functional attributes of both mesenchymal and lymphoid cells which is induced by IL-1{beta}-IL-1R-NFkB pathway for differentiation of cardiac fibroblast cells. These data suggest that cardiac fibroblast transdifferentiation during inflammation may form the basis for maladaptive remodeling during myocardial fibrosis.

Intra-species transcriptional profiling reveals key regulators of Candida albicans pathogenic traits

Wang, J. M. — 2021-03-04

The human commensal and opportunistic fungal pathogen Candida albicans displays extensive genetic and phenotypic variation across clinical isolates. Here, we performed RNA sequencing on 21 well-characterized isolates to examine how genetic variation contributes to gene expression differences, and to link these differences to phenotypic traits. C. albicans adapts primarily through clonal evolution and yet hierarchical clustering of gene expression profiles in this set of isolates did not reproduce their phylogenetic relationship. Strikingly, strain-specific gene expression was prevalent in some strain backgrounds. Association of gene expression with phenotypic data by differential analysis, linear correlation, and assembly of gene networks connected both previously characterized and novel genes with 23 C. albicans traits. Construction of de novo gene modules produced a gene atlas incorporating 67% of C. albicans genes and revealed correlations between expression modules and important phenotypes such as systemic virulence. Furthermore, targeted investigation of two modules that have novel roles in growth and filamentation supported our bioinformatic predictions. Together, these studies reveal widespread transcriptional variation across C. albicans isolates and identify genetic and epigenetic links to phenotypic variation based on co-expression network analysis. ImportanceInfectious fungal species are often treated uniformly despite clear evidence of genotypic and phenotypic heterogeneity being widespread across strains. Identifying the genetic basis for this phenotypic diversity is extremely challenging because of the tens or hundreds of thousands of variants that may distinguish two strains. Here we use transcriptional profiling to determine differences in gene expression that can be linked to phenotypic variation among a set of 21 Candida albicans isolates. Analysis of this transcriptional dataset uncovered clear tends in gene expression characteristics for this species and new genes and pathways that associated with variation in pathogenic processes. Direct investigation confirmed functional predictions for a number of new regulators associated with growth and filamentation, demonstrating the utility of these approaches in linking genes to important phenotypes.

Single cell analysis of the aging hypothalamus

Hajdarovic, K. H. — 2021-03-08

Alterations in metabolism, sleep patterns, body composition, and hormone status are all key features of aging. The hypothalamus is a well-conserved brain region that controls these homeostatic and survival-related behaviors. Despite the importance of this brain region in healthy aging, little is known about the intrinsic features of hypothalamic aging. Here, we utilize single nuclei RNA-sequencing to assess the transcriptomes of 40,064 hypothalamic nuclei from young and aged female mice. We identify cell type-specific signatures of aging in neurons, astrocytes, and microglia, as well as among the diverse collection of neuronal subtypes in this region. We uncover key changes in cell types critical for metabolic regulation and body composition, as well as in an area of the hypothalamus linked to cognition. In addition, our analysis reveals an unexpected female-specific feature of hypothalamic aging. Specifically, we discover that the master regulator of X-inactivation, Xist, is elevated with age, particularly in hypothalamic neurons. Moreover, using machine learning, we show that levels of X-chromosome genes, and Xist itself, are the best predictors of cellular age. Together, this study identifies critical cell-specific changes of the aging hypothalamus in mammals, and uncovers a novel marker of neuronal aging in females.

Multivariate Pattern Analysis (MVPA) Reveals Distinct Neural Profiles of Frontoparietal Networks in Boys with Attention-Deficit/Hyperactivity Disorder and Boys with Persistent Depressive Disorder

Vilgis, V. — 2021-03-10

Working memory deficits are common in attention-deficit/hyperactivity disorder (ADHD) and depression, two common neurodevelopmental disorders with overlapping cognitive profiles but distinct clinical presentation. Multivariate techniques have previously been utilized to understand working memory processes in functional brain networks in healthy adults, but have not yet been applied to investigate how working memory processes within the same networks differ within typical and atypical developing populations. We used multivariate pattern analysis (MVPA) to identify whether brain networks discriminated between spatial vs. verbal working memory processes in ADHD and Persistent Depressive Disorder (PDD). 36 male clinical participants and 19 typically developing (TD) boys participated in a fMRI scan while completing a verbal and a spatial working memory task. Within a priori functional brain networks (frontoparietal, default mode, salience) the TD group demonstrated differential response patterns to verbal and spatial working memory. The PDD group showed weaker differentiation than TD, with lower classification accuracies observed in primarily the left frontoparietal network. The neural profiles of the ADHD and PDD differed specifically in the SN where the ADHD groups neural profile suggests significantly less specificity in neural representations of spatial and verbal working memory. We highlight within-group classification as an innovative tool for understanding the neural mechanisms of how cognitive processes may deviate in clinical disorders, an important intermediary step towards improving translational psychiatry.

Lamb1a regulates atrial growth by limiting excessive, contractility-dependent second heart field addition during zebrafish heart development

Derrick, C. J. — 2021-03-10

During early vertebrate heart development, the heart transitions from a linear tube to a complex asymmetric structure. This process includes looping of the tube and ballooning of the emerging cardiac chambers, which occur simultaneously with growth of the heart. A key driver of cardiac growth is deployment of cells from the Second Heart Field (SHF) into both poles of the heart, with cardiac morphogenesis and growth intimately linked in heart development. Laminin is a core component of extracellular matrix (ECM) basement membranes, and although mutations in specific laminin subunits are linked with a variety of cardiac abnormalities, including congenital heart disease and dilated cardiomyopathy, no role for laminin has been identified in early vertebrate heart morphogenesis. We identified dynamic, tissue-specific expression of laminin subunit genes in the developing zebrafish heart, supporting a role for laminins in heart morphogenesis. lamb1a mutants exhibit cardiomegaly from 2dpf onwards, with subsequent progressive defects in cardiac morphogenesis characterised by a failure of the chambers to compact around the developing atrioventricular canal. We show that loss of lamb1a results in excess addition of SHF cells to the atrium, revealing that Lamb1a functions to limit heart size during cardiac development by restricting SHF addition to the venous pole. lamb1a mutants exhibit hallmarks of altered haemodynamics, and specifically blocking cardiac contractility in lamb1a mutants rescues heart size and atrial SHF addition. Furthermore, we identify that FGF and RA signalling, two conserved pathways promoting SHF addition, are regulated by heart contractility and are dysregulated in lamb1a mutants, suggesting that laminin mediates interactions between SHF deployment, heart biomechanics, and biochemical signalling during heart development. Together, this describes the first requirement for laminins in early vertebrate heart morphogenesis, reinforcing the importance of specialised ECM composition in cardiac development.

The level of synovial human VEGFA, IL-8 and MIP-1α correlate with truncation of lubricin glycans in osteoarthritis

Huang, S. — 2021-03-11

Osteoarthrithis (OA) is an endemic disease due to the increase of the worlds elderly population. Previously thought to be a consequence of an imbalance between cartilage degradation and biosynthesis, it is now recognized as a disease also involving inflammation, hence influencing the level of inflammatory cytokines, growth factors and chemokines. Lubricin is a mucin type molecule where its OA induced glycosylation truncation propels a deteriorating lubrication of the articular cartilage. The objective of this study was to explore the OA driven truncation of O-linked glycosylation of synovial lubricin and its cross talk with systemic and local (synovial fluid, SF) inflammation. We compared the systemic level of cytokines/chemokine in OA patients and controls plasma with their local level in SF using a 44 plex screen. The level of 27 cytokines and chemokines was consistently measured in both plasma and SF. The data showed that the levels of cytokines and chemokines in OA plasma display limited correlation to their counterpart in SF. The level of synovial IL-8 and MIP-1 and VEGFA in OA patients, but not their plasma level, where the only cytokines that displayed a significant correlation to the observed lubricin O-linked glycosylation truncation. These cytokines were also shown to be upregulated exposing fibroblast like synoviocytes from healthy and OA patients to recombinant lubricin with truncated glycans mainly consisting of Tn-antigens, while lubricin with sialylated and non-sialylated T anigens did not have any effect. The data suggest that truncated glycans of lubricin, as found in OA, promotes the synovial cytokine production and exerebate the local synovial inflammation.

Sub-second multi-channel magnetic control of select neural circuits in behaving flies

Sebesta, C. — 2021-03-16

Precisely timed activation of genetically targeted cells is a powerful tool for studying neural circuits and controlling cell-based therapies. Magnetic control of cell activity or "magnetogenetics" using magnetic nanoparticle heating of temperature-sensitive ion channels enables remote, non-invasive activation of neurons for deep-tissue applications and studies of freely behaving animals. However, the in vivo response time of thermal magnetogenetics is currently tens of seconds, which prevents the precise temporal modulation of neural activity similar to light-based optogenetics. Moreover, magnetogenetics has not provided a means to selectively activate multiple channels to drive behavior. Here we produce sub-second behavioral responses in Drosophila melanogaster by combining magnetic nanoparticles with a rate-sensitive thermoreceptor (TRPA1-A). Furthermore, by tuning the properties of magnetic nanoparticles to respond to different magnetic field strengths and frequencies, we can achieve sub-second, multichannel stimulation, analogous to multi-color optogenetic stimulation. These results bring magnetogenetics closer to the temporal resolution and multiplexed stimulation possible with optogenetics while maintaining the minimal invasiveness and deep-tissue stimulation only possible by magnetic control.

Sex-specific transcript diversity is regulated by a maternal pioneer factor in early Drosophila embryos

Ray, M. — 2021-03-19

Co-transcriptional splicing coordinates the processes of transcription and splicing and is driven by transcription factors (TFs) and diverse RNA-binding proteins (RBPs). Yet the mechanisms by which specific TFs and RBPs function together in context-specific ways to drive precise co-transcriptional splicing at each of thousands of genomic loci remains unknown. Therefore, we have used sex-specific splicing in Drosophila as a model to understand how the function of TFs and RBPs is coordinated to transcribe and process specific RNA transcripts at the correct genomic locations. We show widespread sex-specific transcript diversity occurs much earlier than previously thought and present a new pipeline called time2splice to quantify splicing changes over time. We define several mechanisms by which the essential and functionally-conserved CLAMP TF functions with specific RBPs to precisely regulate co-transcriptional splicing: 1) CLAMP links the DNA of gene bodies of sex-specifically spliced genes directly to the RNA of target genes and physically interacts with snRNA and protein components of the splicing machinery; 2) In males, CLAMP regulates the distribution of the highly conserved RBP Maleless (MLE) (RNA Helicase A) to prevent aberrant sex-specific splicing; 3) In females, CLAMP modulates alternative splicing by directly binding to target DNA and RNA and indirectly through regulating the splicing of sex lethal, the master regulator of sex determination. Overall, we provide new insight into how TFs function specifically with RBPs to drive alternative splicing.

Explicit and implicit depth-cue integration: evidence of systematic biases with real objects

Campagnoli, C. — 2021-03-22

In a previous series of experiments using virtual stimuli, we found evidence that 3D shape estimation agrees to a superadditivity rule of depth-cue combination. According to this rule, adding depth cues leads to greater perceived depth magnitudes and, in principle, to depth overestimation. The mechanism underlying the superadditivity effect can be fully accounted for by a normative theory of cue integration, through the adaptation of a model of cue integration termed the Intrinsic Constraint (IC) model. As for its nature, it remains unclear whether superadditivity is a byproduct of the artificial nature of virtual environments, causing explicit reasoning to infiltrate behavior and inflate the depth judgments when a scene is richer in depth cues, or the genuine output of the process of depth-cue integration. In the present study, we addressed this question by testing whether the IC models prediction of superadditivity generalizes beyond VR environments to real world situations. We asked participants to judge the perceived 3D shape of cardboard prisms through a matching task. To assay the potential influence of explicit control over those perceptual estimates, we also asked participants to reach and hold the same objects with their fingertips and we analyzed the in-flight grip size during the reaching. Using physical objects ensured that all visual information was fully consistent with the stimulis 3D structure without computer-generated artifacts. We designed a novel technique to carefully control binocular and monocular 3D cues independently from one another, allowing to add or remove depth information from the scene seamlessly. Even with real objects, participants exhibited a clear superadditivity effect in both explicit and implicit tasks. Furthermore, the magnitude of this effect was accurately predicted by the IC model. These results confirm that superadditivity is an inherent feature of depth estimation.

Ovalbumin antigen-specific activation of T cell receptor closely resembles soluble antibody stimulation as revealed by BOOST phosphotyrosine proteomics

Chua, X. Y. — 2021-03-25

Activation of T cell receptors (TCR) leads to a network of early signaling predominantly orchestrated by tyrosine phosphorylation in T cells. TCR are commonly activated using soluble anti-TCR antibodies, but this approach is not antigen-specific. Alternatively, activating the TCR using specific antigens of a range of binding affinities in the form of peptide-major histocompatibility complex (pMHC) is presumed to be more physiological. However, due to the lack of wide-scale phosphotyrosine (pTyr) proteomic studies directly comparing anti-TCR antibodies and pMHC, a comprehensive definition of these activated states remains enigmatic. Elucidation of the tyrosine phosphoproteome using quantitative pTyr proteomics enables a better understanding of the unique features of these activating agents and the role of ligand binding affinity on signaling. Here, we apply the recently established Broad-spectrum Optimization Of Selective Triggering (BOOST) to examine perturbations in tyrosine phosphorylation of TCR triggered by anti-TCR antibodies and pMHC. Our data reveals that high-affinity ovalbumin (OVA) pMHC activation of the TCR triggers a largely similar, albeit potentially stronger, pTyr-mediated signaling regulatory axis compared to anti-TCR antibody. Signaling output resulting from OVA pMHC variants correlates well with their weaker affinities, enabling affinity-tunable control of signaling strength. Collectively, we provide a framework for applying BOOST to compare pTyr-mediated signaling pathways of T cells activated in an antigen-independent and antigen-specific manner. Abstract Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/436968v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@152f03aorg.highwire.dtl.DTLVardef@19a6e2org.highwire.dtl.DTLVardef@1b48b6org.highwire.dtl.DTLVardef@142ced6_HPS_FORMAT_FIGEXP M_FIG C_FIG

Subthalamic-Cortical Network Reorganization during Parkinson's Tremor

Lauro, P. — 2021-03-29

Tremor, a common and often primary symptom of Parkinsons disease, has been modeled with distinct onset and maintenance dynamics. To identify the neurophysiologic correlates of each state, we acquired intraoperative cortical and subthalamic nucleus recordings from ten patients performing a naturalistic visual-motor task. From this task we isolated short epochs of tremor onset and sustained tremor. Comparing these epochs, we found that the subthalamic nucleus was central to tremor onset, as it drove both motor cortical activity and tremor output. Once tremor became sustained, control of tremor shifted to cortex. At the same time, changes in directed functional connectivity across sensorimotor cortex further distinguished the sustained tremor state. SIGNIFICANCE STATEMENTTremor is a common symptom of Parkinsons disease (PD). While tremor pathophysiology is thought to involve both basal ganglia and cerebello-thalamic-cortical circuits, it is unknown how these structures functionally interact to produce tremor. In this manuscript, we analyzed intracranial recordings from the subthalamic nucleus and sensorimotor cortex in patients with PD undergoing deep brain stimulation (DBS) surgery. Using an intraoperative task, we examined tremor in two separate dynamic contexts: when tremor first emerged, and when tremor was sustained. We believe that these findings reconcile several models of Parkinsons tremor, while describing the short-timescale dynamics of subcortical-cortical interactions during tremor for the first time. These findings may describe a framework for developing proactive and responsive neurostimulation models for specifically treating tremor.

Predicted Utility Modulates Working Memory Fidelity in the Brain

Levin, E. J. — 2021-04-02

The predicted utility of information stored in working memory (WM) is hypothesized to influence the strategic allocation of WM resources. Prior work has shown that when information is prioritized, it is remembered with greater precision relative to other remembered items. However, these paradigms often complicate interpretation of the effects of predicted utility on item fidelity due to a concurrent memory load. Likewise, no fMRI studies have examined whether the predicted utility of an item modulates fidelity in the neural representation of items during the memory delay without a concurrent load. In the current study, we used fMRI to investigate whether predicted utility influences fidelity of WM representations in the brain. Using a generative model multivoxel analysis approach to estimate the quality of remembered representations across predicted utility conditions, we observed that items with greater predicted utility are maintained in memory with greater fidelity, even when they are the only item being maintained. Further, we found that this pattern follows a parametric relationship where more predicted utility corresponded to greater fidelity. These precision differences could not be accounted for based on a redistribution of resources among already-remembered items. Rather, we interpret these results in terms of a gating mechanism that allows for pre-allocation of resources based on predicted value alone. This evidence supports a theoretical distinction between resource allocation that occurs as a result of load and resource pre-allocation that occurs as a result of predicted utility.

Taking aim at the perceptual side of motor learning: Exploring how explicit and implicit learning encode perceptual error information through depth vision

Campagnoli, C. — 2021-04-04

Motor learning in visuomotor adaptation tasks results from both explicit and implicit processes, each responding differently to an error signal. While the motor output side of these processes is extensively studied, their visual input side is relatively unknown. We investigated if and how depth perception affects the computation of error information by explicit and implicit motor learning. Two groups of participants threw virtual darts at a virtual dartboard while receiving perturbed endpoint feedback. The Delayed group was allowed to re-aim and their feedback was delayed to emphasize explicit learning, while the Clamped group received clamped cursor feedback which they were told to ignore, and continued to aim straight at the target to emphasize implicit adaptation. Both groups played this game in a highly detailed virtual environment (Depth condition) and in an empty environment (No-Depth condition). The Delayed group showed an increase in error sensitivity under Depth relative to No-Depth conditions. In contrast, the Clamped group adapted to the same degree under both conditions. The movement kinematics of the Delayed participants also changed under the Depth condition, consistent with the target appearing more distant, unlike the Clamped group. A comparison of the Delayed behavioral data with a perceptual task from the same individuals showed that the effect of the Depth condition on the re-aiming direction was consistent with an increase in the scaling of the error distance and size. These findings suggest that explicit and implicit learning processes may rely on different sources of perceptual information. New & NoteworthyWe leveraged a classic sensorimotor adaptation task to perform a first systematic assessment of the role of perceptual cues in the estimation of an error signal in the 3D space during motor learning. We crossed two conditions presenting different amounts of depth information, with two manipulations emphasizing explicit and implicit learning processes. Explicit learning responded to the visual conditions, consistent with perceptual reports, while implicit learning appeared to be independent of them.

Integrative multi-omics analysis reveals conserved hierarchical mechanisms of FOXO3 pioneer-factor activity

Brown, A. K. — 2021-04-06

FOXO transcription factors are critical for cellular homeostasis and have been implicated in longevity in several species. Yet how these factors directly affect aging, particularly in humans, is not well understood. Here, we take an integrated multi-omics approach to identify the chromatin-level mechanisms by which FOXO3 coordinates transcriptional programs. We find that FOXO3 functions as a pioneer factor in human cells, directly altering chromatin accessibility to regulate gene expression. Unexpectedly, FOXO3s pioneer activity at many sites is achieved through a two-step process, in which chromatin accessibility is initially reduced, then transitions to an open conformation. The direct FOXO3 network comprises chromatin remodelers, including the SWI/SNF remodeling complex, which we find is functionally required for FOXO3 activity. We also identify a novel secondary network of activator protein-1 (AP-1) transcription factors deployed by FOXO3, which orchestrate a neuronal-specific subnetwork. Together, this hierarchical FOXO3 pioneer network regulates key cellular processes including metabolism, proteostasis, epigenetics and proliferation, which must be tightly controlled under changing conditions that accompany aging.

Thunderstruck: The ACDC model of flexible sequences and rhythms in recurrent neural circuits

Calderon, C. B. — 2021-04-08

Adaptive sequential behavior is a hallmark of human cognition. In particular, humans can learn to produce precise spatiotemporal sequences given a certain context. For instance, musicians can not only reproduce learned action sequences in a context-dependent manner, they can also quickly and flexibly reapply them in any desired tempo or rhythm without overwriting previous learning. Existing neural network models fail to account for these properties. We argue that this limitation emerges from the fact that sequence information (i.e., the position of the action) and timing (i.e., the moment of response execution) are typically stored in the same neural network weights. Here, we augment a biologically plausible recurrent neural network of cortical dynamics to include a basal ganglia-thalamic module which uses reinforcement learning to dynamically modulate action. This "associative cluster-dependent chain" (ACDC) model modularly stores sequence and timing information in distinct loci of the network. This feature increases computational power and allows ACDC to display a wide range of temporal properties (e.g., multiple sequences, temporal shifting, rescaling, and compositionality), while still accounting for several behavioral and neurophysiological empirical observations. Finally, we apply this ACDC network to show how it can learn the famous "Thunderstruck" song intro and then flexibly play it in a "bossa nova" rhythm without further training.

Chitinase 3-like-1 Contributes to Acetaminophen-induced Liver Injury by Promoting Hepatic Platelet Recruitment

Shan, Z. — 2021-04-09

Hepatic platelet accumulation contributes to acetaminophen (APAP)-induced liver injury (AILI). However, little is known about the molecular pathways involved in platelet recruitment to the liver and whether targeting such pathways could attenuate AILI. The present study unveiled a critical role of chitinase 3-like-1 (Chi3l1) in hepatic platelet recruitment during AILI. Increased Chi3l1 and platelets in the liver were observed in patients and mice overdosed with APAP. Compared to wild-type (WT) mice, Chi3l1-/- mice developed attenuated AILI with markedly reduced hepatic platelet accumulation. Mechanistic studies revealed that Chi3l1 signaled through CD44 on macrophages to induce podoplanin expression, which mediated platelet recruitment through C-type lectin-like receptor 2. Moreover, APAP treatment of CD44-/- mice resulted in much lower numbers of hepatic platelets and liver injury than WT mice, a phenotype similar to that in Chi3l1-/- mice. Recombinant Chi3l1 could restore hepatic platelet accumulation and AILI in Chi3l1-/- mice, but not in CD44-/- mice. Importantly, we generated anti-Chi3l1 monoclonal antibodies and demonstrated that they could effectively inhibit hepatic platelet accumulation and AILI. Overall, we uncovered the Chi3l1/CD44 axis as a critical pathway mediating APAP-induced hepatic platelet recruitment and tissue injury. We demonstrated the feasibility and potential of targeting Chi3l1 to treat AILI.

Single-cell transcriptomics of peripheral blood in the aging mouse

Teo, Y. V. — 2021-04-10

Compositional and transcriptional changes in the hematopoietic system have been used as biomarkers of immunosenescence and aging. Here, we use single-cell RNA-sequencing to study the aging peripheral blood in mice, and characterize the changes in cell-type composition and transcriptional profiles associated with age. We identified 17 clusters from a total of 14,588 single cells. We detected a general upregulation of antigen processing and presentation and chemokine signaling pathways and a downregulation of genes involved in ribosome pathways with age. We also observed increased percentage of cells expressing markers of senescence, Cdkn1a and Cdkn2a, in old peripheral blood. In addition, we detected a cluster of activated T cells that are exclusively found in old blood, with lower expression of Cd28 and higher expression of Bcl2 and Cdkn2a, suggesting that the cells are senescent and resistant to apoptosis.

Multiple 9-1-1 complexes promote homolog synapsis, DSB repair, and ATR signaling during mammalian meiosis

Pereira, C. — 2021-04-10

DNA damage response mechanisms have meiotic roles that ensure successful gamete formation. While completion of meiotic double-strand break (DSB) repair requires the canonical RAD9A-RAD1-HUS1 (9A-1-1) complex, mammalian meiocytes also express RAD9A and HUS1 paralogs, RAD9B and HUS1B, predicted to form alternative 9-1-1 complexes. The RAD1 subunit is shared by all predicted 9-1-1 complexes and localizes to meiotic chromosomes even in the absence of HUS1 and RAD9A. Here we report that testis-specific RAD1 disruption resulted in impaired DSB repair, germ cell depletion and infertility. Unlike Hus1 or Rad9a disruption, Rad1 loss also caused defects in homolog synapsis, ATR signaling and meiotic sex chromosome inactivation. Comprehensive testis phosphoproteomics revealed that RAD1 and ATR coordinately regulate numerous proteins involved in DSB repair, meiotic silencing, synaptonemal complex formation, and cohesion. Together, these results establish critical roles for both canonical and alternative 9-1-1 complexes in meiotic ATR activation and successful prophase I completion.

Enhanced cell deconvolution of peripheral blood using DNA methylation for high-resolution immune profiling

Salas, L. A. — 2021-04-12

DNA methylation microarrays can be employed to interrogate cell-type composition in complex tissues. Here, we expand reference-based deconvolution of blood DNA methylation to include 12 leukocyte subtypes (neutrophils, eosinophils, basophils, monocytes, B cells, CD4+ and CD8+ naive and memory cells, natural killer, and T regulatory cells). Including derived variables, our method provides up to 56 immune profile variables. The IDOL (IDentifying Optimal Libraries) algorithm was used to identify libraries for deconvolution of DNA methylation data both for current and retrospective platforms. The accuracy of deconvolution estimates obtained using our enhanced libraries was validated using artificial mixtures, and whole-blood DNA methylation with known cellular composition from flow cytometry. We applied our libraries to deconvolve cancer, aging, and autoimmune disease datasets. In conclusion, these libraries enable a detailed representation of immune-cell profiles in blood using only DNA and facilitate a standardized, thorough investigation of the immune system in human health and disease.

Thalamocortical mechanisms regulating the relationship between transient beta events and human tactile perception

Law, R. — 2021-04-18

Transient neocortical events with high spectral power in the 15-29Hz beta band are among the most reliable predictors of sensory perception. Prestimulus beta event rates in primary somatosensory cortex correlate with sensory suppression, most effectively 100-300ms before stimulus onset. However, the neural mechanisms underlying this perceptual association are unknown. We combined human magnetoencephalography (MEG) measurements with biophysical neural modeling to test potential cellular and circuit mechanisms that underlie observed correlations between prestimulus beta events and tactile detection. Extending prior studies, we found that simulated bursts from higher-order, non-lemniscal thalamus were sufficient to drive beta event generation and to recruit slow supragranular inhibition acting on a 300ms time scale to suppress sensory information. Further analysis showed that the same beta generating mechanism can lead to facilitated perception for a brief period when beta events occur simultaneously with tactile stimulation before inhibition is recruited. These findings were supported by close agreement between model-derived predictions and empirical MEG data. The post-event suppressive mechanism explains an array of studies that associate beta with decreased processing, while the during-event faciliatory mechanism may demand a reinterpretation of the role of beta events in the context of coincident timing.

When good mutations go bad: how population size can change the direction of natural selection

Raynes, Y. — 2021-04-19

Classical evolutionary theory holds that the efficiency, but not the direction, of natural selection depends on population size. In small populations, drift overwhelms selection, rendering all fitness-affecting mutations selectively neutral. Yet, beneficial mutations never become deleterious and deleterious mutations never become beneficial. Remarkably, several mutations, including in modifiers of recombination and mutation rate, have now been shown to be favored at some population sizes but disfavored at others, challenging established theory. Previously, we have designated this phenomenon sign inversion. Here we show that, unlike selected mutations in the classical framework, mutations susceptible to sign inversion confer both fitness costs and fitness benefits, that vary among their carriers. Furthermore, all such mutations can be classified based on whether their effects differ between or within mutant lineages. Using computer simulations, we demonstrate that both between-lineage and within-lineage variability can cause sign inversion and elucidate the common underlying mechanism. Our results confirm that variability in the sign of selective effects is necessary for sign inversion, which occurs because drift overwhelms selection on carriers bearing the cost and carriers enjoying the benefit at different population sizes.

Redefining replication in multi-ancestry genome-wide association studies

Smith, S. P. — 2021-04-20

Since 2005, genome-wide association (GWA) datasets have been largely biased toward sampling European ancestry individuals, and recent studies have shown that GWA results estimated from self-identified European individuals are not transferable to non-European individuals due to various confounding challenges. Here, we demonstrate that enrichment analyses which aggregate SNP-level association statistics at multiple genomic scales--from genes to genomic regions and pathways--have been underutilized in the GWA era and can generate biologically interpretable hypotheses regarding the genetic basis of complex trait architecture. We illustrate examples of the robust associations generated by enrichment analyses while studying 25 continuous traits assayed in 566,786 individuals from seven diverse self-identified human ancestries in the UK Biobank and the Biobank Japan, as well as 44,348 admixed individuals from the PAGE consortium including cohorts of African-American, Hispanic and Latin American, Native Hawaiian, and American Indian/Alaska Native individuals. We identify 1,000 gene-level associations that are genome-wide significant in at least two ancestry cohorts across these 25 traits, as well as highly conserved pathway associations with triglyceride levels in European, East Asian, and Native Hawaiian cohorts.

CMOS electrochemical imaging arrays for thedetection and classification of microorganisms

Arcadia, C. E. — 2021-04-22

Microorganisms account for most of the biodiversity on earth. Yet while there are increasingly powerful tools for studying microbial genetic diversity, there are fewer tools for studying microorganisms in their natural environments. In this paper, we present recent advances in CMOS electrochemical imaging arrays for detecting and classifying microorganisms. These microscale sensing platforms can provide non-optical measurements of cell geometries, behaviors, and metabolic markers. We review integrated electronic sensors appropriate for monitoring microbial growth, and present measurements of single-celled algae using a CMOS sensor array with thousands of active pixels. Integrated electrochemical imaging can contribute to improved medical diagnostics and environmental monitoring, as well as discoveries of new microbial populations.

Lipid droplets modulate proteostasis, SQST-1/SQSTM1 dynamics, and lifespan in C. elegans

Kumar, A. V. — 2021-04-22

The ability of organisms to live long depends largely on the maintenance of proteome stability via proteostatic mechanisms including translational regulation, protein chaperoning and degradation machineries. In several long-lived Caenorhabditis elegans strains, such as insulin/IGF-1 receptor daf-2 mutants, enhanced proteostatic mechanisms are accompanied by elevated intestinal lipid stores, but the role of lipid droplets in longevity has remained obscure. Here, while determining the regulatory network of the selective autophagy receptor SQST-1/SQSTM1, we unexpectedly uncovered a novel role for lipid droplets in proteostasis and longevity. Using an unbiased genome-wide RNAi screening approach, we identified several SQST-1 modulators, including proteins found on lipid droplets and those prone to aggregate with age. SQST-1 accumulated on lipid droplets when autophagy was inhibited, suggesting that lipid droplets may serve a role in facilitating selective autophagy. Expansion of intestinal lipid droplets by silencing the conserved cytosolic triacylglycerol lipase gene atgl-1/ATGL enhanced autophagy, and extended lifespan in an HSF-1/HSF1-dependent and CDC-48/VCP-dependent manner. Silencing atgl-1 mitigated the age-related accumulation of SQST-1 and reduced overall ubiquitination of proteins. Reducing atgl-1 also improved proteostasis in proteotoxicity models of neurodegenerative diseases. Subcellular analyses revealed that lipid droplets unexpectedly harbor more soluble ubiquitinated proteins than the cytosol. Accordingly, low lipid droplet levels exacerbated the proteostatic collapse when autophagy or proteasome function was compromised. Altogether, our study uncovers a key role for lipid droplets in C. elegans as a proteostatic mediator that reduces protein ubiquitination, facilitates autophagy, and promotes longevity.

Multi-Ancestry Meta-Analysis yields novel genetic discoveries and ancestry-specific associations

Turley, P. — 2021-04-24

We present a new method, Multi-Ancestry Meta-Analysis (MAMA), which combines genome-wide association study (GWAS) summary statistics from multiple populations to produce new summary statistics for each population, identifying novel loci that would not have been discovered in either set of GWAS summary statistics alone. In simulations, MAMA increases power with less bias and generally lower type-1 error rate than other multi-ancestry meta-analysis approaches. We apply MAMA to 23 phenotypes in East-Asian- and European-ancestry populations and find substantial gains in power. In an independent sample, novel genetic discoveries from MAMA replicate strongly.

DeepRegFinder: Deep Learning-Based Regulatory Elements Finder

Ramakrishnan, A. — 2021-04-28

MotivationEnhancers and promoters are important classes of DNA regulatory elements that control gene expression. Identifying them at the genomic scale is a critical and challenging task in bioinformatics. The most successful method so far is to train machine learning models on known enhancer and promoter sites and predict them at other genomic regions using ChIP-seq and related data. ResultsWe have developed a highly customizable program called DeepRegFinder which automates data processing, model training and genome-wide prediction of enhancers and promoters using convolutional and recurrent neural networks. Our program further classifies the enhancers and promoters into active and poised states to facilitate downstream analysis. Based on mean average precision scores of different classes across multiple cell types, our method significantly outperforms the existing algorithms. Availabilityhttps://github.com/shenlab-sinai/DeepRegFinder

Gain control by sparse, ultra-slow glycinergic synapses

Jain, V. — 2021-05-04

SO_SCPLOWUMMARYC_SCPLOWRetinal ON starburst amacrine cells (SACs) play a critical role in computing stimulus direction, partly in service of image stabilization by optokinetic nystagmus. ON SAC responses are sculpted by rich GABAergic innervation, mostly from neighbouring SACs. Surprisingly, however, we find that glycinergic narrow field amacrine cells (NACs) serve as their dominant source of inhibition during sustained activity. Although NAC inputs constitute only [~]5% of inhibitory synapses to ON SACs, their distinct input patterns enable them to drive glycine inhibition during the both light increments and decrements. NAC-to-ON-SAC inhibition appears to be mediated by ultra-slow non-canonical glycine receptors containing the 4 subunit, which effectively summate during repetitive stimulation. Glycinergic inhibition strongly decreases the output gain of the SACs, ensuring that their direction-selective output is maintained over their operating range. These results reveal an unexpected role for glycinergic pathways and receptor kinetics in modulating direction selectivity in the retina.

Reorganization of molecular networks associated with DNA methylation and changes in the rearing environments of the house wren (Troglodytes aedon)

vonHoldt, B. — 2021-05-04

Environmental change, such as increased rates of urbanization, can induce shifts in phenotypic plasticity with some individuals adapting to city life while others are displaced. A key trait that can facilitate adaptation is the degree at which animals respond to stress. This stress response has a heritable component and exhibits intra- and inter-individual variation. However, the mechanisms behind this variability and whether they might be responsible for adaptation to different environments are not known. Variation in DNA methylation can be a potential mechanism that mediates environmental effects on the stress response. We used an inter- and intra-environmental cross-foster experiment to analyze the contribution of DNA methylation to early-life phenotypic variation. We found that at hatching, urban house wren (Troglodytes aedon) offspring had increased methylation as compared to their rural counterparts, and observed plasticity in methylation as offspring aged, indicating developmental effects of the rearing environment on methylation. Differential methylation analyses showed that cellular respiration genes were differentially expressed at hatching and behavioral and metabolism genes were differentially expressed at fledgling. Lastly, hyper-methylation of a single gene (CNTNAP2) is associated with increased glucocorticoid levels. These differential methylation patterns linked to a specific physiological phenotype suggest that DNA methylation may be a mechanism by which individuals adapt to novel environments. Characterizing genetic and environmental influences on methylation is critical for understanding the role of epigenetic mechanisms in evolutionary adaptation.

Thiolutin is a direct inhibitor of RNA Polymerase II

Qiu, C. — 2021-05-05

Thiolutin is a natural product transcription inhibitor with an unresolved mode of action. Thiolutin and the related dithiolopyrrolone holomycin chelate Zn2+ and previous studies have concluded that RNA Polymerase II (Pol II) inhibition in vivo is indirect. Here, we present chemicogenetic and biochemical approaches to investigate thiolutins mode of action in Saccharomyces cerevisiae. We identify mutants that alter sensitivity to thiolutin. We provide genetic evidence that thiolutin causes oxidation of thioredoxins in vivo and that thiolutin both induces oxidative stress and interacts functionally with multiple metals including Mn2+ and Cu2+, and not just Zn2+. Finally, we show direct inhibition of RNA polymerase II (Pol II) transcription initiation by thiolutin in vitro in support of classical studies that thiolutin can directly inhibit transcription in vitro. Inhibition requires both Mn2+ and appropriate reduction of thiolutin as excess DTT abrogates its effects. Pause prone, defective elongation can be observed in vitro if inhibition is bypassed. Thiolutin effects on Pol II occupancy in vivo are widespread but major effects are consistent with prior observations for Tor pathway inhibition and stress induction, suggesting that thiolutin use in vivo should be restricted to studies on its modes of action and not as an experimental tool.

A murine model of the human CREBRFR457Q obesity-risk variant does not influence energy or glucose homeostasis in response to nutritional stress

Kershaw, E. E. — 2021-05-06

Obesity and diabetes have strong heritable components, yet the genetic contributions to these diseases remain largely unexplained. In humans, a missense variant in Creb3 regulatory factor (CREBRF) [rs373863828 (p.Arg457Gln); CREBRFR457Q] is strongly associated with increased odds of obesity but decreased odds of diabetes. Although virtually nothing is known about CREBRFs mechanism of action, emerging evidence implicates it in the adaptive transcriptional response to nutritional stress downstream of TORC1. The objectives of this study were to generate a murine model with knockin of the orthologous variant in mice (CREBRFR458Q) and to test the hypothesis that this CREBRF variant promotes obesity and protects against diabetes by regulating energy and glucose homeostasis downstream of TORC1. To test this hypothesis, we performed extensive phenotypic analysis of CREBRFR458Q knockin mice at baseline and in response to acute (fasting/refeeding), chronic (low- and high-fat diet feeding), and extreme (prolonged fasting) nutritional stress as well as with pharmacological TORC1 inhibition. The results demonstrate that the murine CREBRFR458Q model of the human CREBRFR457Q variant does not influence energy/glucose homeostasis in response to these interventions. Alternative preclinical models and/or studies in humans will be required to decipher the mechanisms linking this variant to human health and disease.

LYRUS: A Machine Learning Model for Predicting the Pathogenicity of Missense Variants

Lai, J. — 2021-05-11

Single amino acid variations (SAVs) are a primary contributor to variations in the human genome. Identifying pathogenic SAVs can aid in the diagnosis and understanding of the genetic architecture of complex diseases, such as cancer. Most approaches for predicting the functional effects or pathogenicity of SAVs rely on either sequence or structural information. Nevertheless, previous analyses have shown that methods that depend on only sequence or structural information may have limited accuracy. Recently, researchers have attempted to increase the accuracy of their predictions by incorporating protein dynamics into pathogenicity predictions. This study presents < Lai Yang Rubenstein Uzun Sarkar > (LYRUS), a machine learning method that uses an XGBoost classifier selected by TPOT to predict the pathogenicity of SAVs. LYRUS incorporates five sequence-based features, six structure-based features, and four dynamics-based features. Uniquely, LYRUS includes a newly-proposed sequence co-evolution feature called variation number. LYRUSs performance was evaluated using a dataset that contains 4,363 protein structures corresponding to 20,307 SAVs based on human genetic variant data from the ClinVar database. Based on our dataset, the LYRUS classifier has a higher accuracy, specificity, F-measure, and Matthews correlation coefficient (MCC) than alternative methods including PolyPhen2, PROVEAN, SIFT, Rhapsody, EVMutation, MutationAssessor, SuSPect, FATHMM, and MVP. Variation numbers used within LYRUS differ greatly between pathogenic and neutral SAVs, and have a high feature weight in the XGBoost classifier employed by this method. Applications of the method to PTEN and TP53 further corroborate LYRUSs strong performance. LYRUS is freely available and the source code can be found at https://github.com/jiaying2508/LYRUS.

Neural cell injury pathology due to high-rate mechanical loading

Estrada, J. B. — 2021-05-13

Successful detection and prevention of brain injuries relies on the quantitative identification of cellular injury thresholds associated with the underlying pathology. Here, by combining a recently developed inertial microcavitation rheology technique with a 3D in vitro neural tissue model, we quantify and resolve the structural pathology and critical injury strain thresholds of neural cells occurring at high loading rates such as encountered in blast, cavitation or directed energy exposures. We find that neuronal dendritic spines characterized by MAP2 displayed the lowest physical failure strain at 7.3%, whereas microtubules and filamentous actin were able to tolerate appreciably higher strains (14%) prior to injury. Interestingly, while these critical injury thresholds were similar to previous literature values reported for moderate and lower strain rates (< 100 1/s), the pathology of primary injury reported here was distinctly different by being purely physical in nature as compared to biochemical activation during apoptosis or necrosis. TeaserControlled microcavitation enables quantitative identification of injury thresholds in neural cells.

Uncoupling of Chromatin Assembly from DNA Replication in Sciara Reveals a Domain of Postreplicative Immature Chromatin

Urnov, F. D. — 2021-05-12

DNA replication in dividing eukaryotic cells imposes a requirement for the faithful recreation on the newly synthesized chromatids of the nucleoprotein architecture of parent chromosomes. Practically nothing is known about the structure of postreplicative immature chromatin--a very short-lived entity (< 30 min.). We report here the unexpected discovery that during DNA amplification of locus II/9A in salivary gland polytene chromosomes of the fungus fly Sciara coprophila, DNA replication fork passage is uncoupled from postreplicative chromatin assembly; this enables visualization and analysis of chromatin fibers disassembled by DNA replication. We used electron microscopy to visualize a wealth of low nucleosome density immature chromatin fibers in preparations of Sciara chromatin from amplification-stage tissue. Remarkably, as gauged by high sensitivity to micrococcal nuclease and an unusually short length of DNA associated with each histone octamer, we found that locus II/9A which undergoes amplification and is replicated once every 4-6 hrs.--but not the bulk genome or a replicatively quiescent DNA stretch--was maintained in such an ummature fiber for ca. 24 hrs. Following amplification, locus II/9A assumed conventional chromatin organization, indicating that the epigenetic mark targeting nascent DNA to the chromatin assembly machinery is stable for several hours. We propose that this very unusual prolonged maintenance of a segment of the genome in immature chromatin facilitates access by the basal transcriptional machinery to the amplified DNA, and thus is an evolutionary adaptation to the demand for high transcription from genes that reside in the amplified loci.

NEOage clocks - Epigenetic clocks to estimate post-menstrual and postnatal age in preterm infants

Graw, S. — 2021-05-18

Epigenetic clocks based on DNA methylation (DNAm) can to accurately predict chronological age and are thought to capture biological aging. A variety of epigenetic clocks have been developed for different tissue types and age ranges, but none has focused on age prediction for preterm infants. Epigenetic estimators of biological age might be especially informative in epidemiologic studies of neonates, particularly those born preterm, since this is a key developmental window. Neonatal DNAm is dynamic and preterm infants are at heightened risk of developmental impairments. We aimed to fill this gap by developing epigenetic clocks for neonatal aging in preterm infants. As part of the Neonatal Neurobehavior and Outcomes in Very Preterm Infants (NOVI) study, buccal cells were collected at NICU discharge to profile DNAm levels in 542 very preterm infants. We applied elastic net regression to identify four epigenetic clocks (NEOage) predictive of post-menstrual and postnatal age, compatible with the Illumina EPIC and 450K arrays. We observed high correlations between predicted and reported ages (0.93 - 0.94) with root mean squared errors (1.28 - 1.63 weeks). Epigenetic estimators of neonatal aging in preterm infants can be useful tools to evaluate biological maturity and associations with neonatal and long-term morbidities.

Paternal chromosome elimination and X non-disjunction on asymmetric spindles in Sciara male meiosis

de Saint Phalle, B. — 2021-05-14

Meiosis in male Sciara is unique with a single centrosome. A monopolar spindle forms in meiosis I, but a bipolar spindle forms in meiosis II. The imprinted paternal chromosomes are eliminated in meiosis I; there is non-disjunction of the X in meiosis II. Despite differences in spindle construction and chromosome behavior, both meiotic divisions are asymmetric, producing a cell and a small bud. Observations of live spermatocytes made with the LC-PolScope, differential interference contrast optics and fluorescence revealed maternal and paternal chromosome sets on the monopolar spindle in meiosis I and formation of an asymmetric monastral bipolar spindle in meiosis II where all chromosomes except the X congress to the metaphase plate. The X remains near the centrosome after meiosis I and stays with it as the spindle forms in meiosis II. Electron microscopy revealed amorphous material between the X and the centrosome. Immunofluorescence with an antibody against the checkpoint protein Mad2 stains the centromeres of the maternal X dyad in late meiosis I and in meiosis II where it fails to congress to the metaphase plate. Mad2 is also present throughout the paternal chromosomes destined for elimination in meiosis I, suggesting a possible role in chromosome imprinting. If Mad2 on the X dyad mediates a spindle checkpoint in meiosis II, it may delay metaphase to facilitate formation of the second half spindle through a non-centrosomal mechanism.

Density Fluctuations Yield Distinct Growth and Fitness Effects in Single Bacteria

Nemati, S. — 2021-05-17

Single-cells grow by increasing their biomass and size. Here, we report that while mass and size accumulation rates of single Escherichia coli cells are exponential, their density fluctuates during growth. As such, the rates of mass and size accumulation of a single-cell are generally not the same, but rather cells differentiate into increasing one rate with respect to the other. This differentiation yields a previously unknown density homeostasis mechanism, which we support mathematically. Further, growth differentiation challenges ongoing efforts to predict single-cell reproduction rates (or fitness-levels), through the accumulation rates of size or mass. In contrast, we observe that density fluctuations can predict fitness, with only high fitness individuals existing in the high density fluctuation regime. We detail our imaging approach and the invisible microfluidic arrays that critically enabled increased precision and throughput. Biochemical production, infections, and natural communities start from few, growing, cells, thus, underscoring the significance of density-fluctuations when considering non-genetic variability.

Polarized Dishevelled dissolution and condensation drives embryonic axis specification in oocytes

Swartz, Z. — 2021-05-18

The organismal body axes that are formed during embryogenesis are intimately linked to intrinsic asymmetries established at the cellular scale in oocytes [1]. Here, we report an axis-defining event in meiotic oocytes of the sea star Patiria miniata. Dishevelled is a cytoplasmic Wnt pathway effector required for axis development in diverse species [2-4], but the mechanisms governing its function and distribution remain poorly defined. Using time-lapse imaging, we find that Dishevelled localizes uniformly to puncta throughout the cell cortex in Prophase I-arrested oocytes, but becomes enriched at the vegetal pole following meiotic resumption through a dissolution-condensation mechanism. This process is driven by an initial disassembly phase of Dvl puncta, followed by selective reformation of Dvl assemblies at the vegetal pole. Rather than being driven by Wnt signaling, this localization behavior is coupled to meiotic cell cycle progression and influenced by Lamp1+ endosome association and Frizzled receptors pre-localized within the oocyte cortex. Our results reveal a cell cycle-linked mechanism by which maternal cellular polarity is transduced to the embryo through spatially-regulated Dishevelled dynamics.

Genetic structure correlates with ethnolinguistic diversity in eastern and southern Africa

Atkinson, E. G. — 2021-05-19

African populations are the most diverse in the world yet are sorely underrepresented in medical genetics research. Here, we examine the structure of African populations using genetic and comprehensive multigenerational ethnolinguistic data from the Neuropsychiatric Genetics of African Populations-Psychosis study (NeuroGAP-Psychosis) consisting of 900 individuals from Ethiopia, Kenya, South Africa, and Uganda. We find that self-reported language classifications meaningfully tag underlying genetic variation that would be missed with consideration of geography alone, highlighting the importance of culture in shaping genetic diversity. Leveraging our uniquely rich multi-generational ethnolinguistic metadata, we track language transmission through the pedigree, observing the disappearance of several languages in our cohort as well as notable shifts in frequency over three generations. We find suggestive evidence for the rate of language transmission in matrilineal groups having been higher than that for patrilineal ones. We highlight both the diversity of variation within the African continent, as well as how within-Africa variation can be informative for broader variant interpretation; many variants appearing rare elsewhere are common in parts of Africa. The work presented here improves the understanding of the spectrum of genetic variation in African populations and highlights the enormous and complex genetic and ethnolinguistic diversity within Africa.

Differentially methylated regions and methylation QTLs for teen depression and early puberty in the Fragile Families Child Wellbeing Study

De Vito, R. — 2021-05-21

AO_SCPLOWBSTRACTC_SCPLOWThe Fragile Families Child Wellbeing Study (FFCWS) is a longitudinal cohort of ethnically diverse and primarily low socioeconomic status children and their families in the U.S. Here, we analyze DNA methylation data collected from 748 FFCWS participants in two waves of this study, corresponding to participant ages 9 and 15. Our primary goal is to leverage the DNA methylation data from these two time points to study methylation associated with two key traits in adolescent health that are over-represented in these data: Early puberty and teen depression. We first identify differentially methylated regions (DMRs) for depression and early puberty. We then identify DMRs for the interaction effects between these two conditions and age by including interaction terms in our regression models to understand how age-related changes in methylation are influenced by depression or early puberty. Next, we identify methylation quantitative trait loci (meQTLs) using genotype data from the participants. We also identify meQTLs with epistatic effects with depression and early puberty. We find enrichment of our interaction meQTLs with functional categories of the genome that contribute to the heritability of co-morbid complex diseases. We replicate our meQTLs in data from the GoDMC study. This work leverages the important focus of the FFCWS data on disadvantaged children to shed light on the methylation states associated with teen depression and early puberty, and on how genetic regulation of methylation is affected in adolescents with these two conditions.

Correction of a pathogenic mutation in iPSCs derived from a patient with Christianson syndrome using CRISPR/Cas9 genome editing

Ma, L. — 2021-05-20

SLC9A6 (also termed NHE6) encodes the endosomal Na+/H+ exchanger 6 (NHE6). Pathogenic, loss-of-function mutations in NHE6 cause the X-linked neurogenetic disorder Christianson syndrome (CS). We developed induced pluripotent stem cell (iPSC) lines derived from a patient with CS and from a biologically related control. The patient with CS contained the nonsense mutation c.1569G>A (p.(W523X)), which caused a significant reduction in NHE6 mRNA and a lack of detectable NHE6 protein in CS iPSCs in comparison to control iPSCs. To establish a cell model for study of CS with an isogenic control, we corrected the c.1569G>A mutation to the NHE6 reference genome sequence using CRISPR/Cas9-mediated homology directed repair knock-in methodology. Multiple subclonal lines were generated, and notably, NHE6 protein was expressed in all analyzed c.1569G>A (p.(W523X)) genome-corrected iPSC lines. This CS iPSC model together with the associated biologically related and isogenic control cell lines will serve as a valuable resource for both basic and translational studies in CS.

AltumAge: A Pan-Tissue DNA-Methylation Epigenetic Clock Based on Deep Learning

Paulo de Lima, L. — 2021-06-01

Several age predictors based on DNA methylation, dubbed epigenetic clocks, have been created in recent years, with the vast majority based on regularized linear regression. This study explores the improvement in the performance and interpretation of epigenetic clocks using deep learning. First, we gathered 143 publicly available data sets from several human tissues to develop AltumAge, a neural network framework that is a highly accurate and precise age predictor. Compared to ElasticNet, AltumAge performs better for within-data set and cross-data set age prediction, being particularly more generalizable in older ages and new tissue types. We then used deep learning interpretation methods to learn which methylation sites contributed to the final model predictions. We observe that while most important CpG sites are linearly related to age, some highly-interacting CpG sites can influence the relevance of such relationships. Using chromatin annotations, we show that the CpG sites with the highest contribution to the model predictions were related to gene regulatory regions in the genome, including proximity to CTCF binding sites. We also found age-related KEGG pathways for genes containing these CpG sites. Lastly, we perform downstream analyses of AltumAge to explore its applicability and compare its age acceleration with Horvaths model. We show that our neural network approach predicts higher age acceleration for tumors and for cells that exhibit age-related changes in vitro, such as replicative senescence and mitochondrial dysfunction. Altogether, our neural network approach provides significant improvement and flexibility to current epigenetic clocks for both performance and model interpretability.

A subset of CB002 xanthine analogues bypass p53-signaling to restore a p53 transcriptome and target an S-phase cell cycle checkpoint in tumors with mutated-p53

Hernandez Borrero, L. — 2021-06-02

Mutations in TP53 occur commonly in the majority of human tumors and confer aggressive tumor phenotypes including metastasis and therapy resistance. CB002 and structural-analogues restore p53 signaling in tumors with mutant-p53 but we find that unlike other xanthines such as caffeine, pentoxifylline, and theophylline, they do not deregulate the G2-checkpoint. Novel CB002-analogues induce pro-apoptotic Noxa protein in an ATF3/4-dependent manner, whereas caffeine, pentoxifylline, and theophylline do not. By contrast to caffeine, CB002-analogues target an S-phase checkpoint associated with increased p-RPA/RPA2, p-ATR, decreased Cyclin A, p-histone H3 expression and downregulation of essential proteins in DNA-synthesis and -repair. CB002-analogue #4 enhances cell death, and decreases Ki-67 in patient-derived tumor-organoids without toxicity to normal human cells. Preliminary in vivo studies demonstrate anti-tumor efficacy in mice. Thus, a novel class of anti-cancer drugs show activation of p53 pathway signaling in tumors with mutated p53, and target an S-phase checkpoint.

How do packet losses affect measures of averaged neural signals?

Dastin-van Rijn, E. — 2021-06-03

Recent advances in implanted device development have enabled chronic streaming of neural data to external devices allowing for long timescale, naturalistic recordings. However, characteristic data losses occur during wireless transmission. Estimates for the duration of these losses are typically uncertain reducing signal quality and impeding analyses. To characterize the effect of these losses on recovery of averaged neural signals, we simulated neural time series data for a typical event-related potential (ERP) experiment. We investigated how the signal duration and the degree of timing uncertainty affected the offset of the ERP, its duration in time, its amplitude, and the ability to resolve small differences corresponding to different task conditions. Simulations showed that long timescale signals were generally robust to the effects of packet losses apart from timing offsets while short timescale signals were significantly delocalized and attenuated. These results provide clarity on the types of signals that can be resolved using these datasets and provide clarity on the restrictions imposed by data losses on typical analyses.

Proprioceptive Genes as a Source of Genetic Variation Underlying Robustness for Flight Performance in Drosophila

Spierer, A. N. — 2021-06-03

A central challenge of quantitative genetics is partitioning phenotypic variation into genetic and non-genetic components. These non-genetic components are usually interpreted as environmental effects; however, variation between genetically identical individuals in a common environment can still exhibit phenotypic variation. A traits resistance to variation is called robustness, though the genetics underlying it are poorly understood. Accordingly, we performed an association study on a previously studied, whole organism trait: robustness for flight performance. Using 197 of the Drosophila Genetic Reference Panel (DGRP) lines, we surveyed variation across single nucleotide polymorphisms, whole genes, and epistatic interactions to find genetic modifiers robustness for flight performance. There was an abundance of genes involved in the development of sensory organs and processing of external stimuli, supporting previous work that processing proprioceptive cues is important for affecting variation in flight performance. Additionally, we tested insertional mutants for their effect on robustness using candidate genes found to modify flight performance. These results suggest several genes involved in modulating a trait mean are also important for affecting trait variance, or robustness, as well. Article SummaryWe sought to understand the genetic architecture of robustness (variation in a trait caused by non-genetic factors) for flight performance. We used 197 Drosophila Genetic Reference Panel (DGRP) lines to find significant individual variants and pairs of epistatic interactions, many of which were involved in proprioception. Additionally, we validated significant genes identified from a prior study for the mean of flight performance, showing genes affecting trait means may also affect trait robustness.

Rat anterior cingulate cortex continuously signals decision variables in a patchforaging task

Kane, G. A. — 2021-06-08

In patch foraging tasks, animals must decide whether to remain with a depleting resource or to leave it in search of a potentially better source of reward. In such tasks, animals consistently follow the general predictions of optimal foraging theory (the Marginal Value Theorem; MVT): to leave a patch when the reward rate in the current patch depletes to the average reward rate across patches. Prior studies implicate an important role for the anterior cingulate cortex (ACC) in foraging decisions based on MVT: within single trials, ACC activity increases immediately preceding foraging decisions, and across trials, these dynamics are modulated as the value of staying in the patch depletes to the average reward rate. Here, we test whether these activity patterns reflect dynamic encoding of decision-variables and whether these signals are directly involved in decision-making. We developed a leaky accumulator model based on the MVT that generates estimates of decision variables within and across trials, and tested model predictions against ACC activity recorded from rats performing a patch foraging task. Model predicted changes in MVT decision variables closely matched rat ACC activity. Next, we pharmacologically inactivated ACC to test the contribution of these signals to decision-making. ACC inactivation had a profound effect on rats foraging decisions and response times yet rats still followed the MVT decision rule. These findings suggest that the ACC encodes foraging-related variables for reasons unrelated to patch-leaving decisions.

Unified classification of mouse retinal ganglion cells using function, morphology, and gene expression

Goetz, J. — 2021-06-11

Classification and characterization of neuronal types are critical for understanding their function and dysfunction. Neuronal classification schemes typically rely on measurements of electrophysiological, morphological, and molecular features, but aligning such datasets has been challenging. Here, we present a unified classification of mouse retinal ganglion cells (RGCs), the sole retinal output neurons. We used visually-evoked responses to classify 1859 mouse RGCs into 42 types. We also obtained morphological or transcriptomic data from subsets and used these measurements to align the functional classification to publicly available morphological and transcriptomic data sets. We created an online database that allows users to browse or download the data and to classify RGCs from their light responses using a machine learning algorithm. This work provides a resource for studies of RGCs, their upstream circuits in the retina, and their projections in the brain, and establishes a framework for future efforts in neuronal classification and open data distribution.

Cross-species metabolomic analysis of DDT and Alzheimer's disease-associated tau toxicity

Kalia, V. — 2021-06-15

BackgroundThe formation of hyperphosphorylated tau (p-tau) protein tangles in neurons is a pathological marker of Alzheimers disease (AD). Exposure to the pesticide dichlorodiphenyltrichloroethane (DDT) has been associated with increased risk of AD. ObjectivesTo determine if there was a connection between DDT exposure and tau toxicity we investigated whether exposure to DDT can exacerbate tau protein toxicity in C. elegans. In addition, we examined the association between p-tau protein and metabolism in a human population study and in a transgenic C. elegans strain neuronally expressing a mutant tau protein fragment that is prone to aggregation. MethodsIn the human population study, we used a metabolome-wide association framework to determine the association between p-tau measured in the cerebrospinal fluid (CSF) and metabolomic features measured in both plasma (n = 142) and CSF (n = 78) using high-resolution metabolomics (HRM). Using the same HRM method, we determined changes in metabolomic features in the transgenic C. elegans strain compared to its control strain. Metabolites associated with p-tau in both species were analyzed for overlap. We also examined the effect of DDT and aggregating tau protein on growth, swim behavior, mitochondrial function, metabolism, learning, and lifespan in C. elegans. ResultsPlasma and CSF-derived features associated with p-tau level were related to drug, amino acid, fatty acid and mitochondrial metabolism pathways. Five metabolites overlapped between plasma and C. elegans, and 4 between CSF and C. elegans. DDT exacerbated the inhibitory effect of aggregating tau protein on growth and basal respiration. In the presence of aggregating tau protein, DDT induced more curling and was associated with reduced levels of amino acids but increased levels of uric acid and adenosylselenohomocysteine. Developmental exposure to DDT blunted the lifespan reduction caused by aggregating tau protein. ConclusionThe model organism C. elegans can complement human studies by providing a means to study mechanisms of environmental toxicants. Specifically, our C. elegans data show that DDT exposure and tau protein aggregation both inhibit mitochondrial function and DDT exposure can exacerbate the mitochondrial inhibitory effects of tau protein aggregation providing a plausible explanation for the observed human associations.

15 years of introgression studies: quantifying gene flow across Eukaryotes

Dagilis, A. J. — 2021-06-16

With the rise of affordable next generation sequencing technology, introgression -- or the exchange of genetic materials between taxa -- is widely perceived to be a ubiquitous phenomenon in nature. Although this claim is supported by several keystone studies, no thorough assessment on the frequency of introgression in nature has been performed to date. In this manuscript, we aim to address this knowledge gap by providing a meta-analysis of the most comprehensive survey of introgression studies in Eukaryotes to date (724 papers with claims of introgression). We first examined the evidence given to support introgression, and if/how the lines of evidence have changed across time. We then collated a single statistic, Pattersons D, that quantifies the strength of introgression across 123 studies to further assess how taxonomic group, divergence time, and aspects of life history influence introgression. We find three main results. Studies on introgression are much more frequent in plants and mammals than any other taxonomic group. The study of introgression has shifted from a largely qualitative assessment of whether introgression happens, to a focus on when and how much introgression has occurred across taxa. The most often used introgression statistic, Pattersons D, shows several intriguing patterns suggesting introgression reports may be biased by both differences in reporting criteria and sequencing technology, but may also differ across taxonomic systems and throughout the process of speciation. Together, these results suggest the need for a unified approach to quantifying introgression in natural communities, and highlight important areas of future research that can be better assessed once this unified approach is met.

Behavioral and nociceptor states of inflammatory painacross timescales in 2D and 3D

Bohic, M. — 2021-06-17

Ongoing pain is often driven by direct activation of pain-sensing neurons and neuroimmune mediated sensitization. These heightened states of pain alter physiology, reduce motor function, and alter motivation to engage in normal behaviors. The complexity of the pain state has evaded a comprehensive definition, especially in nonverbal animals. Here in mice, we capture the physiological state of sensitized pain neurons at different time points post-inflammation and used computational tools to automatically map behavioral signatures of evoked and spontaneous displays of pain. First, retrograde labeling coupled with electrophysiology of neurons innervating the site of localized inflammation defined critical time points of pain sensitization. Next, we used high-speed videography combined with supervised and unsupervised machine learning tools and uncovered sensory-evoked defensive coping postures to pain. Using 3D pose analytics inspired by natural language processing, we identify movement sequences that correspond to robust representations of ongoing pain states. Surprisingly, with this analytical framework, we find that a commonly used anti-inflammatory painkiller does not return an animals behavior back to a pre-injury state. Together, these findings reveal the previously unidentified signatures of pain and analgesia at timescales when inflammation induces heightened pain states.

Single Cell Transcriptomics Reveals Global Markers of Transcriptional Diversity Across Different Forms of CellularSenescence

Evans, S. A. — 2021-06-17

Cellular Senescence is a state of irreversible cell cycle arrest, and the accumulation of senescent cells contributes to age- related organismal decline. The detrimental effects of cellular senescence are due to the senescence associated secretory phenotype (SASP), an array of signaling molecules and growth factors secreted by senescent cells that contribute to the sterile inflammation associated with aging tissues. Recent studies, both in vivo and in vitro, have highlighted the heterogeneous nature of the senescence phenotype. In particular, single cell transcriptomics has revealed that Oncogene Induced Senescence (OIS) is characterized by the presence of subpopulations of cells expressing different SASP profiles. We have generated a comprehensive dataset via single-cell transcriptional profiling of genetically homogenous clonal cell lines from different forms of senescence, including OIS, Replicative Senescence (RS), and DNA Damage Induced Senescence (DDIS). We identified subpopulations of cells that are common to all three major forms of senescence and show that the expression profiles of these subpopulations are driven by markers formerly identified in individual forms of senescence. These common signatures are characterized by chromatin modifiers, inflammation, extracellular matrix remodeling, and Ribosomal protein expression. The expression patterns of these subpopulations recapitulate primary and secondary senescence, a phenomenon where a preexisting (primary) senescent cell induces senescence in a neighboring (secondary) cell through cell-to-cell contact. Since it is still unclear what type of senescence occurs in-vivo with age, it is important to know that the formation of primary and secondary populations is common to multiple types of senescence since this mechanism could help explain how senescent cells accumulate in aged organisms. Finally, we show that these subpopulations show differential susceptibility to the senolytic agent Navitoclax, suggesting that senolytic agents targeting the apoptotic pathways may be clearing only a subset of senescent cells based on their inflammatory profiles in-vivo.

Dynamic interplay between reward and voluntary attention determines stimulus processing in visual cortex

Grahek, I. — 2021-06-17

Reward enhances stimulus processing in the visual cortex, but the mechanisms through which this effect occurs remain unclear. Reward prospect can both increase the deployment of voluntary attention and increase the salience of previously neutral stimuli. In this study we orthogonally manipulated reward and voluntary attention while human participants performed a global motion detection task. We recorded steady-state visual evoked potentials (SSVEPs) to simultaneously measure the processing of attended and unattended stimuli linked to different reward probabilities, as they compete for attentional resources. The processing of the high rewarded feature was enhanced independently of voluntary attention, but this gain diminished once rewards were no longer available. Neither the voluntary attention nor the salience account alone can fully explain these results. Instead, we propose how these two accounts can be integrated to allow for the flexible balance between reward-driven increase in salience and voluntary attention.

Large-scale neural recordings with single-cell resolution in human cortex using high-density Neuropixels probes

Paulk, A. C. — 2021-06-21

Recent advances in multi-electrode array technology have made it possible to monitor large neuronal ensembles at cellular resolution. In humans, however, current approaches either restrict recordings to only a few neurons per penetrating electrode or combine the signals of thousands of neurons in local field potential (LFP) recordings. Here, we describe a new probe variant and set of techniques which enable simultaneous recording from over 200 well-isolated cortical single units in human participants during intraoperative neurosurgical procedures using silicon Neuropixels probes. We characterized a diversity of extracellular waveforms with eight separable single unit classes, with differing firing rates, positions along the length of the linear electrode array, spatial spread of the waveform, and modulation by LFP events such as inter-ictal discharges and burst suppression. While some additional challenges remain in creating a turn-key recording system, high-density silicon arrays provide a path for studying human-specific cognitive processes and their dysfunction at unprecedented spatiotemporal resolution.

Multivalent interactions with RNA drive RNA binding protein recruitment and dynamics in biomolecular condensates in Xenopus oocytes

Cabral, S. E. — 2021-06-21

RNA localization and biomolecular condensate formation are key biological strategies for organizing the cytoplasm and generating cellular and developmental polarity. While enrichment of RNAs and RNA-binding proteins (RBPs) is a hallmark of both processes, the functional and structural roles of RNA-RNA and RNA-protein interactions within condensates remain unclear. Recent work from our laboratory has shown that RNAs required for germ layer patterning in Xenopus oocytes localize in novel biomolecular condensates, termed Localization bodies (L-bodies). L-bodies are composed of a non-dynamic RNA phase enmeshed in a more dynamic protein-containing phase. However, the interactions that drive the biophysical characteristics of L-bodies are not known. Here, we test the role of RNA-protein interactions using an L-body RNA-binding protein, PTBP3, which contains four RNA-binding domains (RBDs). We find that binding of RNA to PTB is required for both RNA and PTBP3 to be enriched in L-bodies in vivo. Importantly, while RNA binding to a single RBD is sufficient to drive PTBP3 localization to L-bodies, interactions between multiple RRMs and RNA tunes the dynamics of PTBP3 within L-bodies. In vitro, recombinant PTBP3 phase separates into non-dynamic structures in an RNA-dependent manner, supporting a role for RNA-protein interactions as a driver of both recruitment of components to L-bodies and the dynamics of the components after enrichment. Our results point to a model where RNA serves as a concentration-dependent, non-dynamic substructure and multivalent interactions with RNA are a key driver of protein dynamics.

Population structure and introgression among recently differentiated Drosophila melanogaster populations

Coughlan, J. M. — 2021-06-25

Despite a century of genetic analysis, the evolutionary history underlying patterns of exceptional genetic and phenotypic variation in the model organism Drosophila melanogaster remains poorly understood. How genetic and phenotypic variation is partitioned across the range of D. melanogaster, particularly in its putative ancestral range in Subtropical Africa, remains unresolved. Here, we assess patterns of population genetic structure, admixture, mate preference, and genetic incompatibility across a global sample, including 174 new accessions from remote regions within Subtropical Africa. While almost all Out of Africa genomes correspond to a single genetic ancestry, different geographic regions within Africa contain multiple ancestries, with substantial cryptic diversity in Subtropical Africa. Admixture between distinct lineages is prevalent across the range, but admixture rates vary between lineages. Female mate choice within Subtropical Africa is highly polymorphic and behavioral types are not monophyletic. The genetic architecture of mate choice is highly polygenic, including loci associated with neurological development, behavior, olfactory perception, and learning. Finally, we discovered that many segregating putative incompatibilities likely evolved during or after expansion out of Africa. This work contributes to our understanding of the evolutionary history of a key model system, and provides insight into the distribution of polymorphic reproductive barriers.

Candida albicans Isolates 529L and CHN1 Exhibit Stable Colonization of the Murine Gastrointestinal Tract

McDonough, L. — 2021-06-27

Candida albicans is a pathobiont that colonizes multiple niches in the body including the gastrointestinal (GI) tract, but is also responsible for both mucosal and systemic infections. Despite its prevalence as a human commensal, the murine GI tract is generally refractory to colonization with the C. albicans reference isolate SC5314. Here, we identify two C. albicans isolates, 529L and CHN1, that stably colonize the murine GI tract in three different animal facilities under conditions where SC5314 is lost from this niche. Analysis of the bacterial microbiota did not show notable differences between mice colonized with the three C. albicans strains. We compared the genotypes and phenotypes of these three strains and identified thousands of SNPs and multiple phenotypic differences, including their ability to grow and filament in response to nutritional cues. Despite striking filamentation differences under laboratory conditions, however, analysis of cell morphology in the GI tract revealed that the three isolates exhibited similar filamentation properties in this in vivo niche. Notably, we found that SC5314 is more sensitive to the antimicrobial peptide CRAMP, and the use of CRAMP-deficient mice increased the ability of SC5314 to colonize the GI tract relative to CHN1 and 529L. These studies provide new insights into how strain-specific differences impact C. albicans traits in the host and advance CHN1 and 529L as relevant strains to study C. albicans pathobiology in its natural host niche. IMPORTANCEUnderstanding how fungi colonize the GI tract is increasingly recognized as highly relevant to human health. The animal models used to study Candida albicans commensalism commonly rely on altering the host microbiome (via antibiotic treatment or defined diets) to establish successful GI colonization by the C. albicans reference isolate SC5314. Here, we characterize two C. albicans isolates that can colonize the murine GI tract without antibiotic treatment and can therefore be used as tools for studying fungal commensalism. Importantly, experiments were replicated in three different animal facilities and utilized three different mouse strains. Differential colonization between fungal isolates was not associated with alterations in the bacterial microbiome but rather with distinct responses to CRAMP, a host antimicrobial peptide. This work emphasizes the importance of C. albicans intra-species variation as well as host anti-microbial defense mechanisms in defining commensal interactions.

Parametric modulators of sex-biased conditioned fear responding

Mitchell, J. R. — 2021-06-30

Pavlovian fear conditioning is a widely used behavioral paradigm for studying associative learning in rodents. Despite early recognition that subjects may engage in a variety of both conditioned and unconditioned responses, the last several decades have seen the field narrow its focus to measure freezing as the sole indicator of conditioned fear. We previously reported that female rats were more likely than males to engage in darting, an escape-like conditioned response that is associated with heightened shock reactivity. To determine how experimental parameters contribute to the frequency of darting in both males and females, we manipulated factors such as chamber size, shock intensity, and number of trials. To better capture fear-related behavioral repertoires in our animals, we developed ScaredyRat, an open-source custom Python tool that analyzes Noldus Ethovision-generated raw data files to identify Darters and quantify both conditioned and unconditioned responses. We find that like freezing, conditioned darting occurrences scale with experimental alterations. While most darting occurs in females, we find that with an extended training protocol, darting can emerge in males as well. Collectively, our data suggest that darting reflects a behavioral switch in conditioned responding that is a product of both an individual animals sex, shock reactivity, and experimental parameters, underscoring the need for careful consideration of sex as a biological variable in classic learning paradigms.

Dysregulated myogenesis and autophagy in genetically induced pulmonary emphysema

Balnis, J. — 2021-07-09

Patients with chronic obstructive pulmonary disease (COPD)-pulmonary emphysema often develop locomotor muscle dysfunction, which is independently associated with disability and higher mortality in that population. Muscle dysfunction entails reduced muscle mass and force-generation capacity, which are influenced by fibers integrity. Myogenesis, which is muscle turnover driven by progenitor cells such as satellite cells, contributes to the maintenance of muscle integrity in the context of organ development and injury-repair cycles. Injurious events crucially occur in COPD patients skeletal muscles in the setting of exacerbations and infections which lead to acute decompensations for limited periods of time after which, patients typically fail to recover the baseline status they had before the acute event. Autophagy, which is dysregulated in muscles from COPD patients, is a key regulator of satellite cells activation and myogenesis, yet very little research has so far investigated the mechanistic role of autophagy dysregulation in COPD muscles. Using a genetically inducible murine model of COPD-driven muscle dysfunction and confirmed with a second genetic animal model, we found a significant myogenic dysfunction associated with a reduced proliferative capacity of freshly isolated satellite cells. Transplantation experiments followed by lineage tracing suggest that an intrinsic defect in satellite cells, and not in the COPD environment, plays a dominant role in the observed myogenic dysfunction. RNA sequencing analysis of freshly isolated satellite cells suggests dysregulation of transcripts associated with control of cell cycle and autophagy, which is confirmed by a direct observation of COPD mice satellite cells fluorescent-tracked autophagosome formation. Moreover, spermidine-induced autophagy stimulation leads to improved satellite cells autophagosome turnover, replication rate and myogenesis. Our data suggests that pulmonary emphysema causes a disrupted myogenesis, which could be improved with stimulation of autophagy and satellite cells activation, leading to an attenuated muscle dysfunction in this context.

Lipopolysaccharide-induced neuroinflammation disrupts functional connectivity and community structure in primary cortical microtissues

Atherton, E. — 2021-07-09

Three-dimensional (3D) neural microtissues are a powerful in vitro paradigm for studying brain development and disease under controlled conditions, while maintaining many key attributes of the in vivo environment. Here, we used primary cortical microtissues to study the effects of neuroinflammation on neural microcircuits. We demonstrated the use of a genetically encoded calcium indicator combined with a novel live-imaging platform to record spontaneous calcium transients in microtissues from day 14-34 in vitro. We implemented graph theory analysis of calcium activity to characterize underlying functional connectivity and community structure of microcircuits, which are capable of capturing subtle changes in network dynamics during early diseases states. We found that microtissues cultured for 34 days displayed functional remodeling of microcircuits and that community structure strengthened over time. Lipopolysaccharide, a neuroinflammatory agent, significantly increased functional connectivity and disrupted community structure 5-9 days after exposure. These microcircuit-level changes have broad implications for the role of neuroinflammation in functional dysregulation of neural networks.

On the Apportionment of Archaic Human Diversity

Villanea, F. A. — 2021-07-16

The apportionment of human genetic diversity within and between populations has been measured to understand human relatedness and demographic history. Likewise, the distribution of archaic ancestry in modern populations can be leveraged to better understand the interaction between our species and its archaic relatives, and the impact of natural selection on archaic segments of the human genome. Resolving these interactions can be difficult, as archaic variants in modern populations have also been shaped by genetic drift, bottlenecks, and gene flow. Here, we investigate the apportionment of archaic variation in Eurasian populations. We find that archaic genome coverage at the individual- and population-level present unique patterns in modern human population: South Asians have an elevated count of population-unique archaic SNPs, and Europeans and East Asians have a higher degree of archaic SNP sharing, indicating that population demography and archaic admixture events had distinct effects in these populations. We confirm previous observations that East Asians have more Neanderthal ancestry than Europeans at an individual level, but surprisingly Europeans have more Neandertal ancestry at a population level. In comparing these results to our simulated models, we conclude that these patterns likely reflect a complex series of interactions between modern humans and archaic populations.

Cortical spheroids display oscillatory network dynamics

Sevetson, J. L. — 2021-07-18

Three-dimensional brain cultures can facilitate the study of central nervous system function and disease, and one of the most important components that they present is neuronal activity on a network level. Here we demonstrate network activity in rodent cortical spheroids while maintaining the networks intact in their 3D state. Networks developed by nine days in culture and became more complex over time. To measure network activity, we imaged neurons in rat and mouse spheroids labelled with a calcium indicator dye, and in mouse spheroids expressing GCaMP. Network activity was evident when we electrically stimulated spheroids, was abolished with glutamatergic blockade, and was altered by GABAergic blockade or partial glutamatergic blockade. We quantified correlations and distances between somas with micron-scale spatial resolution. Spheroids seeded at as few as 4,000 cells gave rise to emergent network events, including oscillations. These results are the first demonstration that self-assembled rat and mouse spheroids exhibit network activity consistent with in vivo network events. These results open the door to experiments on neuronal networks that require fewer animals and enable high throughput experiments on network-perturbing alterations in neurons and glia.

TAF4b transcription networks regulating early oocyte differentiation

Gura, M. A. — 2021-07-18

Establishment of a healthy ovarian reserve is contingent upon numerous regulatory pathways during embryogenesis. Previously, mice lacking TBP-associated factor 4b (Taf4b) were shown to exhibit a diminished ovarian reserve. However, potential oocyte-intrinsic functions of TAF4b have not been examined. Here we use a combination of gene expression profiling and chromatin mapping to characterize the TAF4b gene regulatory network in mouse oocytes. We find that Taf4b-deficient oocytes display inappropriate expression of meiotic, chromatin, and X-linked genes, and unexpectedly we found a connection with Turner Syndrome pathways. Using Cleavage Under Targets and Release Using Nuclease (CUT&RUN), we observed TAF4b enrichment at genes involved in meiosis and DNA repair, some of which are differentially expressed in Taf4b-deficient oocytes. Interestingly, TAF4b target genes were enriched for Sp/KLF family motifs rather than TATA-box, suggesting an alternate mode of promoter interaction. Together, our data connects several gene regulatory nodes that contribute to the ovarian reserve.

Hierarchical clustering optimizes the tradeoff between compositionality and expressivity of task structures in reinforcement learning

Liu, R. G. — 2021-07-21

A hallmark of human intelligence, but challenging for reinforcement learning (RL) agents, is the ability to compositionally generalise, that is, to recompose familiar knowledge components in novel ways to solve new problems. For instance, when navigating in a city, one needs to know the location of the destination and how to operate a vehicle to get there, whether it be pedalling a bike or operating a car. In RL, these correspond to the reward function and transition function, respectively. To compositionally generalize, these two components need to be transferable independently of each other: multiple modes of transport can reach the same goal, and any given mode can be used to reach multiple destinations. Yet there are also instances where it can be helpful to learn and transfer entire structures, jointly representing goals and transitions, particularly whenever these recur in natural tasks (e.g., given a suggestion to get ice cream, one might prefer to bike, even in new towns). Prior theoretical work has explored how, in model-based RL, agents can learn and generalize task components (transition and reward functions). But a satisfactory account for how a single agent can simultaneously satisfy the two competing demands is still lacking. Here, we propose a hierarchical RL agent that learns and transfers individual task components as well as entire structures (particular compositions of components) by inferring both through a non-parametric Bayesian model of the task. It maintains a factorised representation of task components through a hierarchical Dirichlet process, but it also represents different possible covariances between these components through a standard Dirichlet process. We validate our approach on a variety of navigation tasks covering a wide range of statistical correlations between task components and show that it can also improve generalisation and transfer in more complex, hierarchical tasks with goal/subgoal structures. Finally, we end with a discussion of our work including how this clustering algorithm could conceivably be implemented by cortico-striatal gating circuits in the brain.

The neuropeptide Pth2 modulates social behavior and anxiety in zebrafish

Anneser, L. — 2021-07-28

Animal behavior is strongly context-dependent and behavioral performance is often modulated by internal state. In particular, different social contexts can alter anxiety levels and modulate social behavior. The vertebrate-specific neuropeptide parathyroid hormone 2 (pth2) is directly regulated by the presence or absence of conspecifics in zebrafish. As its cognate receptor, the parathyroid hormone 2 receptor (pth2r), is widely expressed across the brain, we tested fish lacking the functional Pth2 peptide in several anxiety-related and social paradigms. Rodents lacking PTH2 display increased anxiety-related behavior. Here we show that the propensity to react to sudden stimuli with an escape response is increased in pth2-/- zebrafish, consistent with elevated anxiety. While overall social preference for conspecifics is maintained in pth2-/- fish until the early juvenile stage, we found that both social preference and shoaling are altered later in development. The data presented suggest that the neuropeptide Pth2 modulates several conserved behavioral features, and may thus enable the animal to react appropriately in different social contexts.

A Topological Data Analytic Approach for Discovering Biophysical Signatures in Protein Dynamics

Tang, W. S. — 2021-07-29

Identifying structural differences among proteins can be a non-trivial task. When contrasting ensembles of protein structures obtained from molecular dynamics simulations, biologically-relevant features can be easily overshadowed by spurious fluctuations. Here, we present SINATRA Pro, a computational pipeline designed to robustly identify topological differences between two sets of protein structures. Algorithmically, SINATRA Pro works by first taking in the 3D atomic coordinates for each protein snapshot and summarizing them according to their underlying topology. Statistically significant topological features are then projected back onto a user-selected representative protein structure, thus facilitating the visual identification of biophysical signatures of different protein ensembles. We assess the ability of SINATRA Pro to detect minute conformational changes in five independent protein systems of varying complexities. In all test cases, SINATRA Pro identifies known structural features that have been validated by previous experimental and computational studies, as well as novel features that are also likely to be biologically-relevant according to the literature. These results highlight SINATRA Pro as a promising method for facilitating the non-trivial task of pattern recognition in trajectories resulting from molecular dynamics simulations, with substantially increased resolution. Author SummaryStructural features of proteins often serve as signatures of their biological function and molecular binding activity. Elucidating these structural features is essential for a full understanding of underlying biophysical mechanisms. While there are existing methods aimed at identifying structural differences between protein variants, such methods do not have the capability to jointly infer both geometric and dynamic changes, simultaneously. In this paper, we propose SINATRA Pro, a computational framework for extracting key structural features between two sets of proteins. SINATRA Pro robustly outperforms standard techniques in pinpointing the physical locations of both static and dynamic signatures across various types of protein ensembles, and it does so with improved resolution.

A nanopore ion source delivers single amino acid and peptide ions directly into the gas phase

Drachman, N. — 2021-08-15

AbstractWe report a nanopore ion source for mass spectrometry that delivers ions directly into high vacuum from aqueous solutions. The ion source comprises a pulled quartz pipette with a sub-100 nm opening. Ions escape an electrified meniscus by ion evaporation and travel along collisionless trajectories to the ion detector. We measure mass spectra of 16 different amino acid ions, post-translationally modified variants of glutathione, and the peptide angiotensin II, showing that these analytes can be emitted as desolvated ions. The emitted current is composed of ions rather than charged droplets, and more than 90% of the current can be recovered in a distant collector. By circumventing the sample loss mechanisms inherent to conventional electrospray ionization (ESI), where charged droplets are sprayed into a background gas that scatters ions and degrades their transmission, the nanopore ion source could enable more sensitive proteomic analyses.

CREB5 reprograms nuclear interactions to promote resistance to androgen receptor targeting therapies

Hwang, J. H. — 2021-08-18

Metastatic castration resistant prostate cancers (mCRPC) are treated with therapies that antagonize the androgen receptor (AR). Nearly all patients develop resistance to AR-targeted therapies (ART). Our previous work identified CREB5 as an upregulated target gene in human mCRPC that promoted resistance to all clinically-approved ART. The mechanisms by which CREB5 promotes progression of mCRPC or other cancers remains elusive. Integrating ChIP-seq and rapid immunoprecipitation and mass spectroscopy of endogenous proteins (RIME), we report that cells overexpressing CREB5 demonstrate extensive reprogramming of nuclear protein-protein interactions in response to the ART agent enzalutamide. Specifically, CREB5 physically interacts with AR, the pioneering actor FOXA1, and other known co-factors of AR and FOXA1 at transcription regulatory elements recently found to be active in mCRPC patients. We identified a subset of CREB5/FOXA1 co-interacting nuclear factors that have critical functions for AR transcription (GRHL2, HOXB13) while others (TBX3, NFIC) regulated cell viability and ART resistance and were amplified or overexpressed in mCRPC. Upon examining the nuclear protein interactions and the impact of CREB5 expression on the mCRPC patient transcriptome, we found CREB5 was associated with TGF{beta} and Wnt signaling and epithelial to mesenchymal transitions, implicating these pathways in ART resistance. Overall, these observations define the molecular interactions among CREB5, FOXA1, and pathways that promote ART resistance.

Super-Resolution Electrochemical Impedance Imaging with a 100 x 100 CMOS Sensor Array

Hu, K. — 2021-08-19

This paper presents a 100 x 100 super-resolution integrated sensor array for microscale electrochemical impedance spectroscopy (EIS) imaging. The system is implemented in 180 nm CMOS with 10 m x 10 m pixels. Rather than treating each electrode independently, the sensor is designed to measure the mutual capacitance between programmable sets of pixels. Multiple spatially-resolved measurements can then be computationally combined to produce super-resolution impedance images. Experimental measurements of sub-cellular permittivity distributions within single algae cells demonstrate the potential of this new approach.

Redox-dependent Structure and Dynamics of Macrophage Migration Inhibitory Factor Reveal Sites of Latent Allostery

Skeens, E. — 2021-09-02

Macrophage migration inhibitory factor (MIF) is a multifunctional immunoregulatory protein that is a key player in the innate immune response. Given its overexpression at sites of inflammation in a wide range of diseases marked by increasingly oxidative cellular environment, a comprehensive structural understanding of how cellular redox conditions may impact the structure and function of MIF is necessary. We used solution NMR spectroscopy and mass spectrometry to investigate structural and dynamic signatures of MIF under varied solution redox conditions. Our results indicate that the MIF structure is modified and becomes increasingly dynamic in an oxidative environment, which may be a means to alter the MIF functional response in a redox-dependent manner. We identified latent allosteric sites within MIF that are redox-sensitive and mutational analysis reveals that loss of redox-responsive residues attenuates activation of the coreceptor CD74. Leveraging sites of redox-sensitivity therefore reveals an avenue to modulate MIF function in its "disease state" via structure-based drug design.

The visual coupling between neighbors explains 'flocking' in human crowds

Dachner, G. C. — 2021-09-06

Patterns of collective motion or flocking in birds, fish schools, and human crowds are believed to emerge from local interactions between individuals. Most models of collective motion attribute these interactions to hypothetical rules or forces, often inspired by physical systems, and described from an overhead view. We develop a visual model of human flocking from an embedded view, based on optical variables that actually govern pedestrian interactions. Specifically, people control their walking speed and direction by canceling the average optical expansion and angular velocity of their neighbors, weighted by visual occlusion. We test the model by simulating data from experiments with virtual crowds and real human swarms. The visual model outperforms our previous overhead model and explains basic properties of physics-inspired models: repulsion forces reduce to canceling optical expansion, attraction forces to canceling optical contraction, and alignment to canceling the combination of expansion/contraction and angular velocity. Critically, the neighborhood of interaction follows from Euclids Law of perspective and the geometry of occlusion. We conclude that the local interactions underlying human flocking are a natural consequence of the laws of optics. Similar principles may apply to collective motion in other species.

Evidence for a selective link between cooperation and individual recognition

Tumulty, J. P. — 2021-09-08

The ability to recognize and discriminate among others is a frequent assumption of models of the evolution of cooperative behavior. At the same time, cooperative behavior has been proposed as a selective agent favoring the evolution of individual recognition abilities. While theory predicts that recognition and cooperation may co-evolve, data linking recognition abilities and cooperative behavior with fitness or evidence of selection are elusive. Here, we provide evidence of a fitness link between individual recognition and cooperation in the paper wasp Polistes fuscatus. Nest founding females in northern populations frequently form cooperative multiple foundress nests and possess highly variable facial patterns that mediate individual recognition. We describe a dearth of cooperative nesting, low phenotypic diversity, and a lack of individual recognition in southern populations. In a common garden experiment, northern co-foundress associations successfully reared offspring while all cooperative southern groups failed to rear any offspring, suggesting a fitness link between individual recognition and successful cooperation. Consistent with a selective link between individual recognition and cooperation, we find that rates of cooperative co-nesting correlate with identity-signaling color pattern diversity across the species range. Moreover, genomic evidence of recent positive selection on cognition loci likely to mediate individual recognition is substantially stronger in northern compared to southern P. fuscatus populations. Collectively, these data suggest that individual recognition and cooperative nesting behavior have co-evolved in P. fuscatus because recognition helps mediate conflict among co-nesting foundresses. This work provides evidence of a specific cognitive phenotype under selection because of social interactions, supporting the idea that social behavior can be a key driver of cognitive evolution.

Humans reconfigure target and distractor processing to address distinct task demands

Ritz, H. — 2021-09-10

When faced with distraction, we can focus more on goal-relevant information (targets) or focus less goal-conflicting information (distractors). How people use cognitive control to distribute attention across targets and distractors remains unclear. To help address this question, we developed a parametric attentional control task that can index both target discriminability and distractor interference. We find that participants exert independent control over target and distractor processing. We measured control adjustments through the influence of incentives and previous conflict on target and distractor sensitivity, finding that these have dissociable influences on control. Whereas incentives preferentially led to target enhancement, conflict on the previous trial preferentially led to distractor suppression. These distinct drivers of control altered sensitivity to targets and distractors early in the trial, promptly followed by reactive reconfiguration towards task-appropriate feature sensitivity. Finally, we provide a process-level account of these findings by showing that these control adjustments are well-captured by an evidence accumulation model with attractor dynamics over feature weights. These results help establish a process-level account of control reconfiguration that provides new insights into how multivariate attentional signals are optimized to achieve task goals.

A linear neural circuit for light avoidance in Drosophila larvae

Sorkac, A. — 2021-09-11

Understanding how neural circuits underlie behaviour is challenging even in the era of the connectome because it requires a combined approach encompassing anatomical and functional analyses. This is exemplified in studying the circuit underlying the light-avoidance behaviour displayed by the larvae of the fruit fly Drosophila melanogaster. While this behaviour is robust and the nervous system relatively simple, only bits and pieces of the circuit have been delineated1. Indeed, some studies resulted in contradicting conclusions regarding the contributions of various neuronal types to this behaviour2,3. Here we devise trans-Tango MkII, a new version of the transsynaptic circuit tracing and manipulation tool trans-Tango4. We implement trans-Tango MkII in anatomical tracing and combine it with circuit epistasis analysis. We use neuronal inhibition to test necessity of particular neuronal types for light-avoidance. We complement these experiments by selective neuronal activation to examine sufficiency in rescuing light-avoidance deficiencies exhibited by photoreceptor mutants. Together, our studies reveal a four-order, linear circuit for light-avoidance behaviour connecting the light-detecting photoreceptors with a pair of neuroendocrine cells via two types of clock neurons. Our combined approach could be readily expanded to other larval circuits. Further, this strategy provides the framework for studying more complex nervous systems and behaviours.

SILAC phosphoproteomics reveals unique signaling circuits in CAR-T cells and the inhibition of B cell-activating phosphorylation in target cells

Griffith, A. A. — 2021-09-10

Chimeric antigen receptor (CAR) is a single-pass transmembrane receptor designed to specifically target and eliminate cancers. While CARs prove highly efficacious against B cell malignancies, the intracellular signaling events which promote CAR T cell activity remain elusive. To gain further insight into both CAR T cell signaling and the potential signaling response of cells targeted by CAR, we analyzed phosphopeptides captured by two separate phopshoenrichment strategies from third generation CD19-CAR T cells cocultured with SILAC labeled Raji B cells by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Here, we report that CD19-CAR T cells upregulated several key phosphorylation events also observed in canonical T cell receptor (TCR) signaling while Raji B cells exhibited a significant decrease in B cell receptor-signaling related phosphorylation events in response to coculture. Our data suggest that CD19-CAR stimulation activates a mixture of unique CD19-CAR-specific signaling pathways and canonical TCR signaling while global phosphorylation in Raji B cells is reduced after association with the CD19-CAR T cells.

Protective Efficacy of Gastrointestinal SARS-CoV-2 Delivery Against Intranasal and Intratracheal SARS-CoV-2 Challenge in Rhesus Macaques

Yu, J. — 2021-09-14

Live oral vaccines have been explored for their protective efficacy against respiratory viruses, particularly for adenovirus serotypes 4 and 7. The potential of a live oral vaccine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), however, remains unclear. In this study, we assessed the immunogenicity of live SARS-CoV-2 delivered to the gastrointestinal tract in rhesus macaques and its protective efficacy against intranasal and intratracheal SARS-CoV-2 challenge. Post-pyloric administration of SARS-CoV-2 by esophagogastroduodenoscopy resulted in limited virus replication in the gastrointestinal tract and minimal to no induction of mucosal antibody titers in rectal swabs, nasal swabs, and bronchoalveolar lavage. Low levels of serum neutralizing antibodies were induced and correlated with modestly diminished viral loads in nasal swabs and bronchoalveolar lavage following intranasal and intratracheal SARS-CoV-2 challenge. Overall, our data show that post-pyloric inoculation of live SARS-CoV-2 is weakly immunogenic and confers partial protection against respiratory SARS-CoV-2 challenge in rhesus macaques. ImportanceSARS-CoV-2 remains a global threat, despite the rapid deployment but limited coverage of multiple vaccines. Alternative vaccine strategies that have favorable manufacturing timelines, greater ease of distribution and improved coverage may offer significant public health benefits, especially in resource-limited settings. Live oral vaccines have the potential to address some of these limitations; however no studies have yet been conducted to assess the immunogenicity and protective efficacy of a live oral vaccine against SARS-CoV-2. Here we report that oral administration of live SARS-CoV-2 in non-human primates may offer prophylactic benefits, but that formulation and route of administration will require further optimization.

Enhanced Specificity Mutations Perturb Allosteric Signaling in CRISPR-Cas9

Palermo, G. — 2021-09-14

CRISPR-Cas9 is a molecular tool with transformative genome editing capabilities. At the molecular level, an intricate allosteric signaling is critical for DNA cleavage, but its role in the specificity enhancement of the Cas9 endonuclease is poorly understood. Here, solution NMR is combined with multi-microsecond molecular dynamics and graph theory-derived models to probe the allosteric role of key enhancement specificity mutations. We show that the mutations responsible for increasing the specificity of Cas9 alter the allosteric structure of the catalytic HNH domain, impacting the signal transmission from the DNA recognition region to the catalytic sites for cleavage. Specifically, the K855A mutation strongly disrupts the HNH domain allosteric structure, exerting the highest perturbation on the signaling transfer, while K810A and K848A result in more moderate effects on the allosteric intercommunication. This differential perturbation of the allosteric signaling reflects the different capabilities of the single mutants to increase Cas9 specificity, with the mutation achieving the highest specificity also strongly perturbing the signaling transfer. These outcomes reveal that the allosteric regulation is critical for the specificity enhancement of the Cas9 enzyme, and are valuable to harness the signaling network to improve the systems specificity.

A Role for Visual Areas in Physics Simulations

Ahuja, A. — 2021-09-16

To engage with the world, we must regularly make predictions about the outcomes of physical scenes. How do we make these predictions? Recent evidence points to simulation - the idea that we can introspectively manipulate rich, mental models of the world - as one possible explanation for how such predictions are accomplished. While theories based on simulation are supported by computational models, neuroscientific evidence for simulation is lacking and many important questions remain. For instance, do simulations simply entail a series of abstract computations? Or are they supported by sensory representations of the objects that comprise the scene being simulated? We posit the latter and suggest that the process of simulating a sequence of physical interactions is likely to evoke an imagery-like envisioning of those interactions. Using functional magnetic resonance imaging, we demonstrate that when participants predict how a ball will fall through an obstacle-filled display, motion-sensitive brain regions are activated. We further demonstrate that this activity, which occurs even though no motion is being sensed, resembles activity patterns that arise while participants perceive the balls motion. This finding suggests that the process of simulating the balls movement is accompanied by a sensory representation of this movement. These data thus demonstrate that mental simulations recreate sensory depictions of how a physical scene is likely to unfold.

Revealing spatio-temporal dynamics with long-term trypanosomatid live-cell imaging

Muniz, R. S. — 2021-09-15

Trypanosoma brucei, the causative agent of human African trypanosomiasis, employs a flagellum for dissemination within the parasites mammalian and insect hosts. T. brucei cells are highly motile in culture and must be able to move in all three dimensions for reliable cell division. These characteristics have made long-term microscopic imaging of live T. brucei cells challenging, which has limited our understanding of a variety of important cell-cycle events. To address this issue, we have devised an imaging approach that confines cells to small volumes that can be imaged continuously for up to 24 h. This system employs cast agarose microwells generated using a PDMS stamp that can be made with different dimensions to maximize cell viability and imaging quality. Using this approach, we have imaged individual T. brucei through multiple rounds of cell division with high spatial and temporal resolution. We have employed this method to study the differential rate of T. brucei daughter cell division and show that the approach is compatible with loss-of-function experiments such as small molecule inhibition and RNAi. We have also developed a strategy that employs in-well "sentinel" cells to monitor potential toxicity due to imaging. This live-cell imaging method will provide a novel avenue for studying a wide variety of cellular events in trypanosomatids that have previously been inaccessible. ImportanceTrypanosoma brucei causes severe diseases that affect humans and livestock in Sub-Saharan Africa. Efficient strategies for manipulating the T. brucei genome have provided a wealth of information about protein localization and function in diverse cellular processes. However, employing live-cell imaging for phenotypic analysis in T. brucei remains a significant challenge because immobilization of this highly motile parasite rapidly leads to morphologic defects and cell death. While fixed-cell imaging can provide snapshots of cellular events, it cannot provide the direct causal link or precise timing of events that comes from watching a living cell change over time. Our strategy using agarose microwells now allows long-term live cell T. brucei imaging with a simple apparatus that can be adapted for a wide range of experimental conditions.

Chromosome-scale genome assembly of the brown anole (Anolis sagrei), a model species for evolution and ecology

Geneva, A. J. — 2021-09-30

Rapid technological improvements are democratizing access to high quality, chromosome-scale genome assemblies. No longer the domain of only the most highly studied model organisms, now non-traditional and emerging model species can be genome-enabled using a combination of sequencing technologies and assembly software. Consequently, old ideas built on sparse sampling across the tree of life have recently been amended in the face of genomic data drawn from a growing number of high-quality reference genomes. Arguably the most valuable are those long-studied species for which much is already known about their biology; what many term emerging model species. Here, we report a new, highly complete chromosome-scale genome assembly for the brown anole, Anolis sagrei - a lizard species widely studied across a variety of disciplines and for which a high-quality reference genome was long overdue.

Chi3l1 is a modulator of glioma stem cell states and a therapeutic vulnerability for glioblastoma

Guetta-Terrier, C. — 2021-10-01

Chi3l1 (Chitinase 3-like 1) is a secreted protein highly expressed in glioblastoma. Here, we show that exposure of glioma stem cells (GSCs) to Chi3l1 reduces the CD133+/SOX2+ cells and increases the CD44+/Chi3l1+ cells. Chi3l1 binds to CD44 and induces phosphorylation and nuclear translocation of beta-catenin, Akt and STAT3. Single cell RNA-seq and RNA velocity following incubation of GSCs with Chi3l1 show significant changes in GSC state dynamics driving GSCs towards a mesenchymal expression profile and reducing transition probabilities towards terminal cellular states. ATAC-seq reveals that Chi3l1 increases accessibility of promoters containing MAZ transcription factor footprint. Inhibition of MAZ directly regulates genes with highest expression in cellular clusters exhibiting significant cell state transitions. Finally, targeting Chi3l1 in vivo with a blocking antibody, resets the transcriptomic profile of glioblastoma and inhibits tumor growth. Our work implicates Chi3l1 as modulator of GSC cellular states and demonstrates pre-clinical efficacy of anti-Chi3l1 antibody treatment.

Monkey Plays Pac-Man with Compositional Strategies and Hierarchical Decision-making

Yang, Q. — 2021-10-04

Humans can often handle daunting tasks with ease by developing a set of strategies to reduce decision making into simpler problems. The ability to use heuristic strategies demands an advanced level of intelligence and has not been demonstrated in animals. Here, we trained macaque monkeys to play the classic video game Pac-Man. The monkeys decision-making may be described with a strategy-based hierarchical decision-making model with over 90% accuracy. The model reveals that the monkeys adopted the take-the-best heuristic by using one dominating strategy for their decision-making at a time and formed compound strategies by assembling the basis strategies to handle particular game situations. With the model, the computationally complex but fully quantifiable Pac-Man behavior paradigm provides a new approach to understanding animals advanced cognition. One-Sentence SummaryMacaque monkeys play Pac-Man with strategy-based hierarchical decision making, a cognitive capacity hitherto unknown in them.

Amplitude discrimination is predictably affected by echo frequency filtering in wideband echolocating bats

Tuninetti, A. — 2021-10-05

Big brown bats emit wideband frequency modulated (FM) ultrasonic pulses for echolocation. They perceive target range from echo delay and target size from echo amplitude. Their sounds contain two prominent down-sweeping harmonic sweeps (FM1, [~]55-22 kHz; FM2, [~]100-55 kHz), which are affected differently by propagation out to the target and back to the bat. FM2 is attenuated more than FM1 during propagation. Bats anchor target ranging asymmetrically on the low frequencies in FM1, while FM2 only contributes if FM1 is present as well. These experiments tested whether the bats ability to discriminate target size from the amplitude of echoes is affected by selectively attenuating upper or lower frequencies. Bats were trained to perform an echo amplitude discrimination task with virtual echo targets 83 cm away. While echo delay was held constant and echo amplitude was varied to estimate threshold, either lower FM1 frequencies or higher FM2 frequencies were attenuated. The results parallel effects seen in echo delay experiments; bats performance was significantly poorer when the lower frequencies in echoes were attenuated, compared to higher frequencies. The bats ability to distinguish between virtual targets at the same simulated range from echoes arriving at the same delay indicates a high level of focused attention for perceptual isolation of one and suppression of the other.

Investigating lead removal at trace concentrations from water by inactive yeast cells

Stathatou, P. M. — 2021-10-09

Traces of heavy metals found in water resources, due to mining activities and e-waste discharge, pose a global threat. Conventional treatment processes fail to remove toxic heavy metals, such as lead, from drinking water in a resource-efficient manner when their initial concentrations are low. Here, we show that by using the yeast Saccharomyces cerevisiae we can effectively remove trace lead from water via a rapid mass transfer process, achieving an uptake of up to 12 mg lead per gram of biomass in solutions with initial lead concentrations below 1 part per million. We found that the yeast cell wall plays a crucial role in this process, with its mannoproteins and {beta}-glucans being the key potential lead adsorbents. Furthermore, we discovered that biosorption is linked to a significant increase in cell wall stiffness. These findings open new opportunities for using environmentally friendly and abundant biomaterials for advanced water treatment targeting emerging contaminants. One-Sentence SummaryRemoving toxic heavy metals from water at challenging trace levels in an environmentally friendly, resource-efficient manner.

Regulation of Nav1.6-mediated sodium currents underlie the homeostatic control of neuronal intrinsic excitability in the optic tectum of the developing Xenopus laevis tadpole

Thompson, A. C. — 2021-10-08

For individual neurons to function appropriately within a network that is undergoing synaptic reorganization and refinement due to developmental or experience-dependent changes in circuit activity, they must homeostatically adapt their intrinsic excitability to maintain a consistent output despite the changing levels of synaptic input. This homeostatic plasticity of excitability is particularly important for the development of sensory circuits, where subtle deficits in neuronal and circuit function cause developmental disorders including autism spectrum disorder and epilepsy. Despite the critical importance of this process for normal circuit development, the molecular mechanism by which this homeostatic control of intrinsic excitability is regulated is not fully understood. Here, we demonstrate that Xenopus optic tectal neurons express distinct fast, persistent and resurgent Na+ currents. Here, we demonstrate that Xenopus optic tectal neurons express distinct fast, persistent and resurgent Na+ currents. These are regulated with developmental changes in synaptic input, and homeostatically in response to changes in visual input. We show that expression of the voltage-gated Na+ channel subtype Nav1.6 is regulated with changes in intrinsic excitability, that blocking Nav1.6 channels is sufficient to decrease intrinsic excitability. Furthermore, that upregulation of Nav1.6 expression is necessary for experience-dependent increases in Na+ currents and intrinsic excitability. Finally, by examining behaviors that rely on visual and multisensory integration, we extend these findings to show that tight regulation of Na+ channel gene expression during a critical period of tectal circuit development is required for the normal functional development of the tectal circuitry.

Whole genome sequence analysis of blood lipid levels in >66,000 individuals

Selvaraj, M. S. — 2021-10-12

Plasma lipids are heritable modifiable causal factors for coronary artery disease, the leading cause of death globally. Despite the well-described monogenic and polygenic bases of dyslipidemia, limitations remain in discovery of lipid-associated alleles using whole genome sequencing, partly due to limited sample sizes, ancestral diversity, and interpretation of potential clinical significance. Increasingly larger whole genome sequence datasets with plasma lipids coupled with methodologic advances enable us to more fully catalog the allelic spectrum for lipids. Here, among 66,329 ancestrally diverse (56% non-European ancestry) participants, we associate 428M variants from deep-coverage whole genome sequences with plasma lipids. Approximately 400M of these variants were not studied in prior lipids genetic analyses. We find multiple lipid-related genes strongly associated with plasma lipids through analysis of common and rare coding variants. We additionally discover several significantly associated rare non-coding variants largely at Mendelian lipid genes. Notably, we detect rare LDLR intronic variants associated with markedly increased LDL-C, similar to rare LDLR exonic variants. In conclusion, we conducted a systematic whole genome scan for plasma lipids expanding the alleles linked to lipids for multiple ancestries and characterize a clinically-relevant rare non-coding variant model for lipids.

Distinct neocortical mechanisms underlie human SI responses to median nerve and laser evoked peripheral activation

Thorpe, R. V. — 2021-10-12

Magneto- and/or electro-encephalography (M/EEG) are non-invasive clinically-relevant tools that have long been used to measure electromagnetic fields in somatosensory cortex evoked by innocuous and noxious somatosensory stimuli. Two commonly applied stimulation paradigms that produce distinct responses in primary somatosensory cortex (SI) linked to innocuous and noxious sensations are electrical median nerve (MN) stimulation and cutaneous laser-evoked (LE) stimulation to the dorsum of the hand, respectively. Despite their prevalence, the physiological mechanisms that produce stereotypic macroscale MN and LE responses have yet to be fully articulated, limiting their utility in understanding brain dynamics associated with non-painful and/or painful somatosensation. Through a literature review, we detailed features of MN and LE responses source-localized to SI that are robust and reproducible across studies. We showed that the first peak in the MN response at [~]20 ms post-stimulus (i.e., MN N1) corresponds to outward-directed deep-to-superficial electrical current flow through the cortical laminae, which is followed by inward-directed current at [~]30 ms (i.e., MN P1). In contrast, the initial LE response occurs later at [~]170 ms (i.e., LE N1) and is oriented inward and opposite the direction of the MN N1. We then examined the neocortical circuit mechanisms contributing to the robust features of each response using the Human Neocortical Neurosolver (HNN) neural modeling software tool (Neymotin et al., 2020). Using HNN as a hypothesis development and testing tool, model results predicted the MN response can be simulated with a sequence of layer specific thalamocortical and cortico-cortical synaptic drive similar to that previously reported for tactile evoked responses (Jones et al., 2007; Neymotin et al., 2020), with the novel discovery that an early excitatory input to supragranular layers at [~]30 ms is an essential mechanism contributing to the inward current flow of the MN P1. Model results further predicted that the initial [~]170 ms inward current flow of the LE N1 was generated by a burst of repetitive gamma-frequency ([~]40 Hz) excitatory synaptic drive to supragranular layers, consistent with prior reports of LE gamma-frequency activity. These results make novel and detailed multiscale predictions about the dynamic laminar circuit mechanisms underlying temporal and spectral features of MN and LE responses in SI and can guide further investigations in follow-up studies. Ultimately, these findings may help with the development of targeted therapeutics for pathological somatosensation, such as somatic sensitivity and acute neuropathic pain.

Diet and Domestication Drive Evolution of the Gut Holobiome

Ramanan, V. — 2021-10-16

The host microbiome encompasses all microorganisms of a host. Host and microbiome coevolution in the gut result in differing microbial compositions, functionality, and host diet [1]. Host diet modulates what macromolecules are used for gut microbial metabolism, which can determine digestion, health, and behavior [2, 3]. Microbial composition across animals provides data on how microbiomes segregate between species and diets [4]. Here we show that microbiome data from GenBank can model host evolution, providing a "holobiome" insight to the important roles of diet and domestication. The main findings of this study in respect to microbial composition among species were: (1) herbivores are more similar than hosts with other diets; (2) domesticated species are more similar than wild relatives; and (3) humans are distinct from primates. Microbial composition between diets indicates a difference in functionality, where protein and fiber degradation are seen more in carnivores and herbivores respectively. Additionally, herbivores show the most microbial diversity among the diets. Finally, this analysis informs us of gaps in current microbiome data collection, which is biased toward pathogens. Thus, the host-microbiome relationship depicts a complex web of microbial functionality, composition, and diet that impact coevolution.

Blood flow synchronization in renal microcirculation - a high-resolution imaging study.

Postnov, D. D. — 2021-10-16

AimsInternephron signalling and interaction are fundamental for kidney function. Earlier studies have shown that nephrons signal to each other over short distances and adjust their activity accordingly. Micropuncture experiments revealed synchronous clusters of 2-3 nephrons formed from such interactions, while imaging and modelling results suggested the possibility of larger clusters. Such clusters are expected to play an important role in renal autoregulation, but their presence has not been confirmed and their size has not been estimated. In this study, we present methodology for high resolution renal blood flow imaging and apply it to estimate frequency and phase angle differences in kidney blood vessels under normal conditions and after administration of the vasoactive agents angiotensin II and acetylcholine. Methods and resultsTo resolve signals from separate arterioles in a sufficiently large field of view, we developed a method for renal laser speckle contrast imaging. Our setup provides imaging of blood flow in the kidney cortex with a limit of image resolution at 0.8m per pixel and imaging frequency of 160Hz. We used the method to record from 1.5x1.5 mm2 sections of the renal surface in anaesthetised Sprague-Dawley rats in unstimulated conditions and during IV infusion of the vasoconstrictor angiotensin II or the vasodilator acetylcholine. In each section, we resolved and segmented 94.8{+/-}15.66 individual arterioles and venules, and analyzed blood flow using wavelet spectral analysis to identify clusters of synchronized blood vessels. ConclusionsWe observed spatial and temporal evolution of blood vessel clusters of various sizes, including the formation of large (>90 vessels) long-lived clusters (>10 periods) locked at the frequency of the tubular glomerular feedback (TGF) mechanism. The analysis showed that synchronization patterns and thus the co-operative dynamics of nephrons change significantly when either of the vasoactive agents is administered. On average, synchronization was stronger (larger clusters, longer duration) with angiotensin II administration than in the unstimulated state or with acetyl choline. While it weakens with distance, increased synchronization duration spanned the whole field of view, and likely, beyond it. Neighbouring vessels tend to demonstrate in-phase synchronization, especially in the vasoconstricted condition, which is expected to cause locally increased pressure variation. Our results confirm both the presence of the local synchronization in the renal microcirculatory blood flow and the fact that it changes depending on the condition of the vascular network and the blood pressure, which might have further implications for the role of such synchronization in pathologies development.

Cortical Spheroid Model for Studying the Effects of Ischemic Brain Injury

McLaughlin, R. M. — 2021-10-16

Stroke is a devastating neurological disorder and a leading cause of death and long-term disability. Despite many decades of research, there are still very few therapeutic options for patients suffering from stroke or its consequences. This is partially due to the limitations of current research models, including traditional in vitro models which lack the three-dimensional (3D) architecture and cellular make-up of the in vivo brain. 3D spheroids derived from primary postnatal rat cortex provide an in vivo-relevant model containing a similar cellular composition to the native cortex and a cell-synthesized extracellular matrix. These spheroids are costeffective, highly reproducible, and can be produced in a high-throughput manner, making this model an ideal candidate for screening potential therapeutics. To study the cellular and molecular mechanisms of stroke in this model, spheroids were deprived of glucose, oxygen, or both oxygen and glucose for 24 hours. Both oxygen and oxygen-glucose deprived spheroids demonstrated many of the hallmarks of stroke, including a decrease in metabolism, an increase in neural dysfunction, and an increase in reactive astrocytes. Pretreatment of spheroids with the antioxidant agent N-acetylcysteine (NAC) mitigated the decrease in ATP seen after 24 hours of oxygen-glucose deprivation. Together, these results show the utility of our 3D cortical spheroid model for studying ischemic injury and its potential for screening stroke therapeutics. Significance StatementThose who survive after suffering a stroke often have long-term cognitive or physical disabilities. There is currently only one available therapeutic, tissue plasminogen activator (tPA), and it must be administered within a few hours after the onset of stroke. As stroke prevalence increases with our aging population, there is a growing need for therapies to mitigate or reverse the resulting brain damage. Three-dimensional (3D) culture systems have the potential to screen novel therapeutics more reliably than traditional in vitro models. Here we present a novel 3D cortical spheroid ischemia model which replicates many of the characteristics of stroke and has the potential to be an effective tool in therapeutic development.

Roosting ecology and the evolution of bat landing maneuvers

Boerma, D. — 2021-10-22

Biomechanics is poised at the intersection of organismal form, function, and ecology, and forms a practical lens through which to investigate evolutionary linkages among these factors. We conducted the first evolutionary analysis of bat flight dynamics by examining the phylogenetic patterning of landing mechanics. We discovered that bats perform stereotyped maneuvers that are correlated with landing performance quantified as impact force, and that these are linked with roosting ecology, a critical aspect of bat biology. Our findings suggest that bat ancestors performed simple, four-limbed landings, similar to those performed by gliding mammals, and that more complex landings evolved in association with novel roost types. This explicit connection between ecology and biomechanics presents the opportunity to identify traits that are associated with a locomotor behavior of known ecological relevance, thus laying the foundation for a broader understanding of the evolution of flight and wing architecture in this extraordinarily successful mammalian lineage.

Sign inversion in selection on ploidy

Raynes, Y. — 2021-10-28

Ploidy - the number of homologous chromosome sets in a cell - is remarkably variable across the natural world, yet the evolutionary processes that have resulted in such diversity remain poorly understood. Here we use stochastic agent-based simulations to model ploidy evolution under the influence of indirect selection, i.e., selection mediated solely by statistical associations with fitness-affecting mutations. We find that in non-equilibrium asexual populations, the sign of selection on ploidy can change with population size - a phenomenon we have previously termed sign inversion. In large populations, ploidy dynamics are dominated by indirect effects of selection on beneficial mutations, which favors haploids over diploids. However, as population size declines, selection for beneficial mutations is neutralized by random genetic drift before drift can overwhelm selection against the cost of the deleterious mutational load. As a result, in small populations indirect selection is dominated by the cost of the deleterious load, which favors diploids over haploids. Our work adds to the growing body of evidence challenging established evolutionary theory that population size can affect only the efficiency, but not the sign, of natural selection.

Differential Vulnerability of Anterior Cingulate Cortex Cell-Types to Diseases and Drugs

Shukla, R. — 2021-10-28

In psychiatric disorders, mismatches between disease-states and therapeutic strategies are highly pronounced, largely because of unanswered questions regarding specific vulnerabilities of different cell-types and therapeutic responses. Which cellular events (housekeeping or salient) are most affected? Which cell-types succumb first to challenges, and which exhibit the strongest response to drugs? Are these events coordinated between cell-types? How does the disease-state and drug affect this coordination? To address these questions, we analyzed single-nucleus-RNAseq (sn-RNAseq) data from the human anterior cingulate cortex--a region involved in many psychiatric disorders. Density index, a metric for quantifying similarities and dissimilarities across functional profiles, was employed to identify common (housekeeping) or salient functional themes across all cell-types. Cell-specific signatures were integrated with existing disease and drug-specific signatures to determine cell-type-specific vulnerabilities, druggabilities, and responsiveness. Clustering of functional profiles revealed cell-types jointly participating in these events. SST and VIP interneurons were found to be most vulnerable, whereas pyramidal neurons were least vulnerable. Overall, the disease-state is superficial layer-centric, largely influences cell-specific salient themes, strongly impacts disinhibitory neurons, and influences astrocyte interaction with a subset of deep-layer pyramidal neurons. Drug activities, on the other hand, are deep layer-centric and involve activating a distinct subset of deep-layer pyramidal neurons to circumvent the disinhibitory circuit malfunctioning in the disease-state. These findings demonstrate a novel application of sn-RNAseq data to explain drug and disease action at a systems level, suggests a targeted drug development and reevaluate various postmortem-based findings.

Sciara coprophila larvae upregulate DNA repair pathways and downregulate developmental regulators in response to ionizing radiation

Urban, J. M. — 2021-10-28

Robust DNA damage prevention and repair strategies are crucial to faithful reproduction and inheritance of the genetic material. Although many molecular pathways that respond to DNA damage are well conserved through evolution, the quality and effectiveness of these systems can vary between species. Studies dating back for nearly a century document that the dark-winged fungus gnat Sciara coprophila (Order: Diptera; sub-order: Nematocera) is relatively resistant to irradiation-induced mutations that cause visible phenotypes when compared to the fruit fly Drosophila melanogaster (Order: Diptera; sub-order: Brachycera). However, the molecular responses to irradiation for S. coprophila have yet to be analyzed. To address this gap, we first characterized the effects of ionizing radiation on S. coprophila throughout its life cycle. Our data show that developing S. coprophila embryos are highly sensitive to even low doses of gamma-irradiation, whereas larvae can tolerate up to 80 Gy and still retain their ability to develop to adulthood with a developmental delay of 5 to 8 extra days in the larval stage. To survey the genes involved in the early transcriptional response to irradiation, we compared RNA-seq profiles of larvae with and without radiation treatment. Our analysis showed that 327 genes are differentially expressed in irradiated larvae, with 232 genes upregulated and 95 genes downregulated relative to controls. The upregulated genes were enriched for DNA damage response genes, including those involved in DNA repair, cell cycle arrest, and apoptosis, whereas the down-regulated genes were enriched for developmental regulators, consistent with the developmental delay observed in irradiated larvae. Thus, our study has laid the groundwork to further dissect how Sciara copes with radiation-induced damage.

The early embryonic transcriptome of a Hawaiian Drosophila picture-wing fly shows evidence of altered gene expression and novel gene evolution

Chenevert, M. — 2021-10-29

A massive adaptive radiation on the Hawaiian archipelago has produced approximately one quarter of the fly species in the family Drosophilidae. The Hawaiian Drosophila clade has long been recognized as a model system for the study of both the ecology of island endemics and the evolution of developmental mechanisms, but relatively few genomic and transcriptomic datasets are available for this group. We present here a differential expression analysis of the transcriptional profiles of two highly conserved embryonic stages in the Hawaiian picture-wing fly Drosophila grimshawi. When we compared our results to previously published datasets across the family Drosophilidae, we identified cases of both gains and losses of gene representation in D. grimshawi, including an apparent delay in Hox gene activation. We also found high expression of unannotated genes. Most transcripts of unannotated genes with open reading frames do not have homologs in non-Hawaiian Drosophila species, although the vast majority have sequence matches in other genomes of the Hawaiian picture-wing flies. Some of these genes may have arisen from non-coding sequence in the ancestor of Hawaiian flies or during the evolution of the clade. Our results suggests that both the modified use of ancestral genes and the evolution of new ones may occur in rapid radiations. RESEARCH HIGHLIGHTSThe early embryonic transcriptome of the Hawaiian fly Drosophila grimshawi shows a loss of expression of conserved Stage 5 genes, including the Hox genes The de novo evolution of embryonically expressed genes may be occurring in the Hawaiian Drosophila lineage AUTHORS STATEMENTThis paper is not being considered for publication elsewhere. This study formed part of Madeline Cheneverts M.S. thesis.

Selective Control of Synaptically-Connected Circuit Elements by All-Optical Synapses

Prakash, M. — 2021-11-01

Understanding percepts, engrams and actions requires methods for selectively modulating synaptic communication between specific subsets of interconnected cells. Here, we develop an approach to control synaptically connected elements using bioluminescent light: Luciferase-generated light, originating from a presynaptic axon terminal, modulates an opsin in its postsynaptic target. Vesicular-localized luciferase is released into the synaptic cleft in response to presynaptic activity, creating a real-time Optical Synapse. Light production is under experimenter-control by introduction of the small molecule luciferin. Signal transmission across this optical synapse is temporally defined by the presence of both the luciferin and presynaptic activity. We validate synaptic Interluminescence by multi-electrode recording in cultured neurons and in mice in vivo. Interluminescence represents a powerful approach to achieve synapse-specific and activity-dependent circuit control during behavior in vivo.

A retinal circuit that vetoes optokinetic responses to fast visual motion

Mani, A. — 2021-11-03

Optokinetic nystagmus (OKN) complements the vestibulo-ocular reflex (VOR) to stabilize the retinal image during head rotation. OKN is driven by the ON direction-selective ganglion cells (ON DSGCs), a rare class of retinal output neuron that encodes both the direction and speed of global retinal slip. The cells and synaptic circuits that give ON DSGCs their directional tuning are well known, but those dictating their slow-speed preference (and thus OKNs) remain enigmatic. Here, we probe this circuit through patch recordings, functional imaging, genetic manipulation, and serial electron microscopic reconstruction in mouse retina. We confirm earlier evidence that feedforward glycinergic inhibition is the main suppressor of ON DSGC responses to fast motion and reveal a surprising source for this inhibition [boxh] the VGluT3 amacrine cell, a retinal interneuron that releases both glycine and glutamate, exciting some neurons and inhibiting others. We find that VGluT3 cells respond robustly to fast global motion and that their output reaches most RGC types, as well as a diverse group of amacrine and bipolar cells. They enhance the response of ON-OFF DSGCs to fast motion, while suppressing it in ON DSGCs. Together, our results identify a novel role for VGluT3 cells, limiting the range of retinal slip speeds that drive image-stabilizing eye movements. More broadly, they suggest VGluT3 cells shape the response of many RGCs and amacrine cells to fast motion.

A Trypanosoma brucei orphan kinesin employs a convergent microtubule organization strategy to complete cytokinesis

Sladewski, T. E. — 2021-11-04

Many single-celled eukaryotes have complex cell morphologies defined by cytoskeletal elements comprising microtubules arranged into higher-order structures. Trypanosoma brucei (T. brucei) cell polarity is mediated by a parallel array of microtubules that underlie the plasma membrane and define the auger-like shape of the parasite. The subpellicular array must be partitioned and segregated using a microtubule-based mechanism during cell division. We previously identified an orphan kinesin, KLIF, that localizes to the division plane and is essential for the completion of cytokinesis. To gain mechanistic insight into how this novel kinesin functions to complete cleavage furrow ingression, we characterized the biophysical properties of the KLIF motor domain in vitro. We found that KLIF is a non-processive dimeric kinesin that dynamically crosslinks microtubules. Microtubules crosslinked in an antiparallel orientation are translocated relative to one another by KLIF, while microtubules crosslinked parallel to one another remain static, resulting in the formation of organized parallel bundles. In addition, we found that KLIF stabilizes the alignment of microtubule plus ends. These features provide a mechanistic understanding for how KLIF functions to form a new pole of aligned microtubule plus ends that defines the shape of the new posterior, which is a unique requirement for the completion of cytokinesis in T. brucei.

Three-dimensional in vitro model of the device-tissue interface reveals innate neuroinflammation can be mitigated by antioxidant ceria nanoparticles

Atherton, E. — 2021-11-08

The recording instability of neural implants due to neuroinflammation at the device-tissue interface (DTI) is a primary roadblock to broad adoption of brain-machine interfaces. While a multiphasic immune response, marked by glial scaring, oxidative stress (OS), and neurodegeneration, is well-characterized, the independent contributions of systemic and local "innate" immune responses are not well-understood. Three-dimensional primary neural cultures provide a unique environment for studying the drivers of neuroinflammation by decoupling the innate and systemic immune systems, while conserving an endogenous extracellular matrix and structural and functional network complexity. We created a three-dimensional in vitro model of the DTI by seeding primary cortical cells around microwires. Live imaging of microtissues over time revealed independent innate neuroinflammation, marked by increased OS, decreased neuronal density, and increased functional connectivity. We demonstrated the use of this model for therapeutic screening by directly applying drugs to neural tissue, bypassing low bioavailability through the in vivo blood brain barrier. As there is growing interest in long-acting antioxidant therapies, we tested efficacy of "perpetual" antioxidant ceria nanoparticles, which reduced OS, increased neuronal density, and protected functional connectivity. Overall, our avascular in vitro model of the DTI exhibited symptoms of OS-mediated innate neuroinflammation which were mitigated by antioxidant intervention.

Low abundance members of the gut microbiome are potent drivers of immune cell education.

Han, G. G. — 2021-11-10

One of the main goals of microbiome research is to identify bacterial members that significantly affect host phenotypes and understand their contributions to disease pathogenesis. Studies identifying bacterial members that dictate host phenotype have focused mainly on the dominant members, and the role of low abundance microbes in determining host phenotypes and pathogenesis of diseases remains unexplored. In this study, we compared the gut bacterial community of mice with wide-ranging microbial exposure to determine if low abundance bacteria vary based on microbial exposure or remain consistent. We noted that similar to the high abundance bacterial community, a core community of low abundance bacteria made up a significant portion of the gut microbiome irrespective of microbial exposure. To determine the effect of low abundance bacteria on community structure and host gene expression, we devised a microbiome dilution strategy to "delete" out low abundance bacteria and engrafted the diluted microbiomes into germ-free mice. Our approach successfully excluded low abundance bacteria from small and large intestinal bacterial communities and induced global changes in microbial community structure and composition in the large intestine. Gene expression analysis of intestinal tissue revealed that loss of low abundance bacteria resulted in a drastic reduction in expression of multiple genes involved MHC class II antigen presentation pathway and T-cell cytokine production in the small intestine. The effect of low abundance bacteria on MHC class II expression was found specific to the intestinal epithelium at an early timepoint post-colonization and correlated with bacteria belonging to the family Erysipelotrichaceae. We conclude that low abundance bacteria have a significantly higher immuno-stimulatory effect compared to dominant bacteria and are thus potent drivers of early immune education in the gut. Therefore, characterizing immune interaction of low abundance bacteria with the host will offer greater insight into the intestinal immune landscape and disease pathogenesis.

Unsupervised integration of single-cell multi-omics datasets with disparities in cell-type representation

Demetci, P. — 2021-11-11

Integrated analysis of multi-omics data allows the study of how different molecular views in the genome interact to regulate cellular processes; however, with a few exceptions, applying multiple sequencing assays on the same single cell is not possible. While recent unsupervised algorithms align single-cell multi-omic datasets, these methods have been primarily benchmarked on co-assay experiments rather than the more common single-cell experiments taken from separately sampled cell populations. Therefore, most existing methods perform subpar alignments on such datasets. Here, we improve our previous work Single Cell alignment using Optimal Transport (SCOT) by using unbalanced optimal transport to handle disproportionate cell-type representation and differing sample sizes across single-cell measurements. We show that our proposed method, SCOTv2, consistently yields quality alignments on five real-world single-cell datasets with varying cell-type proportions and is computationally tractable. Additionally, we extend SCOTv2 to integrate multiple (M [≥] 2) single-cell measurements and present a self-tuning heuristic process to select hyperparameters in the absence of any orthogonal correspondence information. Available at: http://rsinghlab.github.io/SCOT.

Biophysical and Architectural Mechanisms of Subthalamic Theta under Response Conflict

Moolchand, P. — 2021-11-11

The cortico-basal ganglia circuit is needed to suppress prepotent actions and to facilitate controlled behavior. Under conditions of response conflict, the frontal cortex and subthalamic nucleus [STN] exhibit increased spiking and theta band power, which are linked to adaptive regulation of behavioral output. The electrophysiological mechanisms underlying these neural signatures of impulse control remain poorly understood. To address this lacuna, we constructed a novel large-scale, biophysically principled model of the subthalamopallidal [STN-Globus Pallidus externus (GPe)] network, and examined the mechanisms that modulate theta power and spiking in response to cortical input. Simulations confirmed that theta power does not emerge from intrinsic network dynamics but is robustly elicited in response to cortical input as burst events representing action selection dynamics. Rhythmic burst events of multiple cortical populations, representing a state of conflict where cortical motor plans vacillate in the theta range, led to prolonged STN theta and increased spiking, consistent with empirical literature. Notably, theta band signaling required NMDA, but not AMPA, currents, which were in turn related to a triphasic STN response characterized by spiking, silence and bursting periods. Finally, theta band resonance was also strongly modulated by architectural connectivity, with maximal theta arising when multiple cortical populations project to individual STN "conflict detector" units, due to an NMDA-dependent supralinear response. Our results provide insights into the biophysical principles and architectural constraints that give rise to STN dynamics during response conflict, and how their disruption can lead to impulsivity and compulsivity.

Beyond gradients: Noise correlations control Hebbian plasticity to shape credit assignment

Scott, D. N. — 2021-11-20

Interference and generalization, which refer to counter-productive and useful interactions between learning episodes, respectively, are poorly understood in biological neural networks. Whereas much previous work has addressed these topics in terms of specialized brain systems, here we investigated how learning rules should impact them. We found that plasticity between groups of neurons can be decomposed into biologically meaningful factors, with factor geometry controlling interference and generalization. We introduce a "coordinated eligibility theory" in which plasticity is determined according to products of these factors, and is subject to surprise-based metaplasticity. This model computes directional derivatives of loss functions, which need not align with task gradients, allowing it to protect networks against catastrophic interference and facilitate generalization. Because the models factor structure is closely related to other plasticity rules, and is independent of how feedback is transmitted, it introduces a widely-applicable framework for interpreting supervised, reinforcement-based, and unsupervised plasticity in nervous systems.

LRRC15 is an inhibitory receptor blocking SARS-CoV-2 spike-mediated entry in trans

Song, J. — 2021-11-24

SARS-CoV-2 infection is mediated by the entry receptor ACE2. Although attachment factors and co-receptors facilitating entry are extensively studied, cellular entry factors inhibiting viral entry are largely unknown. Using a surfaceome CRISPR activation screen, we identified human LRRC15 as an inhibitory receptor for SARS-CoV-2 entry. LRRC15 directly binds to the receptor-binding domain (RBD) of spike protein with a moderate affinity and inhibits spike-mediated entry. Analysis of human lung single cell RNA sequencing dataset reveals that expression of LRRC15 is primarily detected in fibroblasts and particularly enriched in pathological fibroblasts in COVID-19 patients. ACE2 and LRRC15 are not co-expressed in the same cell types in the lung. Strikingly, expression of LRRC15 in ACE2-negative cells blocks spike-mediated viral entry in ACE2+ cell in trans, suggesting a protective role of LRRC15 in a physiological context. Therefore, LRRC15 represents an inhibitory receptor for SARS-CoV-2 regulating viral entry in trans.

Social preference in rats not impacted by posterior parietal activity despite overall changes in familiarity-based social behavior

Wise, T. B. — 2021-11-25

Recent literature points to a potential link between the evolution of complex social behavior and the posterior parietal cortex (PPC) in primates including humans (Parkinson & Wheatley, 2013). Thus far, this theory has been overlooked in other highly social animals that may have also evolved due to social selective pressures. In rodents, there is limited knowledge on the involvement of the PPC on sociality, and most studies of such behavior are limited to understanding social preference. We investigated the role of the PPC through two experiments using the 3-Chamber Sociability and Social Novelty test in rats (Crawley, 2004). In Experiment 1, we used a standard 3-Chamber paradigm, which included two novel demonstrators. In Experiment 2, this paradigm was altered to increase the difference in familiarity between demonstrators such that one demonstrator was highly familiar to the subject and the other was entirely novel. Rats with pre-testing permanent neurotoxic lesions were compared to sham surgery control rats, and the same rats were used for both experiments. Experiments 1 and 2 showed that both groups of rats preferred general social interaction, suggesting no deficit in sociability following PPC damage, regardless of demonstrator identity. Further, experimental and control rats showed similar levels of novelty preference following PPC damage, with novelty preferences increasing in Experiment 2. We argue that heightened novelty preference in Experiment 2 may reflect the increased difference in familiarity between demonstrators. Within the confines of the 3-Chamber task, our results suggest that PPC function was not required for general sociability or social novelty recognition. Because the PPC is implicated in abstract cognition, we argue that existing social tests in rodents may not adequately measure the complex cognitive capacities thought to be supported by the PPC. Future studies should investigate the role of the PPC in social cognition by employing behavioral tasks that require higher cognitive demand rather than testing inherent preference for social partners. Outside of our investigation of the PPC, these results show that social novelty preference can be manipulated through changes in familiarity of demonstrators, and that rats can discriminate others social identities.

Archaic introgression and variation in pharmacogenes and implications for local adaptation

Wroblewski, T. H. — 2021-11-28

Modern humans carry both Neanderthal and Denisovan (archaic) genome elements that are part of the human gene pool and affect the life and health of living individuals. The impact of archaic DNA may be particularly evident in pharmacogenes - genes responsible for the processing of exogenous substances such as food, pollutants, and medications. However, the health implications and contribution of archaic ancestry in pharmacogenes of modern humans remains understudied. Here, we explore eleven key cytochrome P450 genes (CYP450) involved in drug metabolizing reactions in three Neanderthal and one Denisovan individuals as well as examine archaic introgression in modern human populations. We infer the metabolizing efficiency of these eleven CYP450 genes in archaic individuals and find important phenotypic differences relative to modern human variants. We identify several single nucleotide variants shared between archaic and modern humans in each gene, including some potentially function-altering mutations in archaic CYP450 genes, which may result in altered metabolism in living people carrying these variants. We highlight three genes which show evidence for archaic introgression into modern humans, as well as one additional gene that shows evidence for a gene duplication found only in Neanderthals and modern Africans.

Clonal hematopoiesis is driven by aberrant activation of TCL1A

Weinstock, J. S. — 2021-12-13

A diverse set of driver genes, such as regulators of DNA methylation, RNA splicing, and chromatin remodeling, have been associated with pre-malignant clonal expansion of hematopoietic stem cells (HSCs). The factors mediating expansion of these mutant clones remain largely unknown, partially due to a paucity of large cohorts with longitudinal blood sampling. To circumvent this limitation, we developed and validated a method to infer clonal expansion rate from single timepoint data called PACER (passenger-approximated clonal expansion rate). Applying PACER to 5,071 persons with clonal hematopoiesis accurately recapitulated the known fitness effects due to different driver mutations. A genome-wide association study of PACER revealed that a common inherited polymorphism in the TCL1A promoter was associated with slower clonal expansion. Those carrying two copies of this protective allele had up to 80% reduced odds of having driver mutations in TET2, ASXL1, SF3B1, SRSF2, and JAK2, but not DNMT3A. TCL1A was not expressed in normal or DNMT3A-mutated HSCs, but the introduction of mutations in TET2 or ASXL1 by CRISPR editing led to aberrant expression of TCL1A and expansion of HSCs in vitro. These effects were abrogated in HSCs from donors carrying the protective TCL1A allele. Our results indicate that the fitness advantage of multiple common driver genes in clonal hematopoiesis is mediated through TCL1A activation. PACER is an approach that can be widely applied to uncover genetic and environmental determinants of pre-malignant clonal expansion in blood and other tissues.

Non-rapid eye movement sleep and wake neurophysiology in schizophrenia

Kozhemiako, N. — 2021-12-16

Motivated by the potential of objective neurophysiological markers to index thalamocortical function in patients with severe psychiatric illnesses, we comprehensively characterized key NREM sleep parameters across multiple domains, their interdependencies, and their relationship to waking event-related potentials and symptom severity. In 130 schizophrenia (SCZ) patients and controls, we confirmed a marked reduction in sleep spindle density in SCZ and extended these findings to show that only slow spindles predicted symptom severity, and that fast and slow spindle properties were largely uncorrelated. We also describe a novel measure of slow oscillation and spindle interaction that was attenuated in SCZ. The main sleep findings were replicated in a demographically distinct sample, and a joint model, based on multiple NREM components, predicted disease status in the replication cohort. Although also altered in patients, auditory event-related potentials elicited during wake were unrelated to NREM metrics. Consistent with a growing literature implicating thalamocortical dysfunction in SCZ, our characterization identifies independent NREM and wake EEG biomarkers that may index distinct aspects of SCZ pathophysiology and point to multiple neural mechanisms underlying disease heterogeneity. This study lays the groundwork for evaluating these neurophysiological markers, individually or in combination, to guide efforts at treatment and prevention as well as identifying individuals most likely to benefit from specific interventions.

Engraftment, fate, and function of HoxB8-conditional neutrophil progenitors in the unconditioned murine host

Cohen, J. T. — 2021-12-21

The development and use of murine myeloid progenitor cell lines that are conditionally immortalized through expression of HoxB8 has provided a valuable tool for studies of neutrophil biology. Recent work has extended the utility of HoxB8-conditional progenitors to the in vivo setting via their transplantation into irradiated mice. Here, we describe the isolation of HoxB8-conditional progenitor cell lines that are unique in their ability to engraft in the naive host in the absence of conditioning of the hematopoietic niche. Our results indicate that HoxB8-conditional progenitors engraft in a {beta}1 integrin-dependent manner and transiently generate donor-derived mature neutrophils. Furthermore, we show that neutrophils derived in vivo from transplanted HoxB8-conditional progenitors are mobilized to the periphery and recruited to sites of inflammation in a manner that depends on the C-X-C chemokine receptor 2 and {beta}2 integrins, the same mechanisms that have been described for recruitment of endogenous primary neutrophils. Together, our studies advance the understanding of HoxB8-conditional neutrophil progenitors and describe an innovative tool that, by virtue of its ability to engraft in the naive host, will facilitate mechanistic in vivo experimentation on neutrophils.

Myofibroblast Senescence Promotes Arrhythmogenic Remodeling in the Aged Infarcted Rabbit Heart

Baggett, B. — 2021-12-23

Progressive tissue remodeling after myocardial infarction (MI) promotes cardiac arrhythmias. This process is well studied in young animals, but little is known about pro-arrhythmic changes in aged animals. Senescent cells accumulate with age and accelerate age-associated diseases. Senescent cells interfere with cardiac function and outcome post-MI with age, but studies have not been performed in large animals, and the mechanisms are unknown. Here, we investigated the role of senescence in regulating inflammation, fibrosis, and arrhythmogenesis in young and aged infarcted rabbits. Aged rabbits exhibited increased peri-procedural mortality and arrhythmogenic electrophysiological remodeling at the infarct border zone (IBZ) compared to young rabbits. Studies of the aged infarct zone revealed persistent myofibroblast senescence and increased inflammatory signaling over a twelve-week timecourse. Senescent IBZ myofibroblasts in aged rabbits appear to be coupled to myocytes, and our computational modeling showed that senescent myofibroblast- cardiomyocyte coupling prolongs action potential duration (APD) and facilitates conduction block permissive of arrhythmias. Aged infarcted human ventricles show levels of senescence consistent with aged rabbits, and senescent myofibroblasts also couple to IBZ myocytes. Our findings suggest that senolytic drugs may mitigate arrhythmias post-MI.

A phase-shifting anterior-posterior network organizes global phase relations

Menceloglu, M. — 2021-12-23

Prior research has identified a variety of task-dependent networks that form through inter-regional phase-locking of oscillatory activity that are neural correlates of specific behaviors. Despite ample knowledge of task-specific functional networks, general rules governing global phase relations have not been investigated. To discover such general rules, we focused on phase modularity, measured as the degree to which global phase relations in EEG comprised distinct synchronized clusters interacting with one another at large phase lags. Synchronized clusters were detected with a standard community-detection algorithm, and the degree of phase modularity was quantified by the index q. Notably, we found that the mechanism controlling phase modularity is remarkably simple. A network comprising anterior-posterior long-distance connectivity coherently shifted phase relations from low-angles (|{Delta}{theta}| < {pi}/4) in low-modularity states (bottom 5% in q) to high-angles (|{Delta}{theta}| > 3{pi}/4) in high-modularity states (top 5% in q), accounting for fluctuations in phase modularity. This anterior-posterior network may play a fundamental functional role as (1) it controls phase modularity across a broad range of frequencies (3-50 Hz examined) in different behavioral conditions (resting with the eyes closed or watching a silent nature video) and (2) neural interactions (measured as power correlations) in beta-to-gamma bands were consistently elevated in high-modularity states. These results may motivate future investigations into the functional roles of phase modularity as well as the anterior-posterior network that controls it.

Efficient learning in children with rapid GABA boosting after training

Frank, S. M. — 2022-01-02

It is unclear why and how children learn more efficiently than adults, although inhibitory systems, which play an important role in stabilizing learning, are immature in children. Here, we found that despite a lower baseline concentration of {gamma}-aminobutyric acid (GABA) in early visual cortical areas in children (8 to 11 years old) than adults (18 to 35 years old), children exhibited a rapid boost of GABA immediately after visual training, whereas the concentration of GABA in adults remained unchanged after training. Moreover, behavioral experiments showed that children stabilized visual learning much faster than adults, showing rapid development of resilience to retrograde interference. These results together suggest that inhibitory systems in childrens brains are more dynamic and adapt more quickly to stabilize learning than in adults. One Sentence SummaryChildren learn more efficiently than adults due to faster stabilization of learning with rapid GABA boosting after training.

Royal knifefish generate powerful suction feeding through large neurocranial elevation and high muscle power output

Li, E. Y. — 2022-01-20

Suction feeding in ray-finned fishes involves powerful buccal cavity expansion to accelerate water and food into the mouth. Previous XROMM studies in largemouth bass (Micropterus salmoides), bluegill sunfish (Lepomis macrochirus), and channel catfish (Ictalurus punctatus) have shown that more than 90% of suction power in high performance strikes comes from the axial musculature. Thus, the shape of the axial muscles and skeleton may impact suction feeding mechanics. Royal knifefish (Chitala blanci) have an unusual postcranial morphology, with a ventrally flexed vertebral column and relatively large mass of epaxial muscle. Based on their body shape, we hypothesized that royal knifefish would generate high power strikes by utilizing large neurocranial elevation, vertebral column extension, and epaxial shortening. As predicted, C. blanci generated high suction expansion power compared to the other three species studied to date (up to 160 W), which was achieved by increasing both the rate of volume change and the intraoral subambient pressure. The large epaxial muscle (25% of body mass) shortened at high velocities to produce large neurocranial elevation and vertebral extension (up to 41 deg, combined), as well as high muscle mass-specific power (up to 800 W kg-1). For the highest power strikes, axial muscles generated 95% of the power, and 64% of the axial muscle mass consisted of the epaxial muscles. The epaxial-dominated suction expansion of royal knifefish supports our hypothesis that postcranial morphology may be a strong predictor of suction feeding biomechanics. SUMMARY STATEMENTRoyal knifefish rely on their distinct postcranial morphology--with a curved vertebral column and large dorsal body muscles--to produce large neurocranial elevation and powerful suction feeding.

Impact of ionizing radiation on the environmental microbiomes of Chernobyl wetlands

Videvall, E. — 2022-01-20

Radioactive contamination in the form of ionizing radiation can be a devastating pollutant because it has the potential to cause damage to DNA and other biomolecules. Anthropogenic sources of ionizing radiation include accidents in nuclear power plants, such as the one in Chernobyl 1986, which caused long-term radioactive pollution. Studies on animals within radioactive zones have provided us with a greater understanding of how wildlife can persevere despite chronic radiation exposure, however, we still know very little about the effects of radiation on the microbial communities in the environment. Here, we examined the impact of ionizing radiation and other environmental factors on the diversity and composition of environmental microbiomes in the wetlands of Chernobyl. We combined extensive field sampling along a gradient of radiation together with 16S rRNA high-throughput metabarcoding (Illumina NovaSeq). While radiation did not affect the alpha diversity of the microbiomes in sediment, soil, or water, it had a strong effect on the beta diversity, indicating that the microbial composition was affected by ionizing radiation. Specifically, we detected several microbial taxa that were more abundant in areas with high radiation levels within the Chernobyl Exclusion Zone, including bacteria and archaea known to be radioresistant. Overall, our results reveal the existence of rich and diverse microbiomes in Chernobyl wetlands, with multiple taxonomic groups that are able to thrive despite the radioactive contamination. Further field and laboratory-based approaches will help to forecast the functionality and re-naturalization dynamics of radiocontaminated environments.

Host Chitinase 3-like-1 is a Universal Therapeutic Target for the Delta, Omicron and Other SARS-CoV-2 Viral Variants in COVID 19

Kamle, S. — 2022-01-24

COVID 19 is the disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2; SC2) which has caused a world-wide pandemic with striking morbidity and mortality. Evaluation of SC2 strains demonstrated impressive genetic variability and many of these viral variants are now defined as variants of concern (VOC) that cause enhanced transmissibility, decreased susceptibility to antibody neutralization or therapeutics and or the ability to induce severe disease. Currently, the delta ({delta}) and omicron (o) variants are particularly problematic based on their impressive and unprecedented transmissibility and ability to cause break through infections. The delta variant also accumulates at high concentrations in host tissues and has caused waves of lethal disease. Because studies from our laboratory have demonstrated that chitinase 3-like-1 (CHI3L1) stimulates ACE2 and Spike (S) priming proteases that mediate SC2 infection, studies were undertaken to determine if interventions that target CHI3L1 are effective inhibitors of SC2 viral variant infection. Here we demonstrate that CHI3L1 augments epithelial cell infection by pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins and that the CHI3L1 inhibitors anti-CHI3L1 and kasugamycin inhibit epithelial cell infection by these VOC pseudovirus moieties. Thus, CHI3L1 is a universal, VOC-independent therapeutic target in COVID 19.

Extraversion is associated with lower brain amyloid deposition in cognitively normal older adults.

Oh, H. — 2022-01-23

Emerging evidence suggests that some personality traits may link to the vulnerability to or protection for Alzheimers disease (AD). A causal mechanism underlying this relationship, however, remains largely unknown. Using 18F-Florbetaben positron emission tomography (PET) binding to beta-amyloid (A{beta}) plaques, a pathological feature of AD, and functional magnetic resonance imaging (fMRI), we investigated pathological and functional correlates of extraversion and neuroticism in a group of healthy young and older subjects. We quantified a level of brain A{beta} deposition in older individuals. Brain activity was measured in young adults using a task-switching fMRI paradigm. When we correlated personality scores of extraversion and neuroticism with these pathological and functional measures, higher extraversion, but not neuroticism, was significantly associated with lower global A{beta} measures among older adults, accounting for age and sex. This association was present across widespread brain regions. Among young subjects, higher extraversion was associated with lower activity during task switching in anterior cingulate cortex, left anterior insular cortex, left putamen, and middle frontal gyrus bilaterally, while higher neuroticism was associated with increased activity throughout the brain. The present results suggest that possibly via efficient neuronal activity, extraversion, one of lifelong personality traits, may confer the protective mechanism against the development of A{beta} pathology during aging.

Sex-Specific Genes Identified in Sea Urchin Gonads are Expressed Prior to Metamorphosis

Pieplow, C. — 2022-01-28

A great collaboration between the germline and somatic cells of an organism is the creation of a functional gonad. Here we begin to test this mechanism in a sea urchin by use of RNA-seq, and quantitative mRNA measurements throughout the life cycle of the sea urchin, Lytechinus variegatus (Lv). We found through de novo analyses that the ovary and testis of this echinoderm contains the many transcripts predicted for gamete morphology and function, but we also discovered many uncharacterized gene products unique to each gonad. We found that transcripts involved in glycolysis are highly enriched in the ovary compared to the testis, and that Lv has an expansion of nanos genes not observed in other sequenced echinoderms. We developed a pipeline integrating timepoint RNA-seq data throughout development to identify hallmark gene expression in gonads. We found activation of meiotic genes surprisingly early in development, and observed that candidate genes involved in sex determination are first expressed during larval growth, well before metamorphosis. We further discovered that individual larvae express varying levels of male- or female-hallmark genes before metamorphosis, including factors for the germ line, oocyte, sperm, and meiosis related genes. Together these data support the hypothesis that sex determination in the sea urchin is initiated prior to metamorphosis, from a shared profile of male and female factors, and that the many uncharacterized genes unique to the gonads may reveal unique pathways and mechanisms in echinoderm reproduction. HighlightO_LITestis and Ovary of the sea urchin, Lytechinus variegatus, have distinct transcriptome profiles C_LIO_LIL. variegatus gonad RNA-seq reveals the expression of 9 nanos genes C_LIO_LIThe ovary in the sea urchin uniquely expresses genes of the glycolytic pathway, including an ovary-specific transcript of GAPDH C_LIO_LIGenes involved in meiosis and in sex determination are expressed early in larval development C_LIO_LIFrom gene expression profiles we conclude here that sex determination is initiated prior to metamorphosis in this echinoderm C_LI

Investigating the performance of deep learning methods for Hi-C resolution improvement

Murtaza, G. — 2022-01-31

MotivationHi-C is a widely used technique to study the 3D organization of the genome. Due to its high sequencing cost, most of the generated datasets are of coarse resolution, which makes it impractical to study finer chromatin features such as Topologically Associating Domains (TADs) and chromatin loops. Multiple deep-learning-based methods have recently been proposed to increase the resolution of these data sets by imputing Hi-C reads (typically called upscaling). However, the existing works evaluate these methods on either synthetically downsampled or a small subset of experimentally generated sparse Hi-C datasets, making it hard to establish their generalizability in the real-world use case. We present our framework - Hi-CY - that compares existing Hi-C resolution upscaling methods on seven experimentally generated low-resolution Hi-C datasets belonging to various levels of read sparsities originating from three cell lines on a comprehensive set of evaluation metrics. Hi-CY also includes four downstream analysis tasks, such as TAD and chromatin loops recall, to provide a thorough report on the generalizability of these methods. ResultsWe observe that existing deep-learning methods fail to generalize to experimentally generated sparse Hi-C datasets showing a performance reduction of up to 57 %. As a potential solution, we find that retraining deep-learning based methods with experimentally generated Hi-C datasets improves performance by up to 31%. More importantly, Hi-CY shows that even with retraining, the existing deep-learning based methods struggle to recover biological features such as chromatin loops and TADs when provided with sparse Hi-C datasets. Our study, through Hi-CY framework, highlights the need for rigorous evaluation in future. We identify specific avenues for improvements in the current deep learning-based Hi-C upscaling methods, including but not limited to using experimentally generated datasets for training. Availabilityhttps://github.com/rsinghlab/Hi-CY Author SummaryWe evaluate deep learning-based Hi-C upscaling methods with our framework Hi-CY using seven datasets originating from three cell lines evaluated using three correlation metrics, four Hi-C similarity metrics, and four downstream analysis tasks, including TAD and chromatin loop recovery. We identify a distributional shift between Hi-C contact matrices generated from downsampled and experimentally generated sparse Hi-C datasets. We use Hi-CY to establish that the existing methods trained with downsampled Hi-C datasets tend to perform significantly worse on experimentally generated Hi-C datasets. We explore potential strategies to alleviate the drop in performance such as retraining models with experimentally generated datasets. Our results suggest that retraining improves performance up to 31 % on five sparse GM12878 datsets but provides marginal improvement in cross cell-type setting. Moreover, we observe that regardless of the training scheme, all deep-learning based methods struggle to recover biological features such as TADs and chromatin loops when provided with very sparse experimentally generated datasets as inputs.

Evidence for independent processing of shape by vision and touch

Miller, R. L. — 2022-02-04

Although visual object recognition is well-studied and relatively well understood, much less is understood about how shapes are recognized by touch and how such haptic stimuli might be compared to visual shapes. One might expect that the processes of visual and haptic object recognition engage identical brain structures given the similarity of the problem and the advantages of avoiding redundant brain circuitry, but it has not yet been established the extent to which this is true. We recruited human participants to perform a one-back same-different visual and haptic shape comparison task both within (i.e., comparing two visual shapes or two haptic shapes) and across (i.e., comparing visual with haptic shapes) modalities. Participants saw or felt a shape and responded according to whether they thought the shape was the same or different from the previously felt or seen shape. We then used various shape metrics to predict performance based on the shape, orientation, and modality of the two stimuli which were being compared on each trial. We found that the fixed orientation of the shape stimuli was an important factor for predicting within-modal behavior, but the orientation of shapes compared across modality did not depend on knowing the presented orientation. We also found that the metrics which best predict shape comparison behavior heavily depended on the modality of the two shapes. We take these results as evidence that object recognition is not necessarily performed in a single, modality-agnostic region.

PELP: accounting for missing data in neural time series by Periodic Estimation of Lost Packets

Dastin-van Rijn, E. — 2022-02-08

ObjectiveImplanted electrical stimulators with sensing capabilities have enabled the development of closed-loop neuromodulation therapies capable of responding to patient needs in real-time. Through a combination of rechargeable technologies and wireless data transmission, it is now possible for researchers to acquire extensive neural recordings from human participants in naturalistic settings using these bidirectional devices. However, data losses during wireless transmission hamper processing and the identification of neural signals of interest, driving the need for methodologies to properly estimate the impact of data loss. ApproachTo accurately reconstruct the timing of data containing losses, we have developed a method called Periodic Estimation of Lost Packets (PELP) to precisely determine the number of samples lost from implanted recordings during active stimulation. PELP leverages a data-driven procedure for determining the period of stimulation and the knowledge that stimulation continues identically during periods where data are missing to accurately account for the number of samples lost. Main resultsUsing simulated stimulation added to collected human EEG data, we show that PELP is robust to a range of stimulation waveforms and noise characteristics. Lastly, we successfully applied PELP to local field potential (LFP) recordings from an implanted, bidirectional device using data recorded in the clinic and the patients own home. SignificanceBy effectively accounting for the timing of missing data, PELP enables the analysis of complex, naturalistic neural time series data from bidirectional implanted devices aiding in the development of novel therapeutic approaches. NCT04806516 (ClinicalTrials.gov).

Distinct mechanisms of germ cell factor regulation for an inductive germ cell fate

Foster, S. — 2022-02-06

Specification of primordial germ cells (PGCs), the lineage which gives rise to eggs and sperm, is essential for sexually reproducing organisms. The mechanism by which animals specify their PGCs generally falls into two categories: inherited or inductive. The inductive mechanism, used by mammals, relies on cell signaling interactions to direct a subset of embryonic cells to a germ cell fate. Previous work suggested that sea star embryos, which develop in simple culture and are markedly transparent, also use inductive mechanisms to specify their germline. The germ cell factors Nanos and Vasa become restricted during early development into a localized region of cells within the posterior enterocoel (PE), the presumptive germline. Nodal signaling was observed to negatively regulate Vasa and Nanos mRNAs outside of the PE and restrict the germline to the PE. Here we employed single cell RNA sequencing to identify the transcriptional program of germ cells and their changes during development. We never see Nodal pathway members within Nanos/Vasa positive cells in the region known to give rise to the PE, and instead see members of the Wnt-signaling pathway and the FoxY family of transcription factors. We learned that Wnt and Delta/Notch signaling enhances expression of both Nanos and Vasa, whereas a test of cell interactions reveals that Nanos and Vasa are regulated distinctly. This work provides insights into the sequence of events that leads to PGC specification and enables deeper mechanistic studies in a tractable in vivo model. HighlightsO_LISingle-cell RNA-sequencing of sea star embryos demonstrates temporal differences in cell fate commitment among echinoderms. C_LIO_LISea urchin and sea star embryos appear to ascribe their germ line by two extreme different mechanisms but share similar pathways in regulation of the germline genes. C_LIO_LIExpression of the germline factors, Vasa and Nanos, is regulated by distinct mechanisms in the sea star. C_LIO_LIGermline induction in the sea star uses similar signaling mechanisms as mammals. C_LI

The Requirement of Ubiquitin C-Terminal Hydrolase L1 (UCHL1) in Mouse Ovarian Development and Fertility

Woodman, M. F. — 2022-02-10

Ubiquitin C-Terminal Hydrolase L1 (UCHL1) is a de-ubiquitinating enzyme enriched in neuronal and gonadal tissues known to regulate the cellular stores of mono-ubiquitin and protein turnover. While its function in maintaining proper motor neuron function is well-established, investigation into its role in the health and function of reproductive processes is only just beginning to be studied. Single-cell-sequencing analysis of all ovarian cells from the murine perinatal period revealed that Uchl1 is very highly expressed in the developing oocyte population, an observation which was corroborated by high levels of oocyte-enriched UCHL1 protein expression in oocytes of all stages throughout the mouse reproductive lifespan. To better understand the role UCHL1 may be playing in oocytes, we utilized a UCHL1-deficient mouse line, finding reduced number of litters, reduced litter sizes, altered folliculogenesis, morphologically abnormal oocytes, disrupted estrous cyclicity and apparent endocrine dysfunction in these animals compared to their wild-type and heterozygous littermates. These data reveal a novel role of UCHL1 in female fertility as well as overall ovarian function, and suggest a potentially essential role for the ubiquitin proteasome pathway in mediating reproductive health. Summary sentenceUbiquitin C-Terminal Hydrolase L1 (UCHL1) is required for proper ovarian folliculogenesis, estrous cyclicity, and fertility in the female mouse.

Decoding Dynamically Shifting States of Parkinson's Disease: Tremor, Bradykinesia, and Effective Motor Control

Lauro, P. M. — 2022-02-14

Parkinsons Disease (PD) is characterized by distinct motor phenomena that are expressed asynchronously. Understanding the neurophysiological correlates of these different motor states could facilitate monitoring of disease progression and allow improved assessments of therapeutic efficacy, as well as enable optimal closed-loop neuromodulation. We examined neural activity in the basal ganglia and cortex of subjects with PD during a quantitative motor task to decode tremor and bradykinesia -- two cardinal motor signs of this disease -- and relatively asymptomatic periods of behavior. Analysis of subcortical and cortical signals revealed that tremor and bradykinesia had distinct, nearly opposite neural signatures, while effective motor control displayed unique, differentiating features. The neurophysiological signatures of these motor states depended on the type of signal recorded as well as the location; cortical decoding accuracy generally outperformed subcortical decoding, while tremor and bradykinesia were better decoded from different portions of the subthalamic nucleus (STN). These results provide a roadmap to leverage real-time neurophysiology to understand and treat PD. One Sentence SummaryMotor signs of Parkinsons disease such as tremor and bradykinesia can be independently decoded from subthalamic and cortical recordings.

Enabling interpretable machine learning for biological data with reliability scores

Ahlquist, K. D. — 2022-02-21

Machine learning has become an important tool across biological disciplines, allowing researchers to draw conclusions from large datasets, and opening up new opportunities for interpreting complex and heterogeneous biological data. Alongside the rapid growth of machine learning, there have also been growing pains: some models that appear to perform well have later been revealed to rely on features of the data that are artifactual or biased; this feeds into the general criticism that machine learning models are designed to optimize model performance over the creation of new biological insights. A natural question thus arises: how do we develop machine learning models that are inherently interpretable or explainable? In this manuscript, we describe reliability scores, a new concept for scientific machine learning studies that assesses the ability of a classifier to produce a reliable classification for a given instance. We develop a specific implementation of a reliability score, based on our work in Sugden et al. 2018 in which we introduced SWIF(r), a generative classifier for detecting selection in genomic data. We call our implementation the SWIF(r) Reliability Score (SRS), and demonstrate the utility of the SRS when faced with common challenges in machine learning including: 1) an unknown class present in testing data that was not present in training data, 2) systemic mismatch between training and testing data, and 3) instances of testing data that are missing values for some attributes. We explore these applications of the SRS using a range of biological datasets, from agricultural data on seed morphology, to 22 quantitative traits in the UK Biobank, and population genetic simulations and 1000 Genomes Project data. With each of these examples, we demonstrate how interpretability tools for machine learning like the SRS can allow researchers to interrogate their data thoroughly, and to pair their domain-specific knowledge with powerful machine-learning frameworks. We hope that this tool, and the surrounding discussion, will aid researchers in the biological machine learning space as they seek to harness the power of machine learning without sacrificing rigor and biological understanding.

Uncertainty Quantification in Variable Selection for Genetic Fine-Mapping using Bayesian Neural Networks

Cheng, W. — 2022-02-25

In this paper, we propose a new approach for variable selection using a collection of Bayesian neural networks with a focus on quantifying uncertainty over which variables are selected. Motivated by fine-mapping applications in statistical genetics, we refer to our framework as an "ensemble of single-effect neural networks" (ESNN) which generalizes the "sum of single-effects" regression framework by both accounting for nonlinear structure in genotypic data (e.g., dominance effects) and having the capability to model discrete phenotypes (e.g., case-control studies). Through extensive simulations, we demonstrate our methods ability to produce calibrated posterior summaries such as credible sets and posterior inclusion probabilities, particularly for traits with genetic architectures that have significant proportions of non-additive variation driven by correlated variants. Lastly, we use real data to demonstrate that the ESNN framework improves upon the state-of-the-art for identifying true effect variables underlying various complex traits.

A novel missense mutation in the proprotein convertase gene furinb causes hepatic cystogenesis during liver development in zebrafish

Ellis, J. L. — 2022-02-26

Hepatic cysts are fluid-filled lesions in the liver that are estimated to occur in 5% of the population. They may cause hepatomegaly and abdominal pain. Progression to secondary fibrosis, cirrhosis, or cholangiocarcinoma can lead to morbidity and mortality. Previous studies of patients and rodent models have associated hepatic cyst formation with increased proliferation and fluid secretion in cholangiocytes, which are partially due to impaired primary cilia. Congenital hepatic cysts are thought to originate from faulty bile duct development, but the underlying mechanisms are not fully understood. In a forward genetic screen, we identified a zebrafish mutant that develops hepatic cysts during larval stages. Cyst formation in these mutants is not due to changes in biliary cell proliferation, bile secretion, or impairment of primary cilia. Instead, time-lapse live imaging data showed that the mutant biliary cells failed to form interconnecting bile ducts because of defects in motility and protrusive activity. Accordingly, immunostaining revealed an excessive and disorganized actin and microtubule cytoskeleton in the mutant biliary cells. By whole-genome sequencing, we determined that the cystic phenotype in the mutant was caused by a missense mutation in the furinb gene which encodes a proprotein convertase. The mutation alters Furinb localization and causes endoplasmic reticulum (ER) stress. The cystic phenotype could be suppressed by treatment with the ER stress inhibitor 4-phenylbutyric acid and exacerbated by treatment with the ER stress inducer tunicamycin. The mutant livers also exhibited increased mTOR signaling and treatment with the mTOR inhibitor rapamycin partially blocked cyst formation by reducing ER stress. Our study has established a novel vertebrate model for studying hepatic cystogenesis and illustrated the role of ER stress in the disease pathogenesis.

STING activation promotes robust immune response and tumor regression in glioblastoma models

Berger, G. — 2022-03-01

Immunotherapy has had a tremendous impact on cancer treatment in the past decade, with hitherto unseen responses at advanced and metastatic stages of the disease. However, the aggressive brain tumor glioblastoma (GBM) is highly immunosuppressive and remains largely refractory to current immunotherapeutic approaches. The cGAS-STING cytoplasmic double stranded DNA (dsDNA) sensing pathway has emerged as a next-generation immunotherapy target with potent local immune stimulatory properties. Here, we investigated the status of the STING pathway in GBM and the modulation of the brain tumor microenvironment (TME) with the STING agonist ADU-S100. Our data reveal the presence of STING in human GBM specimens, where it stains strongly in the tumor vasculature. We show that human GBM explants can respond to STING agonist treatment by secretion of inflammatory cytokines. In murine GBM models, we show a profound shift in the tumor immune landscape after STING agonist treatment, with massive infiltration of the tumor-bearing hemisphere with innate immune cells including inflammatory macrophages, neutrophils and NK populations. Treatment of established murine intracranial GL261 and CT-2A tumors by biodegradable ADU-S100-loaded intracranial implants demonstrated a significant increase in survival in both models and long-term survival with immune memory in GL261. Responses to treatment were abolished by NK cell depletion. This study reveals therapeutic potential and deep remodeling of the TME by STING activation in GBM and warrants the further examination of STING agonists alone or in combination with other immunotherapies such as cancer vaccines, CAR T cells, NK therapies or immune checkpoint blockade. Significance statementModulation of the immune microenvironment is critical for immunosuppressive and therapy refractory tumors like glioblastoma. Activation of the STING pathway deeply remodels the brain tumor environment and attracts innate immune cells and natural killer cell populations, producing a robust antitumor effect with long-term immune memory. We further show that human glioblastoma tissue can respond to the therapy and lay the foundations for combined intracranial immunotherapies by using crosslinked biodegradable brain implants.

On the normative advantages of dopamine and striatal opponency for learning and choice

Frank, M. J. — 2022-03-13

The basal ganglia (BG) contribute to reward-driven behavior, but it is unclear why such behavior should rely on notoriously complex circuitry involving opponent striatal pathways. Dopamine (DA) nonlinearly modulates activity and plasticity in the BG and is itself dynamically modulated across tasks. We develop the OpAL* model to assess the normative advantages of such circuitry. In OpAL*, dynamic DA modulation differentially amplifies the striatal pathway most tuned to the accumulated history of positive or negative outcomes. This efficient coding mechanism avoids a vexing explore-exploit tradeoff that plagues traditional RL models in sparse reward environments. OpAL* exhibits robust advantages over alternative models across a range of environments, particularly with sparse reward. These advantages depend on opponent and nonlinear Hebbian plasticity mechanisms previously thought to be pathological. Finally, OpAL* captures risky choice patterns arising from DA and environmental manipulations across species, suggesting that they result from a normative biological mechanism. Everybody wants the most they can possibly get For the least they can possibly do - Todd Snider, "Easy Money"

Neurons in inferior temporal cortex are sensitive to motion trajectory during degraded object recognition

Burk, D. C. — 2022-03-14

Our brains continuously acquire sensory information and make judgments even when visual information is limited. In some circumstances, an ambiguous object can be recognized from how it moves, such as an animal hopping or a plane flying overhead. Yet it remains unclear how movement is processed by brain areas involved in visual object recognition. Here we investigate whether inferior temporal cortex, an area traditionally known for shape processing, has access to motion information during degraded shape recognition. We developed a matching task that required monkeys to recognize moving shapes with variable levels of shape degradation. Neural recordings in area IT showed that, surprisingly, some IT neurons preferred blurry shapes over clear ones. Further, many of the neurons exhibited motion sensitivity at different times during the presentation of the blurry target. Population decoding analyses showed that motion pattern could be decoded from IT neuron pseudo-populations. Contrary to previous findings, these results suggest that neurons in IT can integrate visual motion and shape information, particularly when shape information is degraded, in a way that has been previously overlooked. Our results highlight the importance of using challenging multi-feature recognition tasks to understand the role of area IT in naturalistic visual object recognition. (Word count: 199)

Leveraging shared ancestral variation to detect local introgression

Lopez Fang, L. — 2022-03-21

Introgression is a common evolutionary phenomenon that results in shared genetic material across non-sister taxa. Existing statistical methods such as Pattersons D statistic can detect introgression by measuring an excess of shared derived alleles between populations. The D statistic is effective to detect genome-wide patterns of introgression but can give spurious inferences of introgression when applied to local regions. We propose a new statistic, D+, that leverages both shared ancestral and derived alleles to infer local introgressed regions. Incorporating both shared derived and ancestral alleles increases the number of informative sites per region, improving our ability to identify local introgression. We use a coalescent framework to derive the expected value of this statistic as a function of different demographic parameters under an instantaneous admixture model and use coalescent simulations to compute the power and precision of D+. While the power of D and D+ is comparable, D+ has better precision than D. We apply D+ to empirical data from the 1000 Genome Project and Heliconius butterflies to infer local targets of introgression in humans and in butterflies.

Connexin36 expression in the cochlear nucleus complex of the echolocating bat, Eptesicus fuscus

Accomando, A. W. — 2022-03-25

Gap junctions and electrical synapses in the central nervous system are associated with rapid temporal processing and coincidence detection. Using histology, immunohistochemistry, and in situ hybridization, we investigated the distribution of Connexin36 (Cx36), a protein that comprises neuronal gap junctions, throughout the cochlear nucleus complex of the echolocating big brown bat, Eptesicus fuscus, a species exhibiting extreme behavioral sensitivity to minute temporal changes in ultrasonic echoes. For comparison, we visualized Cx36 expression in the cochlear nucleus of transgenic Cx36 reporter mice, species that hear ultrasound but do not echolocate. We observed Cx36 expression in the anteroventral and dorsal cochlear nucleus, with more limited expression in the posteroventral cochlear nucleus, of both species. Several different morphological cell types were labeled, including globular and spherical bushy, octopus, stellate, and fusiform cells. Labeled Cx36 puncta were also observed. Cx36 expression in the bat was spread throughout a relatively smaller area of the cochlear nucleus than in the mouse, even though the bat cochlear nucleus is hypertrophied. In the bat, the anteroventral cochlear nucleus showed higher percent area label than the dorsal cochlear nucleus, with a trend towards the opposite result in the mouse. The presence of gap junctions appears to be a conserved feature of the mammalian cochlear nucleus and thus not uniquely tied to the temporal hyperacuity of echolocation.

Neuroimaging within the Dominantly Inherited Alzheimer's Network (DIAN): PET and MRI

McKay, N. S. — 2022-03-30

The Dominantly Inherited Alzheimer Network (DIAN) Observational Study is an international collaboration studying autosomal dominant Alzheimer disease (ADAD). This rare form of Alzheimer disease (AD) is caused by mutations in the presenilin 1 (PSEN1), presenilin 2 (PSEN2), or amyloid precursor protein (APP) genes. As individuals from these families have a 50% chance of inheriting the familial mutation, this provides researchers with a well-matched cohort of carriers vs non-carriers for case-control studies. An important trait of ADAD is that the age at symptom onset is highly predictable and consistent for each specific mutation, allowing researchers to estimate an individuals point in their disease time course prior to symptom onset. Although ADAD represents only a small proportion (approximately 0.1%) of all AD cases, studying this form of AD allows researchers to investigate preclinical AD and the progression of changes that occur within the brain prior to AD symptom onset. Furthermore, the young age at symptom onset (typically 30-60 years) means age-related comorbidities are much less prevalent than in sporadic AD, thereby allowing AD pathophysiology to be studied independent of these confounds. A major goal of the DIAN Observational Study is to create a global resource for AD researchers. To that end, the current manuscript provides an overview of the DIAN magnetic resonance imaging (MRI) and positron emission tomography (PET) protocols and highlights the key imaging results of this study to date.

The Human Motoneuron Expression Signature is Defined by ALS-Related Genes

Yadav, A. — 2022-03-28

The mammalian spinal cord functions as a community of glial and neuronal cell types to accomplish sensory processing, autonomic control, and movement; conversely, the dysfunction of these cell types following spinal cord injury or disease states can lead to chronic pain, paralysis, and death. While we have made great strides in understanding spinal cellular diversity in animal models, it is crucial to characterize human biology directly to uncover specialized features of basic function and to illuminate human pathology. Here, we present a cellular taxonomy of the adult human spinal cord using single nucleus RNA-sequencing with spatial transcriptomics and antibody validation. We observed 29 glial clusters, including rare cell types such as ependymal cells, and 35 neuronal clusters, which we found are organized principally by anatomical location. To demonstrate the potential of this resource for understanding human disease, we analyzed the transcriptome of spinal motoneurons that are prone to degeneration in amyotrophic lateral sclerosis (ALS) and other diseases. We found that, compared with all other spinal neurons, human motoneurons are defined by genes related to cell size, cytoskeletal structure, and ALS, thereby supporting a model of a specialized motoneuron molecular repertoire that underlies their selective vulnerability to disease. We include a publicly available browsable web resource with this work, in the hope that it will catalyze future discoveries about human spinal cord biology.

Small Molecules Restore Azole Activity Against Drug-Tolerant and Drug-Resistant Candida Isolates

Alabi, P. E. — 2022-03-31

Each year, fungi cause more than 1.5 billion infections worldwide and have a devastating impact on human health, particularly in immunocompromised individuals or patients in intensive care units. The limited antifungal arsenal and emerging multidrug resistant species necessitate the development of new therapies. One strategy for combating drug resistant pathogens is the administration of molecules that restore fungal susceptibility to approved drugs. Accordingly, we carried out a screen to identify small molecules that could restore the susceptibility of pathogenic Candida species to azole antifungals. This screening effort led to the discovery of novel 1,4-benzodiazepines that restore fluconazole susceptibility in resistant isolates of Candida albicans, as evidenced by 100-1000-fold potentiation of fluconazole activity. This potentiation effect was also observed in azole-tolerant strains of C. albicans and in other pathogenic Candida species. The 1,4-benzodiazepines selectively potentiated different azoles, but not other approved antifungals. A remarkable feature of the potentiation was that the combination of the compounds with fluconazole was fungicidal, whereas fluconazole alone is fungistatic. Interestingly, the potentiators were not toxic to C. albicans in the absence of fluconazole, but inhibited virulence-associated filamentation of the fungus. We found that the combination of the potentiators and fluconazole significantly enhanced host survival in a Galleria mellonella model of systemic fungal infection. Taken together, these observations validate a strategy wherein small molecules can restore the activity of highly used anti-infectives that have lost potency. IMPORTANCEIn the last decade, we have been witnessing a higher incidence of fungal infections, due to an expansion of the fungal species capable of causing disease (e.g., Candida auris), as well as increased antifungal drug resistance. Among human fungal pathogens, Candida species are a leading cause of invasive infections and are associated with high mortality rates. Infections by these pathogens are commonly treated with azole antifungals, yet the expansion of drug-resistant isolates have reduced their clinical utility. In this work, we describe the discovery and characterization of small molecules that potentiate fluconazole and restore the susceptibility of azole-resistant and azole-tolerant Candida isolates. Interestingly, the potentiating 1,4-benzodiazepines were not toxic to fungal cells but inhibited their virulence-associated filamentous growth. Furthermore, combinations of the potentiators and fluconazole decreased fungal burdens and enhanced host survival in a Galleria mellonella model of systemic fungal infections. Accordingly, we propose the use of novel antifungal potentiators as a powerful strategy for addressing the growing resistance of fungi to clinically approved drugs.

Emergence of distinct neural subspaces in motor cortical dynamics during volitional adjustments of ongoing locomotion

Xing, D. — 2022-04-05

3The brain is capable of simultaneously carrying out multiple functions, such as making different types of movements at the same time. One example is how we are able to both carry out stereotyped walking or running movements, while concurrently performing precise, target-directed movements such as kicking a ball in a soccer match. Recently, research has shown that different computations within the same population of neurons can be carried out without disrupting each other by confining the processes into separate subspaces. Whether this strategy is used to precisely control our limbs while maintaining locomotion is still an open question. Here, we recorded the activity of primary motor cortex in nonhuman primates during obstacle avoidance on a treadmill. We found that the same neural population was active during both basic unobstructed locomotion and volitional obstacle avoidance movements. Additionally, we identified the neural modes spanning the subspace of the low-dimensional dynamics in M1 using both supervised and unsupervised techniques. We found that motor cortex employs a subspace that consistently maintains the same cyclic activity throughout obstacle stepping, despite large changes in the movement itself. All the variance corresponding to the large change in movement during the obstacle avoidance is confined to its own distinct subspace. Our findings suggest that M1 utilizes different activity subspaces to coordinate the maintenance of ongoing locomotor-related neural dynamics and fast volitional gait adjustments during complex locomotion. 4 Significance StatementOur ability to modulate our ongoing walking gait with precise, voluntary adjustments is what allows us to navigate complex terrains. Locomotion and precise, goal-directed movements, such as reaching are two distinct movement modalities and have been shown to have differing requirements of motor cortical input. It is unknown how these two movements are represented in M1 low dimensional dynamics when both are carried out at the same time, such as during obstacle avoidance. We developed a novel obstacle avoidance paradigm in freely-moving non-human primates and discovered that the strategy employed by motor cortex is to confine the rhythmic locomotion-related dynamics and the voluntary, gait-adjustment movement into separate subspaces.

recolorize: improved color segmentation of digital images (for people with other things to do)

Weller, H. I. — 2022-04-05

Color is an important source of biological information in fields ranging from disease ecology to sexual selection. Despite its importance, most metrics for color are restricted to point measurements. Methods for moving beyond point measurements rely on color maps, where every pixel in an image is assigned to one of a set of discrete color classes (color segmentation). Manual methods for color segmentation are slow and subjective, while existing automated methods often fail due to biological variation in pattern, technical variation in images, and poor scalability for batch clustering. As a result, color segmentation is the common bottleneck step for a majority of existing downstream analyses. Here we present recolorize, an R package for color segmentation that succeeds in many cases where existing methods fail. Recolorize has three major components: (1) an effective two-part clustering algorithm where color distributions are binned and combined according to perceived similarity in a frequency-independent manner; (2) a toolkit for minor manual adjustments to automatic output where needed; and (3) flexible export options. This paper illustrates how to use recolorize and compares it to existing methods, including examples where we segment formerly intractable images, and demonstrates the downstream use of methods that rely on color maps.

Neuronal HLH-30/TFEB modulates muscle mitochondrial fragmentation to improve thermoresistance in C. elegans

Wong, S. Q. — 2022-04-10

Transcription factor EB (TFEB) is a conserved master transcriptional activator of autophagy and lysosomal genes that modulates organismal lifespan regulation and stress resistance. As neurons can coordinate organism-wide mechanisms, we investigated the role of neuronal TFEB in stress resistance and longevity. To this end, the C. elegans TFEB orthologue, hlh-30, was rescued panneuronally in hlh-30 loss of function mutants. While important in the long lifespan of daf-2 animals, neuronal hlh-30 was not sufficient to restore normal lifespan in short-lived hlh-30 mutants. However, neuronal HLH-30/TFEB rescue mediated robust improvements in the heat stress resistance of wild-type but not daf-2 animals. Notably, these mechanisms can be uncoupled, as neuronal HLH-30/TFEB regulates longevity and thermoresistance dependently and independently of DAF-16/FOXO respectively. Through transcriptomics profiling and functional analysis, we identified the uncharacterized gene W06A11.1 as a bona fide mediator of heat stress resistance via the induction of mitochondrial fragmentation in distal muscles. Neuron-to-muscle communication occurred through a modulation of neurotransmission. Taken together, this study uncovers a novel mechanism of heat stress protection mediated by neuronal HLH-30/TFEB.

Intrinsically photosensitive retinal ganglion cells evade temporal filtering to encode environmental light intensity

Sabbah, S. — 2022-04-10

The retina encodes environmental light intensity to drive innate physiological responses. The synaptic basis of such coding remains obscure. Intrinsically photosensitive retinal ganglion cells (ipRGCs) are the only retinal output neurons stably encoding intensity. They do so even without their melanopsin photopigment, so specializations in their synaptic drive from bipolar cells (BCs) must also contribute. Here, we shed new light on mechanisms responsible for this unique intensity-coding drive. By ultrastructural reconstruction, we show that specific BC types and unusual ribbon synapses carry photoreceptor signals to ipRGCs. By glutamate imaging and electrophysiology, we show that their light responses are unusually persistent. Still, we find that virtually all BCs encode intensity. Intensity coding becomes restricted to ipRGCs primarily because other RGCs filter out steady-state intensity signals postsynaptically. Thus, neural "pinholes" in global, persistent neural "masking" allow intensity signals to be encoded by ipRGCs and sent to specific centers of the visual brain.

MMR Deficiency Defines Distinct Molecular Subtype of Breast Cancer with Unique Proteomic Networks and Variable Clinical Significance

Hacking, S. — 2022-04-15

Mismatch repair (MMR) alterations are important prognostic and predictive biomarkers in a variety of cancer subtypes including colorectal and endometrial. However, in breast cancer (BC), the distinction and clinical significance of MMR is largely unknown. This may be due in part to the fact that genetic alterations in MMR genes are rare, and only seen to occur in around 3% of BCs. In the present study we analyzed TCGA data using a multi-sample protein-protein interactions (PPI) analysis tool, Proteinarium, and showed a distinct separation in the MMR deficient and intact specific networks. MMR deficient tumor specific networks have a highly connected cluster of histone genes, identified by unique PPI. We also found that distribution of MMR deficient breast cancer is more prevalent in HER2-enriched and triple-negative (TN) BC subtypes compared to luminal BCs. Poorer survival was seen in patients with HER2-enriched BCs with MMR deficiency, whereas an improved survival was seen in TNBCs with MMR deficiency. We recommend defining MMR deficient breast cancer by next generation sequencing (NGS) when any somatic mutation is detected in one of the 7 MMR genes found in our study. Our recommendations include labeling patients with variants of undetermined significance (VUS) as MMR deficient supported by findings from distinct clusters of patients based on our network analysis. MMR may have a role in guiding the use of immunotherapy for both TN as well as HER2-enriched BC.

Predicting A/B compartments from histone modifications using deep learning

Zheng, S. — 2022-04-19

Genomes fold into organizational units in the 3D space that can influence critical biological functions. In particular, the organization of chromatin into A and B compartments segregates its active regions from inactive regions. Compartments, evident in Hi-C contact matrices, have been used to describe cell-type specific changes in the A/B organization. However, obtaining Hi-C data for all cell and tissue types of interest is prohibitively expensive, which has limited the widespread consideration of compartment status. We present a prediction tool called Compartment prediction using Recurrent Neural Network (CoRNN) that models the relationship between the compartmental organization of the genome and histone modification enrichment. Our model predicts A/B compartments, in a cross-cell type setting, with an average area under the ROC curve of 90.9%. Our cell type-specific compartment predictions show high overlap with known functional elements. We investigate our predictions by systematically removing combinations of histone marks and find that H3K27ac and H3K36me3 are the most predictive marks. We then perform a detailed analysis of loci where compartment status cannot be accurately predicted from these marks. These regions represent chromatin with ambiguous compartmental status, likely due to variations in status within the population of cells. These ambiguous loci also show highly variable compartmental status between biological replicates in the same GM12878 cell type. Finally, we demonstrate the generalizability of our model by predicting compartments in independent tissue samples. Our software and trained model are publicly available at https://github.com/rsinghlab/CoRNN.

Development of transformation for genome editing of an emerging model organism

Yamamoto, Y. — 2022-04-19

With the advances in genomic sequencing, many organisms with novel biological properties are ripe for use as emerging model organisms. However, to make full use of them, transformation methods need to be developed to permit genome editing. Here, we present development of transformation for the fungus fly Sciara (Bradysia) coprophila; this may serve as a paradigm for development of transformation for other emerging systems, especially insects. Sciara has a variety of unique biological features including locus-specific developmentally-regulated DNA amplification; chromosome imprinting; a monopolar spindle in male meiosis I; non-disjunction of the X chromosome in male meiosis II; X chromosome elimination in early embryogenesis; germ line limited (L) chromosomes; high resistance to radiation. Mining the unique biology of Sciara requires a transformation system to test mutations of DNA sequences that may play roles for these features. We describe a Sciara transformation system using a modified piggyBac transformation vector and detailed protocols we have developed to accommodate Sciara-specific requirements. This advance will provide a platform for us and others in the growing Sciara community to take advantage of this unique biological system. In addition, the versatile piggyBac vectors described here and transformation methods will be useful for other emerging model systems. Author BiographiesSusan A. Gerbi (Ph.D. with Joseph Gall at Yale University 1970) is the George Eggleston, Professor of Biochemistry at Brown University. Her research includes chromosomes, DNA replication and ribosomal RNA. She was President and is a Fellow of ASCB, a Fellow of AAAS and received the RI Governors Award for Scientific Achievement. Other honors include RNA Society/CSHL Press Distinguished Research Mentor award; GSA George Beadle award; ASCB Senior Leadership/ Mentoring Award. She is a national leader in graduate education, including member of the National Academy of Sciences Panel on Bridges to Independence that led to the NIH K99 program, Chair of the AAMC Graduate Research Education Training Group; Chair of the FASEB Consensus Conference on Graduate Education. Yutaka Yamamoto (M.D. Kansai Medical University 1990; Ph.D. with Walter Gehring at Biozentrum - Basel 1995; postdoc with David Glover at Dundee University and University of Cambridge) is a research associate at Brown University. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=65 SRC="FIGDIR/small/488772v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@17fce9aorg.highwire.dtl.DTLVardef@1d4e96forg.highwire.dtl.DTLVardef@129a689org.highwire.dtl.DTLVardef@1748fd8_HPS_FORMAT_FIGEXP M_FIG C_FIG

Disruption of anterior temporal lobe reduces distortions in memory from category knowledge

Tompary, A. — 2022-04-20

Memory retrieval does not provide a perfect recapitulation of past events, but instead an imperfect reconstruction of event-specific details and general knowledge. However, it remains unclear whether this reconstruction relies on mixtures of signals from different memory systems, including one supporting general knowledge. Here, we investigate whether the anterior temporal lobe (ATL) distorts new memories due to prior category knowledge. In this experiment (N=36), participants encoded and retrieved image-location associations. Most images locations were clustered according to their category, but some were in random locations. With this protocol, we previously demonstrated that randomly located images were retrieved closer to their category cluster relative to their encoded locations, suggesting an influence of category knowledge. We combined this procedure with transcranial magnetic stimulation (TMS) delivered to the left ATL before retrieval. We separately examined event-specific details (error) and category knowledge (bias) to identify distinct signals attributable to different memory systems. We found that TMS to ATL attenuated bias in location memory, but only for atypical category members. The magnitude of error was not impacted, suggesting that a memorys fidelity can be decoupled from its distortion by category knowledge. This raises the intriguing possibility that retrieval is jointly supported by separable memory systems.

TAF4b Transcriptional Regulation During Cellular Quiescence of Developing Prospermatogonia

Gura, M. — 2022-04-20

Prospermatogonia (ProSpg) link the embryonic development of male primordial germ cells to the healthy establishment of postnatal spermatogonia and long-term mammalian spermatogenesis. While these spermatogenic precursor cells undergo the characteristic transitions of cycling and quiescence, the transcriptional events underlying these developmental hallmarks remain unknown. Here we investigated the expression and function of TAF4b in the timely development of mouse ProSpg using an integration of gene expression profiling and chromatin mapping. We find that Taf4b mRNA expression is elevated during the transition of M-to-T1 ProSpg and Taf4b-deficient ProSpg are delayed in their entry into quiescence. Gene ontology, protein network analysis, and chromatin mapping demonstrate that TAF4b is both a direct and indirect regulator of cell cycle-related gene expression programs during ProSpg quiescence. By comparing the transcriptome changes in male and female Taf4b-deficient embryonic germ cells, we revealed that TAF4b promotes sex-independent and -dependent gene expression pathways, highlighting its unique and critical role in the fertility of both sexes.

Deceptive learning in histopathology

Shahamatdar, S. — 2022-04-22

Deep learning holds immense potential for histopathology, automating tasks that are simple for expert pathologists, and revealing novel biology for tasks that were previously considered difficult or impossible to solve by eye alone. However, the extent to which the visual strategies learned by deep learning models in histopathological analysis are trustworthy or not has yet to be systematically analyzed. In this work, we address this problem and discover new limits on the histopathological tasks for which deep learning models learn trustworthy versus deceptive solutions. While tasks that have been extensively studied in the field like tumor detection are reliable and trustworthy, recent advances demonstrating the ability to learn molecular profiling from hematoxylin and eosin (H&E) stained slides do not hold up to closer scrutiny. Our analysis framework represents a new approach in understanding the capabilities of deep learning models, which should be incorporated into the computational pathologists toolkit.

The germline factor DDX4 contributes to the chemoresistance of small cell lung cancer cells

Noyes, C. — 2022-04-22

Human cancers often re-express germline factors, yet their mechanistic role in oncogenesis and cancer progression remains unknown. Here we demonstrate that DDX4, a germline factor and RNA helicase conserved in all multicellular organisms, contributes to epithelial mesenchyme transition (EMT)-like features and cisplatin resistance in small cell lung cancer (SCLC) cells. DDX4 depletion in H69AR and SHP77 cell lines decreased motility and resistance to cisplatin, whereas its overexpression increased these features. Proteomic analysis suggests that DDX4 upregulates metabolic protein expression related to DNA repair and immune/inflammatory response, suggesting its fundamental function may be in regulating cellular metabolism. Consistent with these trends in cell lines, DDX4 depletion compromised in vivo tumor development while its overexpression enhanced tumor growth even after cisplatin treatment in nude mice. Although the DDX4 expression level in somatic tumors is generally low compared to that in the germline, the relatively higher DDX4 expression in SCLC patients correlates with decreased survival and shows increased expression of EMT and cisplatin resistance markers. Taken together, we conclude that DDX4 influences the survival of SCLC patients by altering cellular metabolism in response to environmental cues such as drug treatments. This fundamental function of DDX4 as a germline factor might be applicable in other cancer types that express DDX4 and may serve as a key to combat specific tumors that are highly resistant to treatments. HighlightsO_LIDDX4 contributes to cellular motility and drug resistance in SCLC cells. C_LIO_LIDDX4-overexpression globally alters the proteome and suppresses cytokine production. C_LIO_LIDDX4 promotes tumorigenesis and drug resistance in vitro and in vivo. C_LIO_LIDDX4 expression correlates with survival in SCLC patients and with immune/inflammatory response both in cell lines and patient samples. C_LI

Vinculin plays a role in neutrophil stiffening and transit through model capillary segments

Neumann, B. M. — 2022-04-24

Neutrophils are rapidly mobilized from the circulation to sites of inflammation. The mechanisms of neutrophil trafficking in the lung are distinct from those in the periphery, in part because the pulmonary capillaries are the primary site of neutrophil emigration rather than postcapillary venules. Since the diameter of a neutrophil is greater than the width of most pulmonary capillary segments, they must deform to transit through this capillary network, even at homeostasis. Resistance to deformation is primarily due to cortical actin that is rapidly assembled when a neutrophil is exposed to a priming or activation stimulus, resulting in neutrophil stiffening and subsequent sequestration within the pulmonary capillary network. In the current study, we use a microfluidic assay to characterize neutrophil transit through model capillary-like channels. Using techniques from single-particle tracking, we analyzed the cumulative distribution of neutrophil transit times and resolve population-based effects. We found that vinculin, an actin-binding adaptor protein, plays an essential role in neutrophil stiffening in response to formyl-Met-Leu-Phe (fMLP). Vinculin-deficient neutrophils lack the development of a population with slow transit through narrow channels that was observed in both wild-type murine bone marrow neutrophils and HoxB8-conditional progenitor-derived neutrophils. Atomic force microscopy studies provide further evidence that vinculin is required for neutrophil stiffening. Consistent with these findings, we observed that neutrophil sequestration in the lungs of mice is attenuated in the absence of vinculin. Together, our studies indicate that vinculin mediates actin-dependent neutrophil stiffening that leads to their sequestration in capillaries.

Efficacy and Specificity of Melanopsin Reporters for Retinal Ganglion Cells

Maloney, R. T. — 2022-04-27

Intrinsically photosensitive retinal ganglion cells (ipRGCs) are specialized retinal output neurons that mediate behavioral, neuroendocrine, and developmental responses to environmental light. There are diverse molecular strategies for marking ipRGCs, especially in mice, making them among the best characterized retinal ganglion cells. With the development of more sensitive reporters, new subtypes of ipRGCs have emerged. We therefore tested high-sensitivity reporter systems to see whether we could reveal yet more. Substantial confusion remains about which of the available methods, if any, label all and only ipRGCs. Here, we compared many different methods for labeling of ipRGCs, including anti-melanopsin immunofluorescence, Opn4-GFP BAC transgenic mice, and Opn4cre mice crossed with three different Cre-specific reporters (Z/EG; Ai9; and Ai14) or injected with Cre-dependent (DIO) AAV2. We show that Opn4cre mice, when crossed with sensitive Cre-reporter mice, label numerous ganglion cell types that lack intrinsic photosensitivity. Though other methods label ipRGCs specifically, they do not label the entire population of ipRGCs. We conclude that no existing method labels all and only ipRGCs. We assess the appropriateness of each reporter for particular applications and integrate findings across reporters to estimate that the overall abundance of ipRGCs among mouse retinal ganglion cells may approach 11%.

Covalent Docking and Molecular Dynamics Simulations Reveal the Specificity-Shifting Mutations Ala237Arg and Ala237Lys in TEM Beta-Lactamase

Monteiro da Silva, G. — 2022-05-01

The rate of modern drug discovery using experimental screening methods still lags behind the rate at which pathogens mutate, underscoring the need for fast and accurate predictive simulations of protein evolution. Multidrug-resistant bacteria evade our defenses by expressing a series of proteins, the most famous of which is the 29-kilodalton enzyme, TEM {beta}-lactamase. Considering these challenges, we applied a covalent docking heuristic to measure the effects of all possible alanine 237 substitutions in TEM due to this codons importance for catalysis and effects on the binding affinities of commercially-available {beta}-lactam compounds. In addition to the usual mutations that reduce substrate binding due to steric hindrance, we identified two distinctive specificity-shifting TEM mutations, Ala237Arg and Ala237Lys, and their respective modes of action. Notably, we discovered and verified through minimum inhibitory concentration assays that, while these mutations and their bulkier side chains lead to steric clashes that curtail ampicillin binding, these same groups foster salt bridges with the negatively-charged side-chain of the cephalosporin cefixime, widely used in the clinic to treat multi-resistant bacterial infections. To measure the stability of these unexpected interactions, we used molecular dynamics simulations and found the binding modes to be stable despite the application of biasing forces. Finally, we found that both TEM mutants also bind strongly to other drugs containing negatively-charged R-groups, such as carumonam and ceftibuten. As with cefixime, this increased binding affinity stems from a salt bridge between the compounds negative moieties and the positively-charged side chain of the arginine or lysine, suggesting a shared mechanism. In addition to reaffirming the power of using simulations as molecular microscopes, our results can guide the rational design of next-generation {beta}-lactam antibiotics and bring the community closer to retaking the lead against the recurrent threat of multidrugresistant pathogens. Author SummaryResistance to antibiotics is a major public health threat. Microorganisms are able to resist commonly used drugs by evolving and expressing enzymes capable of neutralizing antibiotics. Understanding the relationships between structural elements in these enzymes and their drug-clearing functions can lead to crucial insights for the discovery of next-generation antibiotics. In this study, we have used cutting-edge computational modeling methods to predict the effects of all naturally-occurring variations of an important region of the binding site of TEM {beta}-lactamase, one of the major resistance-granting enzymes in bacteria. In an effort to identify patterns that could be useful for drug discovery, our simulations sought to understand how chemical changes in the tested region can affect resistance against a collection of over 90 widely used antibiotics. Crucially, through our simulations, we have identified a pathway for bacterial resistance against {beta}-lactam antibiotics containing a negatively-charged moiety. We have also elucidated the mechanism behind the gain of resistance, which involves strong interactions between the drugs negative moieties and the positively-charged chemical shifts resulting from the mutation. Finally, we validated our predictions against fitness experiments for two commonly used antibiotics, which qualitatively corroborated our most important findings.

EGR2 promoter antisense RNA coordinates chromatin remodeling and spatial genome organization in Schwann cells.

Moreno, M.-M. — 2022-05-02

The EGR2 promoter antisense RNA (AS-RNA) recruits chromatin remodeling complexes to inhibit EGR2 transcription following peripheral nerve injury. Here we show that the EGR2-AS-RNA modulates chromatin accessibility and interacts with two distinct histone modification complexes. It binds to EZH2 and WDR5 and enables targeting of H3K27me3 and H3K4me3 to promoters of EGR2and C-JUN respectively. Expression of the AS-RNA results in reorganization of the global chromatin landscape and quantitative changes in loop formation and in contact frequency at domain boundaries exhibiting enrichment for AP-1 genes. In addition, the EGR2-AS-RNA induces changes in hierarchical TADs and increases transcription factor occupancy on an inter-TAD loop between a super-enhancer regulatory hub and the promoter of mTOR. Our results show that the EGR2-AS-RNA may serve as regulator of chromatin remodeling and spatial genome organization in Schwann cells.

High-resolution mass spectrometry reveals environmentally relevant uptake, elimination, and metabolic alterations following early embryonic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin in zebrafish

Kossack, M. E. — 2022-05-04

Dioxin and dioxin-like compounds are ubiquitous environmental contaminants that induce toxicity by binding to the aryl hydrocarbon receptor (AHR), a ligand activated transcription factor. The zebrafish model has been used to define the developmental toxicity observed following exposure to exogenous AHR ligands such as the potent agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin, TCDD). While the model has successfully identified cellular targets of TCDD and molecular mechanisms mediating TCDD-induced phenotypes, fundamental information such as the body burden produced by standard exposure paradigms is still unknown. We performed targeted gas chromatography (GC) high-resolution mass spectrometry (HRMS) in tandem with non-targeted liquid chromatography (LC) HRMS to quantify TCDD uptake, model the elimination dynamics of TCDD, and determine how TCDD exposure affects the zebrafish metabolome. We found that 10 ppb, 1 ppb, and 50 ppt waterborne exposures during early embryogenesis produced environmentally relevant body burden of TCDD: 38 {+/-} 4.34, 26.6 {+/-} 1.2, and 8.53 {+/-} 0.341 pg/embryo, respectively, at 24 hours post fertilization. In addition, we discovered that TCDD exposure was associated with the dysregulation of several metabolic pathways that are critical for brain development and function including glutamate metabolism, chondroitin sulfate biosynthesis, and tyrosine metabolism pathways. Together, these data demonstrate that existing exposure paradigms produce environmentally relevant body burdens of TCDD in zebrafish and provide insight into the biochemical pathways impacted by toxicant-induced AHR activation. HIGHLIGHTSO_LIHistorical TCDD exposure paradigms produce environmentally relevant body burdens in zebrafish embryos. C_LIO_LITCDD elimination for high doses can be modeled using an exponential regression. C_LIO_LIExposure to TCDD alters metabolic pathways that are essential for brain development and function. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/490602v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@17ba806org.highwire.dtl.DTLVardef@1543d58org.highwire.dtl.DTLVardef@1878204org.highwire.dtl.DTLVardef@168f21d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Molecular insights into the effect of alkanediols on FUS liquid-liquid phase separation

Perdikari, T. M. — 2022-05-06

1,6-hexanediol disrupts many phase-separated condensates in cells and in test tubes. In this study, we use a combination of microscopy, nuclear magnetic resonance (NMR) spectroscopy, molecular simulation, and biochemical assays to probe how alkanediols suppress phase separation and why certain isomers are more effective. Alkanediols of different lengths and configurations are all capable of disrupting phase separation of the RNA-binding protein Fused in Sarcoma (FUS), though potency varies depending on both geometry and hydrophobicity, which we measure directly. Alkanediols induce a shared pattern of changes to the protein chemical environment though to differing extents. Consistent with the view that alkanediols disrupt phase separation driven by hydrophobic groups, they decrease the thermal stability of a model globular protein. Conversely, 1,6-hexanediol does not disrupt charge-mediated phase separation, such as FUS RGG-RNA and poly-lysine/poly-aspartic acid condensates. All-atom simulations show that hydroxyl groups in alkanediols mediate interaction with protein backbone and polar amino acid side chains, while the aliphatic chain allows contact with hydrophobic and aromatic residues, providing a molecular picture of how amphiphilic interactions disrupt FUS phase separation.

Defining the HIV Capsid Binding Site of Nucleoporin 153

Li, S. — 2022-05-07

The interaction between the HIV-1 capsid (CA) and human nucleoporin 153 (NUP153) is vital for delivering the HIV-1 preintegration complex into the nucleus via the nuclear pore complex. The interaction with CA requires a phenylalanine/glycine-containing motif in the C-terminus of NUP153. This study used molecular modeling and biochemical assays to determine the amino acids of NUP153 that are essential for its interactions with CA. Molecular dynamics, FoldX, and PyRosetta simulations delineated the minimal CA binding motif of NUP153 based on the known structure of NUP153 bound to the HIV-1 CA hexamer. Computational predictions were experimentally validated by testing the interaction of NUP153 with CA using an in vitro binding assay and a cell-based TRIM-NUP153C restriction assay. This multidisciplinary approach identified eight amino acids from P1411 to G1418 that stably engage with CA, with significant correlations between molecular models and empirical experiments. Specifically, P1411, V1414, F1415, T1416, F1417, and G1418 were confirmed as critical amino acids required to interact NUP153 with CA. IMPORTANCEHuman immunodeficiency virus (HIV) can infect non-dividing cells by interacting with host nuclear pores. The host nuclear pore protein NUP153 directly interacts with the HIV capsid to promote viral nuclear entry. This study used a multidisciplinary approach combining computational and experimental techniques to map the essential amino acids of NUP153 required for HIV capsid interaction. This approach revealed that the HIV capsid interacts specifically with only six amino acids of NUP153, suggesting other FG-containing motifs could also interact with the capsid. Based on molecular modeling, naturally occurring polymorphisms in human and non-human primates would be predicted to prevent NUP153 interaction with capsid, potentially protecting from HIV infection.

The Role of Conjunctive Representations in Prioritizing and Selecting Planned Actions

Kikumoto, A. — 2022-05-10

For flexible goal-directed behavior, prioritizing and selecting a specific action among multiple candidates is often important. Working memory has long been assumed to play a role in prioritization and planning, while bridging cross-temporal contingencies during action selection. However, studies of working memory have mostly focused on memory for single components of an action plan, such as a rule or a stimulus, rather than management of all of these elements during planning. Therefore, it is not known how post-encoding prioritization and selection operate on the entire profile of representations for prospective actions. Here, we assessed how such control processes unfold over action representations, highlighting the role of conjunctive representations that nonlinearly integrate task-relevant features during maintenance and prioritization of action plans. For each trial, participants prepared two independent rule-based actions simultaneously, then they were retro-cued to select one as their response. Prior to the start of the trial, one rule-based action was randomly assigned to be high priority by cueing that it was more likely to be tested. We found that both full action plans were maintained as conjunctive representations during action preparation, regardless of priority. However, during output selection, the conjunctive representation of the high priority action plan was more enhanced and readily selected as an output. Further, the strength of conjunctive representation was related to behavioral interference when the low priority action was tested. Thus, multiple integrated representations were maintained for upcoming actions and served as the target of post-encoding attentional selection mechanisms to prioritize and select an action from those in working memory.

Mouse primary T cell phosphotyrosine proteomics enabled by BOOST

Chua, X. Y. — 2022-05-13

The Broad Spectrum Optimization of Selective Triggering (BOOST) approach was recently developed to increase the quantitative depth of the tyrosine phosphoproteome by mass spectrometry-based proteomics. While BOOST has been demonstrated in the Jurkat T cell line, it has not been demonstrated in scarce mice primary T cells. Here, we show the first phosphotyrosine proteomics experiment performed in mice primary T cells using BOOST. We identify and precisely quantify more than 2,000 unique pTyr sites from more than 3,000 unique pTyr peptide PSMs using only 1 mg of protein from T cell receptor-stimulated primary T cells from mice. We further reveal the importance of the phase-constrained spectrum deconvolution method ({Phi}SDM) parameter on Orbitrap instruments that, when disabled, enhances quantitation depth, accuracy, and precision in low-abundance samples. Using samples with contrived ratios, we find that disabling {Phi}SDM allows for up to a two-fold increase in the number of statistically significant intensity ratios detected while enabling {Phi}SDM degrades quantitation, especially in low-abundance samples. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/491817v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@4a2742org.highwire.dtl.DTLVardef@57645borg.highwire.dtl.DTLVardef@17dbfacorg.highwire.dtl.DTLVardef@b2df00_HPS_FORMAT_FIGEXP M_FIG C_FIG

Estimation of Periodic Signals with Applications to Deep Brain Stimulation

Chen, P. — 2022-05-24

Deep brain stimulation (DBS) therapies have shown clinical success in the treatment of a number of neurological illnesses, including obsessive-compulsive disorder, depression, and Parkinsons disease. An emerging strategy for increasing the efficacy of DBS therapies is to develop closed-loop, adaptive DBS systems that can sense biomarkers associated with particular symptoms and in response, adjust DBS parameters in realtime. The development of such systems requires extensive analysis of the underlying neural signals while DBS is on, so that candidate biomarkers can be identified and the effects of varying the DBS parameters can be better understood. However, DBS creates high amplitude, high frequency stimulation artifacts that prevent the underlying neural signals and thus the biological mechanisms underlying DBS from being analyzed. Additionally, DBS devices often require low sampling rates, which alias the artifact frequency, and rely on wireless data transmission methods that can create signal recordings with missing data of unknown length. Thus, traditional artifact removal methods cannot be applied to this setting. We present a novel periodic artifact removal algorithm for DBS applications that can accurately remove stimulation artifacts in the presence of missing data and in some cases where the stimulation frequency exceeds the Nyquist frequency. The numerical examples suggest that, if implemented on dedicated hardware, this algorithm has the potential to be used in embedded closed-loop DBS therapies to remove DBS stimulation artifacts and hence, to aid in the discovery of candidate biomarkers in real-time. Code for our proposed algorithm is publicly available on Github.

coiaf: directly estimating complexity of infection with allele frequencies

Paschalidis, A. — 2022-05-28

In malaria, individuals are often infected with different parasite strains; the complexity of infection (COI) is defined as the number of genetically distinct parasite strains in an individual. Changes in the mean COI in a population have been shown to be informative of changes in transmission intensity with a number of probabilistic likelihood and Bayesian models now developed to estimate the COI. However, rapid, direct measures based on heterozygosity or FwS do not properly represent the COI. In this work, we present two new methods that use easily calculated measures to directly estimate the COI from allele frequency data. Using a simulation framework, we show that our methods are computationally efficient and comparably accurate to current methods in the literature. Through a sensitivity analysis, we characterize how the bias and accuracy of our two methods are impacted by the distribution of parasite densities and the assumed sequencing depth and number of sampled loci. We further estimate the COI globally from Plasmodium falciparum sequencing data using our developed methods and compare the results against the literature. We show significant differences in estimated COI globally between continents and a weak relationship between malaria prevalence and COI. Author summaryComputational models, used in conjunction with rapidly advancing sequencing technologies, are increasingly being used to help inform surveillance efforts and understand the epidemiological dynamics of malaria. One such important metric, the complexity of infection (COI), indirectly quantifies the level of transmission. Existing "gold-standard" COI measures rely on complex probabilistic likelihood and Bayesian models. As an alternative, we have developed the statistics and software package coiaf, which features two rapid, direct measures to estimate of the number of genetically distinct parasite strains in an individual (the COI). Our methods were evaluated using simulated data and subsequently compared to current "state-of-the-art" methods, yielding comparable results. Lastly, we examined the distribution of the COI in several locations across the world, identifying significant differences in COI between continents. coiaf, therefore, provides a new, promising framework for rapidly characterizing polyclonal infections.

Suction feeding of West African lungfish (Protopterus annectens): An XROMM analysis of jaw mechanics, cranial kinesis, and hyoid mobility.

Gartner, S. M. — 2022-05-30

Suction feeding in fishes is characterized by rapid cranial movements, but extant lungfishes (Sarcopterygii: Dipnoi) exhibit a reduced number and mobility of cranial bones relative to actinopterygian fishes. Despite fusion of cranial elements, lungfishes are proficient at suction feeding, though the impacts of novel cranial morphology and reduced cranial kinesis on feeding remain poorly understood. We used X-ray Reconstruction of Moving Morphology (XROMM) to study the kinematics of seven mobile skeletal elements (neurocranium, upper jaw, lower jaw, tongue, ceratohyal, clavicle, and cranial rib) and two muscles (costoclavicular portion of the hypaxialis and rectus cervicis) during the feeding strikes of West African lungfish (Protopterus annectens). We found that feeding by P. annectens on non-evasive prey is relatively slow, with a mean time to peak gape of 273 ms. Lower jaw depression and clavicular rotation were hingelike, with one degree of freedom, but the ceratohyals rotated in a complex motion involving depression and long-axis rotation. We quantified the relative contributions to oral cavity volume change (RCVC) and found that oral cavity expansion is created primarily by ceratohyal and clavicle motion. P. annectens suction feeds relatively slowly but successfully through muscle shortening of hypaxial and rectus cervicis muscles contributing to hyoid mobility. Summary StatementThree-dimensional hyoid movements and clavicle retraction generate suction during the relatively slow, but successful, feeding strikes of the West African lungfish (Protopterus annectens).

Language exposure and brain myelination in early development

Fibla, L. — 2022-06-02

The language environment to which children are exposed has an impact on later language and cognitive abilities as well as on brain development; however, it is unclear how early such impacts emerge. This study investigates the effects of childrens early language environment and socioeconomic status (SES) on brain structure in infancy at both 6 and 30 months of age. We used magnetic resonance imaging (MRI) to quantify concentrations of myelin in specific fiber tracts in the brain. Our central question was whether Language ENvironment Analysis (LENA) measures from in-home recording devices and SES measures of maternal education and family income predicted myelin concentrations over development. Results show relationships between amount of in-home adult input and myelination in the white matter tracts most associated with language. Right hemisphere regions also show an association with SES, with older children from a higher SES background who were exposed to more adult input showing greater concentrations of myelin in language-related areas. We discuss these results in relation with the current literature and implications for future language research and intervention. Significance StatementThis is the first study to look at how brain myelination is impacted by language input and socioeconomic status early in development. We find robust relationships of both factors in language-related brain areas at 30 months of age.

Non-invasive auditory brainstem responses to FM sweeps in awake big brown bats

Simmons, A. M. — 2022-06-04

We introduce two EEG techniques, one based on conventional monopolar electrodes and one based on a novel tripolar electrode, to record for the first time auditory brainstem responses (ABRs) from the scalp of unanesthetized, unrestrained big brown bats. Stimuli were frequency-modulated (FM) sweeps varying in sweep direction, sweep duration, and harmonic structure. As expected from previous invasive ABR recordings, upward-sweeping FM signals evoked larger amplitude responses (peak-to-trough amplitude in the latency range of 3-5 ms post-stimulus onset) than downward-sweeping FM signals. Scalp-recorded responses displayed amplitudelatency trading effects as expected from invasive recordings. These two findings validate the reliability of our noninvasive recording techniques. The feasibility of recording noninvasively in unanesthetized, unrestrained bats will energize future research uncovering electrophysological signatures of perceptual and cognitive processing of biosonar signals in these animals, and allows for better comparison with ABR data from echolocating cetaceans, where invasive experiments are heavily restricted. Because experiments can be repeated in the same animal over time without confounds of stress or anesthesia, our technique requires fewer captures of wild bats, thus helping to preserve natural populations and addressing the goal of reducing animal numbers used for research purposes.

Conserved reduction of m6A marks during aging and neurodegeneration is linked to altered translation of synaptic transcripts

Castro-Hernandez, R. — 2022-06-10

N6-methyladenosine (m6A) plays diverse roles in the regulation of mRNA metabolism. In the mammalian brain it has been linked to developmental processes and memory function. However, the precise role of m6A in the context synaptic plasticity and especially during impaired cognition are not fully understood. Here, we describe the mouse and human brain m6A epi-transcriptome in a tissue-specific manner. We furthermore show that m6A levels undergo a massive decrease across mouse brain regions as a consequence of aging. In addition, Alzheimer's disease in humans correlates with decreased N6-methylation in a similar population of transcripts that are linked to synaptic function and localized to synapses, such as the calcium/calmodulin-dependent kinase II (CaMKII). We furthermore show that reduced m6A levels impair synaptic protein-synthesis of CAMKII. Our results suggest that m6A-RNA-methylation is an important mechanism to control synaptic protein synthesis which is affected early in cognitive diseases. Significance statementThe addition of N6-methyladenosine (m6A) to RNA plays a role in various cellular processes and its de-regulation has been linked to several devastating diseases. The precise role of m6A RNA-methylation in the adult brain is, however, not well understood. In our study, we describe the genome-wide m6A epi-transcriptome in the healthy and diseased brains of mice and humans. Our data demonstrate that a substantial amount of m6A transcripts are conserved. These transcripts are linked to the regulation of synaptic processes and are localized to synapses. In the diseases brain we detect RNA hypomethylation across multiple transcripts in all investigated brain regions and across species. At the mechanistic level we find that reduced m6A levels specifically impairs synaptic protein-synthesis.

Denoising Approach Affects Diagnostic Differences in Brain Connectivity across the Alzheimer's Disease Continuum

Blujus, J. K. — 2022-06-21

Graph theory provides a promising technique to investigate Alzheimers disease (AD)-related alterations in brain connectivity. However, discrepancies exist in the reported disruptions that occur to network topology across the AD continuum, which may be attributed to differences in the denoising approach used in fMRI processing to remove the effect of non-neuronal sources from signal. The current study aimed to determine if diagnostic differences in graph metrics were dependent on nuisance regression strategy. Sixty cognitively normal (CN), 60 MCI, and 40 AD matched for age, sex, and motion, were selected from the ADNI database for analysis. Resting state images were preprocessed using AFNI (v21.2.04) and 16 nuisance regression approaches were employed, which included the unique combination of four nuisance regressors (derivatives of the realignment parameters, motion censoring [euclidean norm > 0.3mm], outlier censoring [outlier fraction > .10], bandpass filtering [0.01 - 0.1 Hz]). Graph metrics representing network segregation (clustering coefficient, local efficiency, modularity), network integration (largest connected component, path length, local efficiency), and small-worldness (clustering coefficient/path length) were calculated. The results showed a significant interaction between diagnosis and nuisance approach on path length, such that diagnostic differences were only evident when motion derivatives and censoring of both motion and outlier volumes were applied. Further, regardless of the denoising approach, AD patients exhibited less segregated networks and lower small-worldness than CN and MCI. Finally, independent of diagnosis, denoising strategy significantly affected the magnitude of nearly all metrics (except local efficiency), such that models including bandpass filtering had higher graph metrics than those without. These findings suggest the relative robustness of network segregation and small-worldness properties to denoising strategy. However, caution should be taken when interpreting path length findings across studies, as subtle variations in regression approach may account for discrepancies. Continued efforts should be taken towards harmonizing preprocessing pipelines across studies to aid replication efforts and build consensus towards understanding the mechanisms underlying pathological aging.

Functional Redundancy of the Posterior Hippocampi, but not Anterior Hippocampi or Left Frontal Cortex, is Disrupted in Pathological Brain Aging

Blujus, J. — 2022-06-20

As prevalence rates of Alzheimers disease (AD), the leading cause of dementia, are projected to more than double by 2050, emphasis has been placed on early intervention strategies that target resilience mechanisms to delay or prevent the onset of clinical symptoms. Several neural mechanisms underlying brain resilience to AD have been proposed, including redundant neural connections between the posterior hippocampi (HC) and all other brain regions, and global functional connectivity of the left frontal cortex (LFC). It remains unknown, however, if regional redundancy of the HC and LFC underscores neural resilience in the presence of AD pathologies. From the ADNI database, 363 cognitively normal older adults (CN) (N = 220; 36% A{beta}+) and patients with Mild Cognitive Impairment (MCI) (N = 143; 51% A{beta}+) were utilized. Regional redundancy was calculated from resting state fMRI data using a graph theoretical approach by summing the direct and indirect paths (path lengths=1-4) between each ROI and its 262 functional connections. The results showed that A{beta}-status significantly disrupted posterior HC, but not anterior HC or LFC, redundancy. A{beta}- groups showed higher redundancy of the bilateral posterior HC than A{beta}+. In regard to redundancy-cognition relationships, higher posterior HC redundancy was related to better episodic memory performance, an effect which was primarily driven by the A{beta}- group. Despite the positive relationship between posterior HC redundancy and cognition, we did not find compelling evidence that redundancy of the posterior HC serves in a resilience manner, as posterior HC redundancy did not moderate the potentially deleterious relationship between A{beta} deposition and cognition. No relationships were found between anterior HC or LFC redundancy and cognitive performance. Together, these findings suggest that redundancy of the LFC does not underpin its role in resilience and that posterior HC redundancy may capture disruptions to network connectivity that occur as a result of A{beta} deposition.

Demographic history and genetic structure in pre-Hispanic Central Mexico

Villa-Islas, V. — 2022-06-20

Aridoamerica and Mesoamerica are two distinct cultural areas that hosted numerous pre-Hispanic civilizations between 2,500 BCE and 1,521 CE. The division between these regions shifted southward due to severe droughts ca. 1,100 years ago, allegedly driving demographic changes and population replacement in some sites in central Mexico. Here, we present shotgun genome-wide data from 12 individuals and 26 mitochondrial genomes from eight pre-Hispanic archaeological sites across Mexico, including two at the shifting border of Aridoamerica and Mesoamerica. We find population continuity spanning the climate change episode and a broad preservation of the genetic structure across present-day Mexico for the last 2,300 years. Lastly, we identify a contribution to pre-Hispanic populations of northern and central Mexico from an ancient unsampled ghost population.

Beyond Drift Diffusion Models: Fitting a broad class of decision and RL models with HDDM

Fengler, A. — 2022-06-20

Computational modeling has become a central aspect of research in the cognitive neurosciences. As the field matures, it is increasingly important to move beyond standard models to quantitatively assess models with richer dynamics that may better reflect underlying cognitive and neural processes. For example, sequential sampling models (SSMs) are a general class of models of decision making intended to capture processes jointly giving rise to reaction time distributions and choice data in n-alternative choice paradigms. A number of model variations are of theoretical interest, but empirical data analysis has historically been tied to a small subset for which likelihood functions are analytically tractable. Advances in methods designed for likelihood-free inference have recently made it computationally feasible to consider a much larger spectrum of sequential sampling models. In addition, recent work has motivated the combination of SSMs with reinforcement learning (RL) models, which had historically been considered in separate literatures. Here we provide a significant addition to the widely used HDDM Python toolbox and include a tutorial for how users can easily fit and assess a (user extensible) wide variety of SSMs, and how they can be combined with RL models. The extension comes batteries included, including model visualization tools, posterior predictive checks, and ability to link trial-wise neural signals with model parameters via hierarchical Bayesian regression.

ct2vl: Converting Ct Values to Viral Loads for SARS-CoV-2 RT-qPCR Test Results

Hill, E. — 2022-06-21

RT-qPCR is the de facto reference method for detecting the presence of SARS-CoV-2 genomic material in infected individuals (1). Although RT-qPCR is inherently quantitative and despite SARS-CoV-2 viral loads varying by 10 orders of magnitude and therefore being potentially highly clinically informative, in practice SARS-CoV-2 RT-qPCR results are usually reported qualitatively as simply positive or negative. This is both because of the mathematical complexity of converting from Ct values to viral loads and because the same Ct value can correspond to orders-of-magnitude differences in viral load depending on the testing platform (2, 3, 4). To address this problem, here we present ct2vl, a Python package designed to help individual clinical laboratories, investigators, and test developers convert from Ct values to viral loads on their own platforms, using only the data generated during validation of those platforms. It allows any user to convert Ct values to viral loads and is readily applicable to other RT-qPCR tests. ct2vl is open source, has 100% code coverage, and is freely available via the Python Package Index (PyPI). IMPORTANCEUp to now, COVID-19 test results have been reported as positive vs. negative, even though "positive" can mean anywhere from 1 copy of SARS-CoV-2 virus per milliliter of transport media to over 1 billion copies/mL, with attendant clinical consequences. Democratizing access to this quantitative data is the first step toward its eventual incorporation into test development, the research literature, and clinical care.

Reprogramming brain immunosurveillance with engineered cytokines

Tabet, A. — 2022-06-24

Immune surveillance of the brain is regulated by resident non-neuronal cells and the blood-brain barrier.1 Dys-regulation of immunosurveillance is a hallmark feature of several diseases2-5 including brain tumors6 that interact with and rely heavily on immune cells,7 suggesting that disrupting the neuroimmunology of tumors could slow their progression. Yet few tools are available to control brain immunology in vivo with local precision, and fewer yet are used for therapeutic intervention. 2 Here, we propose engineered cytokines as a neuroimmune-modulation platform. We demonstrate that the residence time of cytokines in the brain can be tuned by binding them to the extracellular matrix or synthetic scaffolds. We then show that the aluminum hydroxide adjuvant (alum) is retained in the brain >2 weeks. Tethering of inflammatory cytokines such as interleukins (IL) 2 and 12 to alum yields extended neuroinflammation and brain immunosurveillance after intracranial administration, while avoiding systemic toxicity. In mouse models of both immunologically hot and cold brain tumors, the intracranial deposition of alum-tethered cytokines causes significant delay in tumor progression. RNA profiling reveals that engineered cytokines engage both innate and adaptive immunity in the brain. These findings suggest that engineered cytokines can reprogram brain immunosurveillance, informing the development of future therapies for neuroimmune diseases.

CHI3L1 Enhances Melanoma Lung Metastasis via Regulation of T cell Co-stimulators and CTLA-4/B7 Axis

Ma, B. — 2022-06-26

ICOS/ICOSL and CD28/B7-1/B7-2 are T cell co-stimulators and CTLA-4 is an immune checkpoint inhibitor that play critical roles in the pathogenesis of neoplasia. Chitinase 3-like-1 (CHI3L1) is induced in many cancers where it portends a poor prognosis and contributes to tumor metastasis. Here we demonstrate that CHI3L1 inhibits the expression of ICOS, ICOSL and CD28 while stimulating CTLA-4 and the B7 moieties in melanoma lung metastasis. We also demonstrate that RIG-like helicase innate immune activation augments T cell co-stimulation, inhibits CTLA-4 and suppresses pulmonary metastasis. At least additive antitumor responses were seen in melanoma lung metastasis treated with anti-CTLA-4 and anti-CHI3L1 antibodies in combination. Synergistic cytotoxic T cell-induced tumor cell death and the heightened induction of the tumor suppressor PTEN were seen in co-cultures of T and tumor cells treated with bispecific antibodies that target both CHI3L1 and CTLA-4. Thus, CHI3L1 contributes to pulmonary metastasis by inhibiting T cell co-stimulation and stimulating CTLA-4. The simultaneous targeting of CHI3L1 and the CTLA-4 axis with individual and, more powerfully with bispecific antibodies, represent promising therapeutic strategies for pulmonary metastasis.

NEURONS IN THE POSTRHINAL CORTEX ENCODE NONSPATIAL CONTEXT DURING VISUAL BICONDITIONAL DISCRIMINATION

Heimer-McGinn, V. R. — 2022-06-26

Spatial context, or the physical surroundings that form the background of an experience, is an essential component of episodic memory. The rodent postrhinal cortex and its primate homolog, the parahippocampal cortex, are thought to preferentially process visuospatial information to represent the spatial features of contexts and scenes. In this study, we addressed open questions about postrhinal function and about how context modulates behavior and cognition. The first question was whether the postrhinal cortex also represents nonspatial contexts. The second question was how representations of context might interact with other cues in the environment. We recorded postrhinal neurons as rats performed a visual nonspatial biconditional discrimination task in which the pattern on the floor determined which object in a pair was correct. Critically, this task design allowed dissociation of location from non-spatial context. We found that postrhinal ensembles and neurons signaled changes in non-spatial context and coded for conjunctions of non-spatial context and objects. Importantly, postrhinal neurons coded for conjunctions of context and objects more often than they coded for conjunctions of location and object. The pattern of findings suggests that postrhinal representations of context may behave like occasion setters by modulating the meaning of other cues in the environment.

Expression Microdissection for use in qPCR based analysis of miRNA in a single cell type

Jenike, A. E. — 2022-06-28

Cell-specific microRNA (miRNA) expression estimates are important in characterizing the localization of miRNA signaling within tissues. Much of this data is obtained from cultured cells, a process known to significantly alter miRNA expression levels. Thus, our knowledge of in vivo cell miRNA expression estimates is poor. We previously demonstrated expression microdissection-miRNA-sequencing (xMD-miRNA-seq) as a means to acquire in vivo estimates, directly from formalin fixed tissues, albeit with limited yield. Here we optimized each step of the xMD process including tissue retrieval, tissue transfer, film preparation, and RNA isolation to increase RNA yields and ultimately show strong enrichment for in vivo miRNA expression by qPCR array. These method improvements, including the development of a non-crosslinked ethylene vinyl acetate (EVA) membrane, resulted in a 23-45 fold increase in miRNA yield, depending on cell type. By qPCR, miR-200a was increased 14-fold in xMD-derived small intestine epithelial cells, with a concurrent 336-fold reduction in miR-143, relative to the matched non-dissected duodenal tissue. xMD is now an optimized method to obtain robust in vivo miRNA expression estimates from cells.

Synapses, predictions, and prediction errors: a neocortical computational study of MDD using the temporal memory algorithm of HTM.

Sherif, M. — 2022-07-03

BackgroundSynapses and spines are central in major depressive disorder (MDD) pathophysiology, recently highlighted by ketamines and psilocybins rapid antidepressant effects. The Bayesian brain and interoception perspectives formalize MDD as being "stuck" in affective states constantly predicting negative energy balance. We examined how synaptic atrophy relates to the predictive function of the neocortex and thus to symptoms, using temporal memory (TM), an unsupervised machine-learning algorithm. TM represents a single neocortical layer, learns in real-time using local Hebbian-learning rules, and extracts and predicts temporal sequences. MethodsWe trained a TM model on random sequences of upper-case alphabetical letters, representing sequences of affective states. To model depression, we progressively destroyed synapses in the TM model and examined how that affected the predictive capacity of the network. ResultsDestroying 50% of the synapses slightly reduced the number of predictions, followed by a marked drop with further destruction. However, reducing the synapses by 25% dropped the confidence in the predictions distinctly. So even though the network was making accurate predictions, the network was no longer confident about these predictions. ConclusionsThese findings explain how interoceptive cortices could be stuck in limited affective states with high prediction error. Growth of new synapses, e.g., with ketamine and psilocybin, would allow representing more futuristic predictions with higher confidence. To our knowledge, this is the first study to use the TM model to connect changes happening at synaptic levels to the Bayesian formulation of psychiatric symptomatology, making it possible to understand treatment mechanisms and possibly, develop new treatments. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

Identification and Mechanistic Basis of non-ACE2 Blocking Neutralizing Antibodies from COVID-19 Patients with Deep RNA Sequencing and Molecular Dynamics Simulations

Fredericks, A. M. — 2022-06-30

Variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continue to cause disease and impair the effectiveness of treatments. The therapeutic potential of convergent neutralizing antibodies (NAbs) from fully recovered patients has been explored in several early stages of novel drugs. Here, we identified initially elicited NAbs (Ig Heavy, Ig lambda, Ig kappa) in response to COVID-19 infection in patients admitted to the intensive care unit at a single center with deep RNA sequencing (>100 million reads) of peripheral blood as a diagnostic tool for predicting the severity of the disease and as a means to pinpoint specific compensatory NAb treatments. Clinical data were prospectively collected at multiple time points during ICU admission, and amino acid sequences for the NAb CDR3 segments were identified. Patients who survived severe COVID-19 had significantly more of a Class 3 antibody (C135) to SARS-CoV-2 compared to non-survivors (16,315 reads vs 1,412 reads, p=0.02). In addition to highlighting the utility of RNA sequencing in revealing unique NAb profiles in COVID-19 patients with different outcomes, we provided a physical basis for our findings via atomistic modeling combined with molecular dynamics simulations. We established the interactions of the Class 3 NAb C135 with the SARS-CoV-2 spike protein, proposing a mechanistic basis for inhibition via multiple conformations that can effectively prevent ACE2 from binding to the spike protein, despite C135 not directly blocking the ACE2 binding motif. Overall, we demonstrate that deep RNA sequencing combined with structural modeling offers the new potential to identify and understand novel therapeutic(s) NAbs in individuals lacking certain immune responses due to their poor endogenous production. Our results suggest a possible window of opportunity for administration of such NAbs when their full sequence becomes available. A method involving rapid deep RNA sequencing of patients infected with SARS-CoV-2 or its variants at the earliest infection time could help to develop personalized treatments using the identified specific NAbs.

Partitioning Marginal Epistasis Distinguishes Nonlinear Effects from Polygenicity and Other Biases in GWA Summary Statistics

Darnell, G. — 2022-07-22

LD score regression (LDSC) is a method to estimate narrow-sense heritability from genome-wide association study (GWAS) summary statistics alone, making it a fast and popular approach. In this work, we present interaction-LD score (i-LDSC) regression: an extension of the original LDSC framework that accounts for interactions between genetic variants. By studying a wide range of generative models in simulations, and by re-analyzing 25 well-studied quantitative phenotypes from 349,468 individuals in the UK Biobank and up to 159,095 individuals in BioBank Japan, we show that the inclusion of a cis-interaction score (i.e., interactions between a focal variant and proximal variants) recovers genetic variance that is not captured by LDSC. For each of the 25 traits analyzed in the UK Biobank and BioBank Japan, i-LDSC detects additional variation contributed by genetic interactions. The i-LDSC software and its application to these biobanks represent a step towards resolving further genetic contributions of sources of non-additive genetic effects to complex trait variation.

Increased homozygosity due to endogamy results in fitness consequences in a human population

Swinford, N. — 2022-07-26

Recessive alleles have been shown to directly affect both human Mendelian disease phenotypes and complex traits like height. Pedigree studies also suggest that consanguinity results in increased childhood mortality and adverse health phenotypes, presumably through penetrance of recessive mutations. Here, we test whether the accumulation of homozygous, recessive alleles decreases reproductive success in a human population. We address this question among the Namibian Himba, an endogamous agro-pastoralist population, who until very recently practiced natural fertility. Using a sample of 681 individuals, we show that Himba exhibit elevated levels of "inbreeding", calculated as the fraction of the genome in runs of homozygosity (FROH). Many individuals contain multiple long segments of ROH in their genomes, indicating that their parents had high kinship coefficients. However, we did not find evidence that this is explained by first-cousin consanguinity, despite a reported social preference for cross-cousin marriages. Rather, we show that elevated haplotype sharing in the Himba is due to a bottleneck, likely in the past 60 generations. We test whether increased recessive mutation load results in observed fitness consequences by assessing the effects of FROH on completed fertility in a cohort of post-reproductive women (n=69). We find that higher FROH is significantly associated with lower fertility among women who have had at least one child (p<0.006). Our data suggest a multi-locus genetic effect on fitness driven by the expression of deleterious recessive alleles, especially those in long ROH. However, these effects are not the result of consanguinity but rather elevated background identity by descent.

Revising transcriptome assemblies with phylogenetic information in Agalma1.0

Guang, A. — 2017-10-12

MotivationOne of the most common transcriptome assembly errors is to mistake different transcripts of the same gene as transcripts from multiple closely related genes. It is difficult to identify these errors during assembly, but in a phylogenetic analysis these errors can be diagnosed from gene trees containing clades of tips from the same species with improbably short branch lengths.nnResultstreeinform is a module implemented in Agalma1.0 that uses phylogenetic analyses across species to refine transcriptome assemblies. It identifies transcripts of the same gene that were incorrectly assigned to multiple genes and reassign them as transcripts of the same gene.nnAvailability and Implementationtreeinform is implemented in Agalma1.0, available at https://bitbucket.org/caseywdunn/agalma.nnContactaugust_guang@brown.edunnSupplementary informationSupplementary information is available at bioRxiv.

A control theoretic model of adaptive behavior in dynamic environments

Ritz, H. — 2017-10-16

To behave adaptively in environments that are noisy and non-stationary, humans and other animals must monitor feedback from their environment and adjust their predictions and actions accordingly. An under-studied approach for modeling these adaptive processes comes from the engineering field of control theory, which provides general principles for regulating dynamical systems, often without requiring a generative model. The proportional-integral-derivative (PID) controller is one of the most popular models of industrial process control. The proportional term is analogous to the "delta rule" in psychology, adjusting estimates in proportion to each successive error in prediction. The integral and derivative terms augment this update to simultaneously improve accuracy and stability. Here, we tested whether the PID algorithm can describe how people sequentially adjust their predictions in response to new information. Across three experiments, we found that the PID controller was an effective model of participants decisions in noisy, changing environments. In Experiment 1, we re-analyzed a change-point detection experiment, and showed that participants behavior incorporated elements of PID updating. In Experiments 2-3 we developed a task with gradual transitions that we optimized to detect PID-like adjustments. In both experiments, the PID model offered better descriptions of behavioral adjustments than both the classical delta-rule model and its more sophisticated variant, the Kalman filter. We further examined how participants weighted different PID terms in response to salient environmental events, finding that these control terms were modulated by reward, surprise, and outcome entropy. These experiments provide preliminary evidence that adaptive behavior in dynamic environments resembles PID control.

Clearing the Fungal FoG: Perseverance, a property distinct from resistance, is associated with clinical persistence

Rosenberg, A. — 2017-10-20

Drug susceptibility, defined by the minimal inhibitory concentration (MIC), often does not predict whether fungal infections will respond to therapy in the clinic. Tolerance at supra-MIC antifungal drug concentrations is rarely quantified and current clinical recommendations suggest it be ignored. Here, we measured and characterized drug-response variables that could influence the outcomes of fungal infections and be generalizable across major clades of Candida albicans, one of the most frequently isolated human fungal pathogens. We quantified antifungal tolerance as the fraction of growth (FoG) above the MIC and found that it is clearly distinct from susceptibility/resistance measured as MIC. Instead, tolerance is due to the slow growth of subpopulations of cells that overcome drug stress more efficiently than the rest of the population, and correlates inversely with the accumulation of intracellular drug. Importantly, many adjuvant drugs used together with fluconazole, a fungistatic drug, reduce tolerance without affecting resistance. These include inhibitors of major stress response hubs such as Hsp90, calcineurin, PKC1 and TOR. Accordingly, in an invertebrate infection model, adjuvant combination therapy was significantly more effective than fluconazole alone in treating highly tolerant isolates and did not improve the treatment of isolates with low tolerance levels. Furthermore, isolates recovered from immunocompetent patients with persistent candidemia displayed significantly higher tolerance than isolates that were readily cleared by fluconazole. Thus, tolerance correlates with the response to fluconazole therapy in patients and may help predict whether infections will respond to fluconazole alone. Similarly, measuring tolerance may provide a useful clinical parameter for choosing appropriate therapeutic strategies to overcome persistent clinical candidemia.

Left-Lateralized Contributions of Saccades to Cortical Activity during a One-Back Word Recognition Task

Chang, Y.-C. C. — 2017-12-23

Saccadic eye movements are an inherent component of natural reading, yet their contribution to information processing at subsequent fixation remains elusive. Here we use anatomically-constrained magnetoencephalography (MEG) to examine cortical activity following saccades as healthy human subjects engaged in a one-back word recognition task. This activity was compared with activity following external visual stimulation that mimicked saccades. A combination of procedures were employed to eliminate saccadic ocular artifacts from the MEG signal. Both saccades and saccade-like external visual stimulation produced early-latency responses beginning ~70 ms after onset in occipital cortex and spreading through the ventral and dorsal visual streams to temporal, parietal and frontal cortices. Robust differential activity following the onset of saccades versus similar external visual stimulation emerged during 150-350 ms in a left-lateralized cortical network. This network included (i) left lateral occipitotemporal and nearby inferotemporal cortex, (ii) left posterior Sylvian fissure and nearby multimodal cortex, and (iii) medial parietooccipital, posterior cingulate and retrosplenial cortices. Moreover, this left-lateralized network colocalized with word repetition priming effects. Together, results suggest that central saccadic mechanisms influence a left-lateralized language network in occipitotemporal and temporal cortex above and beyond saccadic influences at preceding stages of information processing during visual word recognition.

Neural systems of cognitive demand avoidance

Sayali, C. — 2017-10-31

Cognitive effort is typically aversive, evident in peoples tendency to avoid cognitively demanding tasks. The cost of control hypothesis suggests that engagement of cognitive control systems of the brain makes a task costly and the currency of that cost is a reduction in anticipated rewards. However, prior studies have relied on binary hard versus easy task subtractions to manipulate cognitive effort and so have not tested this hypothesis in "dose-response" fashion. In a sample of 50 participants, we parametrically manipulated the level of effort during fMRI scanning by systematically increasing cognitive control demands during a demand-selection paradigm over six levels. As expected, frontoparietal control network (FPN) activity increased, and reward network activity decreased, as control demands increased across tasks. However, avoidance behavior was not attributable to the change in FPN activity, lending only partial support to the cost of control hypothesis. By contrast, we unexpectedly observed that the deactivation of a task-negative brain network corresponding to the Default Mode Network (DMN) across levels of the cognitive control manipulation predicted the change in avoidance. In summary, we find partial support for the cost of control hypothesis, while highlighting the role of task-negative brain networks in modulating effort avoidance behavior.

Localization of adaptive variants in human genomes using averaged one-dependence estimation

Sugden, L. A. — 2017-12-04

Statistical methods for identifying adaptive mutations from population-genetic data face several obstacles: assessing the significance of genomic outliers, integrating correlated measures of selection into one analytic framework, and distinguishing adaptive variants from hitchhiking neutral variants. Here, we introduce SWIF(r), a probabilistic method that detects selective sweeps by learning the distributions of multiple selection statistics under different evolutionary scenarios and calculating the posterior probability of a sweep at each genomic site. SWIF(r) is trained using simulations from a user-specified demographic model and explicitly models the joint distributions of selection statistics, thereby increasing its power to both identify regions undergoing sweeps and localize adaptive mutations. Using array and exome data from 45 {ddagger}Khomani San hunter-gatherers of southern Africa, we identify an enrichment of adaptive signals in genes associated with metabolism and obesity. SWIF(r) provides a transparent probabilistic framework for localizing beneficial mutations that is extensible to a variety of evolutionary scenarios.

Feature-specific awake reactivation in human V1 after visual training

Bang, J. W. — 2017-12-08

Converging human studies have demonstrated that brain activity patterns observed during task performance reemerge in the following restful awake state. Such "awake reactivation" has been demonstrated across higher-order cortex for complex images or associations. However, it remains unclear what specific training components are reactivated in these studies. Here we sought to provide evidence for the reactivation of a particular visual feature - Gabor orientation. Following extensive training on a visual task, we found robust reactivation in human V1 that lasted at least eight minutes. This effect was not present in higher retinotopic areas such as V2, V3, V3A, or V4v, demonstrating that the effects in V1 are not due to top-down processes such as conscious rehearsal. Furthermore, the amount of awake reactivation predicted the amount of performance improvement on the visual task. These results demonstrate that functionally-relevant awake reactivation of specific visual features occurs in early sensory cortex.

Impaired expected value computations coupled with overreliance on prediction error learning in schizophrenia

Hernaus, D. — 2017-12-22

BackgroundWhile many have emphasized impaired reward prediction error (RPE) signaling in schizophrenia, multiple studies suggest that some decision-making deficits may arise from overreliance on RPE systems together with a compromised ability to represent expected value. Guided by computational frameworks, we formulated and tested two scenarios in which maladaptive representation of expected value should be most evident, thereby delineating conditions that may evoke decision-making impairments in schizophrenia.nnMethodsIn a modified reinforcement learning paradigm, 42 medicated people with schizophrenia (PSZ) and 36 healthy volunteers learned to select the most frequently rewarded option in a 75-25 pair: once when presented with more deterministic (90-10) and once when presented with more probabilistic (60-40) pairs. Novel and old combinations of choice options were presented in a subsequent transfer phase. Computational modeling was employed to elucidate contributions from RPE systems ("actor-critic") and expected value ("Q-leaming").nnResultsPSZ showed robust performance impairments with increasing value difference between two competing options, which strongly correlated with decreased contributions from expected value-based ("Q-leaming") learning. Moreover, a subtle yet consistent contextual choice bias for the "probabilistic" 75 option was present in PSZ, which could be accounted for by a context-dependent RPE in the "actor-critic".nnConclusionsWe provide evidence that decision-making impairments in schizophrenia increase monotonically with demands placed on expected value computations. A contextual choice bias is consistent with overreliance on RPE-based learning, which may signify a deficit secondary to the maladaptive representation of expected value. These results shed new light on conditions under which decisionmaking impairments may arise.

Efficient graph-color compression with neighborhood-informed Bloom filters

Schilken, I. — 2017-12-26

MotivationTechnological advancements in high-throughput DNA sequencing have led to an exponential growth of sequencing data being produced and stored as a byproduct of biomedical research. Despite its public availability, a majority of this data remains hard to query to the research community due to a lack of efficient data representation and indexing solutions. One of the available techniques to represent read data is a condensed form as an assembly graph. Such a representation contains all sequence information but does not store contextual information and metadata.nnResultsWe present two new approaches for a compressed representation of a graph coloring: a lossless compression scheme based on a novel application of wavelet tries as well as a highly accurate lossy compression based on a set of Bloom filters. Both strategies retain a coloring with dynamically changing graph topology. We present construction and merge procedures for both methods and evaluate their performance on a wide range of different datasets. By dropping the requirement of a fully lossless compression and using the topological information of the underlying graph, we can reduce memory requirements by up to three orders of magnitude. Representing individual colors as independently stored modules, our approaches are fully dynamic and can be efficiently parallelized. These properties allow for an easy upscaling to the problem sizes common to the biomedical domain.nnAvailabilityWe provide prototype implementations in C++, summaries of our experiments as well as links to all datasets publicly at https://github.com/ratschlab/graph_annotation.nnContactandre.kahles@inf.ethz.ch, carsten@brown.edu, Gunnar.Ratsch@ratschlab.org

IMPROVEMENTS TO BAYESIAN GENE ACTIVITY STATE ESTIMATION FROM GENOME-WIDE TRANSCRIPTOMICS DATA

Disselkoen, C. — 2017-12-29

An important question in many biological applications, is to estimate or classify gene activity states (active or inactive) based on genome-wide transcriptomics data. Recently, we proposed a Bayesian method, titled MultiMM, which showed superior results compared to existing methods. In short, MultiMM performed better than existing methods on both simulated and real gene expression data, confirming well-known biological results and yielding better agreement with fluxomics data. Despite these promising results, MultiMM has numerous limitations. First, MultiMM leverages co-regulatory models to improve activity state estimates, but information about co-regulation is incorporated in a manner that assumes that networks are known with certainty. Second, MultiMM assumes that genes that change states in the dataset can be distinguished with certainty from those that remain in one state. Third, the model can be sensitive to extreme measures (outliers) of gene expression. In this manuscript, we propose a modified Bayesian approach, which addresses these three limitations by improving outlier handling and by explicitly modeling network and other uncertainty yielding improved gene activity state estimates when compared to MultiMM.

Generalizing Genetic Risk Scores from Europeans to Hispanics/Latinos

Grinde, K. E. — 2018-01-04

Genetic risk scores (GRSs) are weighted sums of risk allele counts of single nucleotide polymorphisms (SNPs) associated with a disease or trait. Construction of GRSs is typically based on published results from Genome-Wide Association Studies (GWASs), the majority of which have been performed in large populations of European ancestry (EA) individuals. While many genotype-trait associations have been shown to generalize from EA populations to other populations, such as Hispanics/Latinos, the optimal choice of SNPs and weights for GRSs may differ between populations due to different linkage disequilibrium (LD) and allele frequency patterns. This is further complicated by the fact that different Hispanic/Latino populations may have different admixture patterns, so that LD and allele frequency patterns may not be the same among non-EA populations. Here, we compare various approaches for GRS construction, using GWAS results from both large EA studies and a smaller study in Hispanics/Latinos, the Hispanic Community Health Study/Study of Latinos (HCHS/SOL, n = 12, 803). We consider multiple ways to select SNPs from association regions and to calculate the SNP weights. We study the performance of the resulting GRSs in an independent study of Hispanics/Latinos from the Woman Health Initiative (WHI, n = 3, 582). We support our investigation with simulation studies of potential genetic architectures in a single locus. We observed that selecting variants based on EA GWASs generally performs well, as long as SNP weights are calculated using Hispanics/Latinos GWASs, or using the meta-analysis of EA and Hispanics/Latinos GWASs. The optimal approach depends on the genetic architecture of the trait.

Galleria mellonella as an Insect Model for P. destructans, the Cause of White-Nose Syndrome in Bats

Beekman, C. N. — 2018-01-08

Pseudogymnoascus destructans is the fungal pathogen responsible for White-nose Syndrome (WNS), a disease that has killed millions of bats in North America over the last decade. A major obstacle to research on P. destructans has been the lack of a tractable infection model for monitoring virulence. Here, we establish a high-throughput model of infection using larvae of Galleria mellonella, an invertebrate used to study host-pathogen interactions for a wide range of microbial species. We demonstrate that P. destructans can kill G. mellonella larvae in an inoculum-dependent manner when infected larvae are housed at 13{degrees}C or 18{degrees}C. Larval killing is an active process, as heat-killed P. destructans spores caused significantly decreased levels of larval death compared to live spores. We also show that fungal spores that were germinated prior to inoculation were able to kill larvae 3-4 times faster than non-germinated spores. Lastly, we identified chemical inhibitors of P. destructans and used G. mellonella to evaluate these inhibitors for their ability to reduce virulence. We demonstrate that two chemicals, trifluoperazine and amphotericin B, can effectively block larval killing by P. destructans and thereby establish that this infection model can be used to screen biocontrol agents against this fungal pathogen.

Improved phylogenetic resolution within Siphonophora (Cnidaria) with implications for trait evolution

Munro, C. — 2018-01-20

Siphonophores are a diverse group of hydrozoans (Cnidaria) that are found at all depths of the ocean - from the surface, like the familiar Portuguese man of war, to the deep sea. Siphonophores play an important role in ocean ecosystems, and are among the most abundant gelatinous predators. A previous phylogenetic study based on two ribosomal RNA genes provided insight into the internal relationships between major siphonophore groups, however there was little support for many deep relationships within the clade Codonophora. Here, we present a new siphonophore phylogeny based on new transcriptome data from 30 siphonophore species analyzed in combination with 13 publicly available genomic and transcriptomic datasets. We use this new phylogeny to reconstruct several traits that are central to siphonophore biology, including sexual system (monoecy vs. dioecy), gain and loss of zooid types, life history traits, and habitat. The phylogenetic relationships in this study are largely consistent with the previous phylogeny, but we find strong support for new clades within Codonophora that were previously unresolved. These results have important implications for trait evolution within Siphonophora, including favoring the hypothesis that monoecy arose twice.

The Eighty Five Percent Rule for Optimal Learning

Wilson, R. C. — 2018-01-27

Researchers and educators have long wrestled with the question of how best to teach their clients be they human, animal or machine. Here we focus on the role of a single variable, the difficulty of training, and examine its effect on the rate of learning. In many situations we find that there is a sweet spot in which training is neither too easy nor too hard, and where learning progresses most quickly. We derive conditions for this sweet spot for a broad class of learning algorithms in the context of binary classification tasks, in which ambiguous stimuli must be sorted into one of two classes. For all of these gradient-descent based learning algorithms we find that the optimal error rate for training is around 15.87% or, conversely, that the optimal training accuracy is about 85%. We demonstrate the efficacy of this Eighty Five Percent Rule for artificial neural networks used in AI and biologically plausible neural networks thought to describe human and animal learning.

Emergence of Human Amygdala Functional Networks: 3 Months to 5 Years of Age

Gabard-Durnam, L. J. — 2018-02-07

Although the amygdalas role in shaping social behavior is especially important during early post-natal development, very little is known of human amygdala functional development before childhood. To address this important gap, this study used resting-state fMRI to examine early functional network development of the amygdala and its subregions in 80 participants from 3-months to 5-years of age. Whole brain functional connectivity with the whole amygdala and its laterobasal and superficial nuclear groups were largely similar to those seen in older children and adults, and functional distinctions between subregion networks exist already. These patterns suggest many amygdala functional circuits are intact from infancy, especially those that are part of larger motor, visual, auditory and subcortical (basal ganglia especially) networks. Notably, these observed robust amygdala functional networks in infancy precede reports to date of elicited amygdala reactivity in development. Developmental changes in connectivity were observed between the laterobasal nucleus and bilateral ventral temporal and motor cortex as well as between the superficial nuclei and medial thalamus, occipital cortex and a different region of motor cortex. These results show amygdala-subcortical and sensory-cortex connectivity begins refinement prior to childhood, though connectivity changes with associative and frontal cortical areas, seen after early childhood, were not evident in this age range. These findings represent early steps in understanding amygdala network dynamics across infancy through early childhood, an important period of emotional and cognitive development.

Measurement error and variant-calling in deep Illumina sequencing of HIV

Howison, M. — 2018-03-05

MotivationNext-generation deep sequencing of viral genomes, particularly on the Illumina platform, is increasingly applied in HIV research. Yet, there is no standard protocol or method used by the research community to account for measurement errors that arise during sample preparation and sequencing. Correctly calling high and low frequency variants while controlling for erroneous variant calls is an important precursor to downstream interpretation, such as studying the emergence of HIV drug-resistance mutations, which in turn has clinical applications and can improve patient care.nnResultsWe developed a new variant-calling pipeline, hivmmer, for Illumina sequences from HIV viral genomes. First, we validated hivmmer by comparing it to other variant-calling pipelines on real HIV plasmid data sets, which have known sequences. We found that hivmmer achieves a lower rate of erroneous variant calls, and that all methods agree on the frequency of correctly called variants. Next, we compared the methods on an HIV plasmid data set that was sequenced using an amplicon-tagging protocol called Primer ID, which is designed to reduce errors and amplification bias during library preparation. We show that the Primer ID consensus does indeed have fewer erroneous variant calls compared to the variant-calling pipelines, and that hivmmer more closely approaches this low error rate compared to the other pipelines. Surprisingly, the frequency estimates from the Primer ID consensus do not differ significantly from those of the variant-calling pipelines. Finally, we built a predictive model for classifying errors in the hivmmer alignment, and show that it achieves high accuracy for identifying erroneous variant calls.nnAvailabilityhivmmer is freely available for non-commercial use from https://github.com/mhowison/hivmmer.nnContactmhowison@brown.edu

Genome wide association analysis identifies genetic variants associated with reproductive variation across domestic dog breeds and uncovers links to domestication

Phillips, J. B. — 2018-03-20

The diversity of eutherian reproductive strategies has led to variation in many traits, such as number of offspring, age of reproductive maturity, and gestation length. While reproductive trait variation has been extensively investigated and is well established in mammals, the genetic loci contributing to this variation remain largely unknown. The domestic dog, Canis lupus familiaris is a powerful model for studies of the genetics of inherited disease due to its unique history of domestication. To gain insight into the genetic basis of reproductive traits across domestic dog breeds, we collected phenotypic data for four traits - cesarean section rate (n = 97 breeds), litter size (n = 60), stillbirth rate (n = 57), and gestation length (n = 23) - from primary literature and breeders handbooks. By matching our phenotypic data to genomic data from the Cornell Veterinary Biobank, we performed genome wide association analyses for these four reproductive traits, using body mass and kinship among breeds as co-variates. We identified 14 genome-wide significant associations between these traits and genetic loci, including variants near CACNA2D3 with gestation length, MSRB3 with litter size, SMOC2 with cesarean section rate, MITF with litter size and still birth rate, KRT71 with cesarean section rate, litter size, and stillbirth rate, and HTR2C with stillbirth rate. Some of these loci, such as CACNA2D3 and MSRB3, have been previously implicated in human reproductive pathologies. Many of the variants that we identified have been previously associated with domestication-related traits, including brachycephaly (SMOC2), coat color (MITF), coat curl (KRT71), and tameness (HTR2C). These results raise the hypothesis that the artificial selection that gave rise to dog breeds also shaped the observed variation in their reproductive traits. Overall, our work establishes the domestic dog as a system for studying the genetics of reproductive biology and disease.

A Global Analysis of Mutations Accompanying Microevolution in the Heterozygous Diploid Pathogen Candida albicans

Ene, I. V. — 2018-03-25

Candida albicans is a heterozygous diploid yeast that is a commensal of the human gastrointestinal (GI) tract and a prevalent opportunistic pathogen. Here, whole-genome sequencing was performed on multiple C. albicans isolates passaged in different niches to characterize the complete spectrum of mutations arising during microevolution. We reveal that evolution during short time-scales (<600 generations) is driven by both de novo base substitutions and short-tract loss of heterozygosity (LOH) events. In contrast, large-scale chromosomal changes are relatively rare, although chromosome 7 trisomies repeatedly emerged during passaging in one GI colonization model. Both strain background and chromosomal features affected mutational patterns, with mutation rates being greatly elevated in regions adjacent to emergent LOH tracts. Mutation rates were also elevated during host infection where genomes showed strong evidence of purifying selection. These results establish the genetic events driving C. albicans evolution and that this heterozygous diploid is extensively shaped by purifying selection.

Geometry-dependent instabilities in electrically excitable tissues

McNamara, H. M. — 2018-04-02

Little is known about how individual cells sense the macroscopic geometry of their tissue environment. Here we explore whether long-range electrical signaling can convey information on tissue geometry to influence electrical dynamics of individual cells. First, we studied an engineered electrically excitable cell line where all voltage-gated channels were well characterized. Cells grown in patterned islands of different shapes showed remarkably diverse firing patterns under otherwise identical conditions, including regular spiking, period-doubling alternans, and arrhythmic firing. A Hodgkin-Huxley numerical model quantitatively reproduced these effects, showing how the macroscopic geometry affected the single-cell electrophysiology via the influence of gap junction-mediated electrical coupling. Qualitatively similar geometry dependent dynamics were experimentally observed in human induced pluripotent stem cell (iPSC)-derived cardiomyocytes. The cardiac results urge caution in translating observations of arrhythmia in vitro to predictions in vivo where the tissue geometry is very different. We present simulation results and scaling arguments which explore how to extrapolate electrophysiological measurements between tissues with different geometries and different gap junction couplings.

Prognostic models for Ebola virus disease derived from data collected at five treatment units in Sierra Leone and Liberia: performance, external validation, and risk visualization

Colubri, A. — 2018-04-06

BackgroundWe created a family of prognostic models for Ebola virus disease from the largest dataset of EVD patients published to date. We incorporated these models into an app, "Ebola Care Guidelines", that provides access to recommended, evidence-based supportive care guidelines and highlights the signs/symptoms with the largest contribution to prognosis.nnMethodsWe applied multivariate logistic regression on 470 patients admitted to five Ebola treatment units in Liberia and Sierra Leone during the 2014-16 outbreak. We validated the models with two independent datasets from Sierra Leone.nnFindingsViral load and age were the most important predictors of death. We generated a parsimonious model including viral load, age, body temperature, bleeding, jaundice, dyspnea, dysphagia, and referral time recorded at triage. We also constructed fallback models for when variables in the parsimonious model are unavailable. The performance of the parsimonious model approached the predictive power of observational wellness assessments by experienced health workers, with Area Under the Curve (AUC) ranging from 0.7 to 0.8 and overall accuracy of 64% to 74%.nnInterpretationMachine-learning models and mHealth tools have the potential for improving the standard of care in low-resource settings and emergency scenarios, but data incompleteness and lack of generalizable models are major obstacles. We showed how harmonization of multiple datasets yields prognostic models that can be validated across different cohorts. Similar performance between the parsimonious model and those incorporating expert wellness assessments suggests that clinically-guided machine learning approaches can recapitulate clinical expertise, and thus be useful when such expertise is unavailable. We also demonstrated with our guidelines app how integration of those models with mobile technologies enables deployable clinical management support tools that facilitate access to comprehensive bodies of medical knowledge.nnFundingHoward Hughes Medical Institute, US National Institutes of Health

Antimicrobial Efficacy and Safety of a Novel Gas Plasma-Activated Catheter Lock Solution

Bhatt, S. — 2018-04-17

Antimicrobial lock solutions are important for prevention of microbial colonization and infection of long-term central venous catheters. We investigated the efficacy and safety of a novel antibiotic-free lock solution formed from gas plasma-activated disinfectant (PAD). Using a luminal biofilm model, viable cells of methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Candida albicans in mature biofilms were reduced by 6 - 8 orders of magnitude with a PAD lock for 60 minutes. Subsequent 24-hour incubation of PAD-treated samples resulted in no detectable regrowth of viable bacteria or fungi. As a comparison, the use of a minocycline/EDTA/ethanol lock solution for 60 minutes led to regrowth of bacteria and fungi, up to 107 - 109 CFU/ml, in 24 hours. The PAD lock solution had minimal impact on human umbilical vein endothelial cell viability, whereas the minocycline/EDTA/ethanol solution elicited cell death in nearly half of human endothelial cells. Additionally, PAD treatment caused little topological change to catheter materials. In conclusion, PAD represents a novel antibiotic-free, non-cytotoxic lock solution that elicits rapid and broad-spectrum eradication of biofilm-laden microbes and which shows promise for the prevention and treatment of intravascular catheter infections.

Counting with DNA in metabarcoding studies: how should we convert sequence reads to dietary data?

Deagle, B. — 2018-04-18

Advances in DNA sequencing technology have revolutionised the field of molecular analysis of trophic interactions and it is now possible to recover counts of food DNA barcode sequences from a wide range of dietary samples. But what do these counts mean? To obtain an accurate estimate of a consumers diet should we work strictly with datasets summarising the frequency of occurrence of different food taxa, or is it possible to use the relative number of sequences? Both approaches are applied in the dietary metabarcoding literature, but occurrence data is often promoted as a more conservative and reliable option due to taxa-specific biases in recovery of sequences. Here, we point out that diet summaries based on occurrence data overestimate the importance of food consumed in small quantities (potentially including low-level contaminants) and are sensitive to the count threshold used to define an occurrence. Our simulations indicate that even with recovery biases incorporated, using relative read abundance (RRA) information can provide a more accurate view of population-level diet in many scenarios. The ideas presented here highlight the need to consider all sources of bias and to justify the methods used to interpret count data in dietary metabarcoding studies. We encourage researchers to continue to addressing methodological challenges, and acknowledge unanswered questions to help spur future investigations in this rapidly developing area of research.

The Genome of the Human Pathogen Candida albicans is Shaped by Mutation and Cryptic Sexual Recombination

Wang, J. M. — 2018-04-28

The opportunistic fungal pathogen Candida albicans lacks a conventional sexual program and is thought to evolve, at least primarily, through the clonal acquisition of genetic changes. Here, we performed an analysis of heterozygous diploid genomes from 21 clinical isolates to determine the natural evolutionary processes acting on the C. albicans genome. Consistent with a model of inheritance by descent, most single nucleotide polymorphisms (SNPs) were shared between closely related strains. However, strain-specific SNPs and insertions/deletions (indels) were distributed non-randomly across the genome. For example, base substitution rates were higher in the immediate vicinity of indels, and heterozygous regions of the genome contained significantly more strain-specific polymorphisms than homozygous regions. Loss of heterozygosity (LOH) events also contributed substantially to genotypic variation, with most long-tract LOH events extending to the ends of the chromosomes suggestive of repair via break-induced replication. Importantly, some isolates contained highly mosaic genomes and failed to cluster closely with other isolates within their assigned clades. Mosaicism is consistent with strains having experienced inter-clade recombination during their evolutionary history and a detailed examination of nuclear and mitochondrial genomes revealed striking examples of recombination. Together, our analyses reveal that both (para)sexual recombination and mitotic mutational processes drive evolution of this important pathogen in nature. To further facilitate the study of genome differences we also introduce an online platform, SNPMap, to examine SNP patterns in sequenced C. albicans genomes.nnAUTHOR SUMMARYMutations introduce variation into the genome upon which selection can act. Defining the nature of these changes is critical for determining species evolution, as well as for understanding the genetic changes driving important cellular processes such as carcinogenesis. The fungus Candida albicans is a heterozygous diploid species that is both a frequent commensal organism and a prevalent opportunistic pathogen. Prevailing theory is that C. albicans evolves primarily through the gradual build-up of mutations, and a pressing question is whether sexual or parasexual processes also operate within natural populations. Here, we determine the evolutionary patterns of genetic change that have accompanied species evolution in nature by examining genomic differences between clinical isolates. We establish that the C. albicans genome evolves by a combination of base-substitution mutations, insertions/deletion events, and both short-tract and long-tract loss of heterozygosity (LOH) events. These mutations are unevenly distributed across the genome, and reveal that non-coding regions and heterozygous regions are evolving more quickly than coding regions and homozygous regions, respectively. Furthermore, we provide evidence that genetic exchange has occurred between isolates, establishing that sexual or parasexual processes have transpired in C. albicans populations and contribute to the diversity of both nuclear and mitochondrial genomes.

Bayesian Estimation of 3D Chromosomal Structure from Single Cell Hi-C Data

Rosenthal, M. — 2018-05-07

The problem of 3D chromosome structure inference from Hi-C datasets is important and challenging. While bulk Hi-C datasets contain contact information derived from millions of cells, and can capture major structural features shared by the majority of cells in the sample, they do not provide information about local variability between cells. Single cell Hi-C can overcome this problem, but contact matrices are generally very sparse, making structural inference more problematic. We have developed a Bayesian multiscale approach, named SIMBA3D, to infer 3D structures of chromosomes from single cell Hi-C while including the bulk Hi-C data and some regularization terms as a prior. We study the landscape of solutions for each single-cell Hi-C dataset as a function of prior strength and demonstrate clustering of solutions using data from the same cell.

Genetic determinants of risk and survival in pulmonary arterial hypertension

Rhodes, C. — 2018-05-16

BackgroundPulmonary arterial hypertension (PAH) is a rare disorder leading to premature death. Rare genetic variants contribute to disease etiology but the contribution of common genetic variation to disease risk and outcome remains poorly characterized.nnMethodsWe performed two separate genome-wide association studies of PAH using data across 11,744 European-ancestry individuals (including 2,085 patients), one with genotypes from 5,895 whole genome sequences and another with genotyping array data from 5,849 further samples. Cross-validation of loci reaching genome-wide significance was sought by meta-analysis. We functionally annotated associated variants and tested associations with duration of survival.nnFindingsA locus at HLA-DPA1/DPB1 within the class II major histocompatibility (MHC) region and a second near SOX17 were significantly associated with PAH. The SOX17 locus contained two independent signals associated with PAH. Functional and epigenomic data indicate that the risk variants near SOX17 alter gene regulation via an enhancer active in endothelial cells. PAH risk variants determined haplotype-specific enhancer activity and CRISPR-inhibition of the enhancer reduced SOX17 expression. Analysis of median survival showed that PAH patients with two copies of the HLA-DPA1/DPB1 risk variant had a two-fold difference (>16 years versus 8 years), compared to patients homozygous for the alternative allele.nnInterpretationWe have found that common genetic variation at loci in HLA-DPA1/DPB1 and an enhancer near SOX17 are associated with PAH. Impairment of Sox17 function may be more common in PAH than suggested by rare mutations in SOX17. Allelic variation at HLA-DPB1 stratifies PAH patients for survival following diagnosis, with implications for future therapeutic trial design.nnFundingUK NIHR, BHF, UK MRC, Dinosaur Trust, NIH/NHLBI, ERS, EMBO, Wellcome Trust, EU, AHA, ACClinPharm, Netherlands CVRI, Dutch Heart Foundation, Dutch Federation of UMC, Netherlands OHRD and RNAS, German DFG, German BMBF, APH Paris, Inserm, Universite Paris-Sud, and French ANR.

Positive reward prediction errors strengthen incidental memory encoding

Jang, A. — 2018-05-21

The dopamine system is thought to provide a reward prediction error signal that facilitates reinforcement learning and reward-based choice in corticostriatal circuits. While it is believed that similar prediction error signals are also provided to temporal lobe memory systems, the impact of such signals on episodic memory encoding has not been fully characterized. Here we develop an incidental memory paradigm that allows us to 1) estimate the influence of reward prediction errors on the formation of episodic memories, 2) dissociate this influence from other factors such as surprise and uncertainty, 3) test the degree to which this influence depends on temporal correspondence between prediction error and memoranda presentation, and 4) determine the extent to which this influence is consolidation-dependent. We find that when choosing to gamble for potential rewards during a primary decision making task, people encode incidental memoranda more strongly even though they are not aware that their memory will be subsequently probed. Moreover, this strengthened encoding scales with the reward prediction error, and not overall reward, experienced selectively at the time of memoranda presentation (and not before or after). Finally, this strengthened encoding is identifiable within a few minutes and is not substantially enhanced after twenty-four hours, indicating that it is not consolidation-dependent. These results suggest a computationally and temporally specific role for putative dopaminergic reward prediction error signaling in memory formation.

Base-pair Ambiguity and the Kinetics of RNA Folding: a Hypothesis-Driven Statistical Analysis

Zhou, G. — 2018-05-25

BackgroundA folding RNA molecule encounters multiple opportunities to form non-native yet energetically favorable pairings of nucleotide sequences. Given this forbidding free-energy landscape, mechanisms have evolved that contribute to a directed and efficient folding process, including catalytic proteins and error-detecting chaperones. Among structural RNA molecules we make a distinction between "bound" molecules, which are active as part of ribonucleoprotein (RNP) complexes, and "unbound," with physiological functions performed without necessarily being bound in RNP complexes. We hypothesized that unbound molecules, lacking the partnering structure of a protein, would be more vulnerable than bound molecules to kinetic traps that compete with native stem structures. We defined an "ambiguity index"--a normalized function of the primary and secondary structure of an individual molecule that measures the number of kinetic traps available to nucleotide sequences that are paired in the native structure, presuming that unbound molecules would have lower indexes. The ambiguity index depends on the purported secondary structure, and was computed under both the comparative ("gold standard") and an equilibrium-based prediction which approximates the minimum free energy (MFE) structure. Arguing that kinetically accessible metastable structures might be more biologically relevant than thermodynamic equilibrium structures, we also hypothesized that MFE-derived ambiguities would be less effective in separating bound and unbound molecules.nnResultsWe have introduced an intuitive and easily computed function of primary and secondary structures that measures the availability of complementary sequences that could disrupt the formation of native stems on a given molecule--an ambiguity index. Using comparative secondary structures, the ambiguity index is systematically smaller among unbound than bound molecules, as expected. Furthermore, the effect is lost when the presumably more accurate comparative structure is replaced instead by the MFE structure.nnConclusionsA statistical analysis of the relationship between the primary and secondary structures of non-coding RNA molecules suggests that stem-disrupting kinetic traps are substantially less prevalent in molecules not participating in RNP complexes. In that this distinction is apparent under the comparative but not the MFE secondary structure, the results highlight a possible deficiency in structure predictions when based upon assumptions of thermodynamic equilibrium.

Forebrain dopamine value signals arise independently from midbrain dopamine cell firing.

Mohebi, A. — 2018-05-30

The mesolimbic dopamine projection from the ventral tegmental area (VTA) to nucleus accumbens (NAc) is a key pathway for reward-driven learning, and for the motivation to work for more rewards. VTA dopamine cell firing can encode reward prediction errors (RPEs1,2), vital learning signals in computational theories of adaptive behavior. However, NAc dopamine release more closely resembles reward expectation (value), a motivational signal that invigorates approach behaviors3-7. This discrepancy might be due to distinct behavioral contexts: VTA dopamine cells have been recorded under head-fixed conditions, while NAc dopamine release has been measured in actively-moving subjects. Alternatively the mismatch may reflect changes in the tonic firing of dopamine cells8, or a fundamental dissociation between firing and release. Here we directly compare dopamine cell firing and release in the same adaptive decision-making task. We show that dopamine release covaries with reward expectation in two specific forebrain hotspots, NAc core and ventral prelimbic cortex. Yet the firing rates of optogenetically-identified VTA dopamine cells did not correlate with reward expectation, but instead showed transient, error-like responses to unexpected cues. We conclude that critical motivation-related dopamine dynamics do not arise from VTA dopamine cell firing, and may instead reflect local influences over forebrain dopamine varicosities.

Data-Constrained Biophysical Modeling Of Human VIM To Assess Contributions Of Intrinsic Oscillatory Activity To Essential Tremor

Lee, S. — 2018-06-07

Essential tremor (ET) is the most common movement disorder, in which the primary symptom is a prominent, involuntary 4-10 Hz rhythmic movement. The presence of tremor frequency oscillations (TFOs) in the ventral intermediate nucleus of the thalamus (VIM) is well-established, but it is often assumed that it is driven by cerebellar tremor frequency activity, while the role of intrinsic oscillatory activity in VIM is not clear. An improved understanding of the mechanisms of tremor and non-tremor frequency activity in VIM is critical to the development of improved pharmacological and neuromodulatory therapies. Starting from a canonical model of thalamus, we developed a biophysically-principled computational model of tremor field activity in the VIM, coupled with the thalamic reticular nucleus (TRN). We simulated TFOs in the model generated either by extrinsic tremor-periodic drive or intrinsic VIM-TRN interaction to understand whether these networks exhibited distinct biophysical properties, which may impact the efficacy of pharmacological or stimulation treatment for TFOs. Extrinsic and intrinsic TFOs in the model depended on T-type Ca2+ channels in different ways. Each also depended on GABA modulation in a site- and type-specific manner. These results suggested that efficacy of pharmacological manipulations may depend upon the mechanisms generating TFOs in VIM. Simulated non-tremor-related motor activity from cerebellum decreased extrinsic but increased intrinsic TFOs. Our results suggest that both mechanisms may be important to understand the emergence and cessation of TFOs in VIM and lead to experimentally testable predictions on how to modulate tremor frequency activity to improve treatment strategies for ET.nnSignificance StatementEssential Tremor (ET) is a movement disorder in which the primary symptom is a prominent, involuntary, and rhythmic shaking, often of the hands. Electrical activity in many areas of the brain exhibit rhythmicity related to the patients tremor. One such area resides in a structure called the thalamus, but it is not fully known what gives rise to tremor-related activity. We created a computational model of this activity, which suggested how to differentiate tremor mechanisms and how these differences may contribute to other impairments in ET. Knowledge of the biophysical mechanisms contributing to tremor can ultimately lead to improvements in treatments to alleviate symptoms of ET.

Quantifying platelet margination in diabetic blood flow

Chang, H.-Y. — 2018-06-12

Patients with type 2 diabetes mellitus (T2DM) develop thrombotic abnormalities strongly associated with cardiovascular diseases. In addition to the changes of numerous coagulation factors such as elevated levels of thrombin and fibrinogen, the abnormal rheological effects of red blood cells (RBCs) and platelets flowing in blood are crucial in platelet adhesion and thrombus formation in T2DM. An important process contributing to the latter is the platelet margination. We employ the dissipative particle dynamics method to seamlessly model cells, plasma, and vessel walls. We perform a systematic study on the RBC and platelet transport in cylindrical vessels by considering different cell shapes, sizes and RBC deformabilities in healthy and T2DM blood, as well as variable flowrates and hematocrit. In particular, we use cellular-level RBC and platelet models with parameters derived from patient-specific data and present a sensitivity study. We find T2DM RBCs, which are less deformable compared to normal RBCs, lower the transport of platelets toward the vessel walls whereas platelets with higher mean volume (often observed in T2DM) lead to enhanced margination. Furthermore, increasing the flowrate or hematocrit enhances platelet margination. We also investigated the effect of platelet shape and observed a non-monotonic variation with the highest near-wall concentration corresponding to platelets with moderate aspect ratio of 0.38. We examine the role of white blood cells (WBCs), whose count is increased notably in T2DM patients. We find that WBC rolling or WBC adhesion tend to decrease platelet margination due to hydrodynamic effects. To the best of our knowledge, such simulations of blood including all blood cells have not been performed before, and our quantitative findings can help separate the effects of hydrodynamic interactions from adhesive interactions, and potentially shed light on the associated pathological processes in T2DM such as increased inflammatory response, platelet activation and adhesion, and ultimately thrombus formation.

Getting there and staying there: supporting and enabling persistent human life on Mars using synthetic natural rubber, self-healing materials, and biological batteries.

Acharya, N. — 2018-06-12

Planetary exploration requires a balance between preemptive planning and financial feasibility. The risk of mid-mission equipment failure, power shortages, or supply depletion incentivizes precautionary measures, but the financial strain of sending unnecessary mass into space limits this practice.nnTo balance the two, our team explored the advantages of biological solutions, namely the self-sustaining abilities of low-mass organisms, to make planetary exploration more self-sufficient and economical. Prioritizing repair over replacement, we are developing self-healing materials embedded with Bacillus subtilis. For longer-lasting energy, we are designing a "biobactery" using linearly oriented Escherichia coli to generate power. For renewable materials, we are engineering bacteria to synthesize and degrade rubber. Individually, these projects offer sustainable alternatives for repair, power, and materials. But when combined, these consolidated insights can provide us with the power to get to Mars and resources to sustain us while were there.

Smoking induces coordinated DNA methylation and gene expression changes in adipose tissue with consequences for metabolic health

Tsai, P.-C. — 2018-06-21

Tobacco smoking is a risk factor for multiple diseases, including cardiovascular disease and diabetes. Many smoking-associated signals have been detected in the blood methylome, but the extent to which these changes are widespread to metabolically relevant tissues, and impact gene expression or cardio-metabolic health, remains unclear.nnWe investigated smoking-associated DNA methylation and gene expression variation in adipose tissue from 542 healthy female twins with available well-characterized cardio-metabolic phenotype profiles. We identified 42 smoking-methylation and 42 smoking-expression signals, where five genes (AHRR, CYP1A1, CYP1B1, CYTL1, F2RL3) were both hypo-methylated and up-regulated in smokers. We replicated and validated a proportion of the signals in blood, adipose, skin, and lung tissue datasets, identifying tissue-shared effects. Smoking leaves systemic imprints on DNA methylation after smoking cessation, with stronger but shorter-lived effects on gene expression. We tested for associations between the observed smoking signals and several adiposity phenotypes that constitute cardio-metabolic disease risk. Visceral fat and android/gynoid ratio were associated with methylation at smoking-markers with functional impacts on expression, such as CYP1A1, and in signals shared across tissues, such as NOTCH1. At smoking-signals BHLHE40 and AHRR DNA methylation and gene expression levels in current smokers were predictive of future gain in visceral fat upon smoking cessation.nnOur results provide the first comprehensive characterization of coordinated DNA methylation and gene expression markers of smoking in adipose tissue, a subset of which link to human cardio-metabolic health and may give insights into the wide ranging risk effects of smoking across the body.nnAuthor SummaryTobacco smoking is the strongest environmental risk factor for human disease. Here, we investigate how smoking systemically changes methylome and transcriptome signatures in multiple tissues in the human body. We observe strong and coordinated epigenetic and gene expression changes in adipose tissue, some of which are mirrored in blood, skin, and lung tissue. Smoking leaves a strong short-lived impact on gene expression levels, while methylation changes are long-lasting after smoking cessation. We investigated if these changes observed in a metabolically-relevant (adipose) tissue had impacts on human disease, and observed strong associations with cardio-metabolic disease traits. Some of the smoking signals could predict future gain in obesity and cardio-metabolic disease risk in current smokers who subsequently go on to quit smoking. Our results provide novel insights into understanding the widespread health consequence of smoking outside the lung.

Signals of polygenic adaptation on height have been overestimated due to uncorrected population structure in genome-wide association studies

Sohail, M. — 2018-06-25

Genetic predictions of height differ among human populations and these differences are too large to be explained by genetic drift. This observation has been interpreted as evidence of polygenic adaptation. Differences across populations were detected using SNPs genome-wide significantly associated with height, and many studies also found that the signals grew stronger when large numbers of subsignificant SNPs were analyzed. This has led to excitement about the prospect of analyzing large fractions of the genome to detect subtle signals of selection and claims of polygenic adaptation for multiple traits. Polygenic adaptation studies of height have been based on SNP effect size measurements in the GIANT Consortium meta-analysis. Here we repeat the height analyses in the UK Biobank, a much more homogeneously designed study. Our results show that polygenic adaptation signals based on large numbers of SNPs below genome-wide significance are extremely sensitive to biases due to uncorrected population structure.

Dissociable forms of uncertainty-driven representational change across the human brain.

Nassar, M. R. — 2018-07-07

Environmental change can lead decision makers to shift rapidly among different behavioral regimes. These behavioral shifts can be accompanied by rapid changes in the firing pattern of neural networks. However, it is unknown what the populations of neurons that participate in such "network reset" phenomena are representing. Here we examined 1) whether and where rapid changes in multivariate activity patterns are observable with fMRI during periods of rapid behavioral change, and 2) what types of representations give rise to these phenomena. We did so by examining fluctuations in multi-voxel patterns of BOLD activity from human subjects making sequential inferences about the state of a partially observable and discontinuously changing variable. We found that, within the context of this sequential inference task, the multivariate patterns of activity in a number of cortical regions contain representations that change more rapidly during periods of uncertainty following a change in behavioral context. In motor cortex, this phenomenon was indicative of discontinuous change in behavioral outputs, whereas in visual regions the same basic phenomenon was evoked by tracking of salient environmental changes. In most other cortical regions, including dorsolateral prefrontal and anterior cingulate cortex, the phenomenon was most consistent with directly encoding the degree of uncertainty. However, in a few other regions, including orbitofrontal cortex, the phenomenon was best explained by representations of a shifting context that evolve more rapidly during periods of rapid learning. These representations may provide a dynamic substrate for learning that facilitates rapid disengagement from learned responses during periods of change.

Long term impact of intensive post graduate laboratory training at the Cold Spring Harbor Neurobiology of Drosophila summer course

Ly, S. — 2018-07-16

Intensive postgraduate courses provide an opportunity for junior and senior level scientists to learn concepts and techniques that will advance their training and research programs. It is commonly assumed that short intensive courses have positive impacts within fields of research; however, these assumptions are rarely tested. Here we describe the framework of a long running postgraduate summer course at Cold Spring Harbor and attempt to quantify the impact made over its history. For over three decades, the Drosophila Neurobiology: Genes, Circuits & Behavior Summer Course at Cold Spring Harbor Laboratories (CSHL) has provided participants with intense instruction on a wide variety of topics and techniques in integrative neuroscience using Drosophila as a model organism. Students are introduced to the latest approaches for studying nervous system development, activity and connectivity, as well as complex behaviors and diseases. The course has a long history of successful alumni, many of whom describe participation in the course as foundational to their training. Student surveys of recent participants indicate a high level of satisfaction, improved career outcomes, and direct impact on publications. Analysis of student success reveals that over 64% of participants obtain independent faculty positions. Further, we describe ongoing efforts to enhance diversity and encourage access to scientific research at undergraduate-focused institutions. Together, our findings suggest that laboratory-intensive postgraduate courses provide a highly effective mechanism for scientific training that has lasting positive impacts on trainees.

Fast and general-purpose linear mixed models for genome-wide genetics

Runcie, D. E. — 2018-07-23

Linear mixed effect models are powerful tools used to account for population structure in genome-wide association studies (GWASs) and estimate the genetic architecture of complex traits. However, fully-specified models are computationally demanding and common simplifications often lead to reduced power or biased inference. We describe Grid-LMM (https://github.com/deruncie/GridLMM), an extendable algorithm for repeatedly fitting complex linear models that account for multiple sources of heterogeneity, such as additive and non-additive genetic variance, spatial heterogeneity, and genotype-environment interactions. Grid-LMM can compute approximate (yet highly accurate) frequentist test statistics or Bayesian posterior summaries at a genome-wide scale in a fraction of the time compared to existing general-purpose methods. We apply Grid-LMM to two types of quantitative genetic analyses. The first is focused on accounting for spatial variability and non-additive genetic variance while scanning for QTL; and the second aims to identify gene expression traits affected by non-additive genetic variation. In both cases, modeling multiple sources of heterogeneity leads to new discoveries.nnAuthor summaryThe goal of quantitative genetics is to characterize the relationship between genetic variation and variation in quantitative traits such as height, productivity, or disease susceptibility. A statistical method known as the linear mixed effect model has been critical to the development of quantitative genetics. First applied to animal breeding, this model now forms the basis of a wide-range of modern genomic analyses including genome-wide associations, polygenic modeling, and genomic prediction. The same model is also widely used in ecology, evolutionary genetics, social sciences, and many other fields. Mixed models are frequently multi-faceted, which is necessary for accurately modeling data that is generated from complex experimental designs. However, most genomic applications use only the simplest form of linear mixed methods because the computational demands for model fitting can be too great. We develop a flexible approach for fitting linear mixed models to genome scale data that greatly reduces their computational burden and provides flexibility for users to choose the best statistical paradigm for their data analysis. We demonstrate improved accuracy for genetic association tests, increased power to discover causal genetic variants, and the ability to provide accurate summaries of model uncertainty using both simulated and real data examples.

Persistent Gamma Spiking in Non-Sensory Fast-Spiking Cells Predicts Perceptual Success

Shin, H. — 2018-07-25

Persistent gamma oscillations (30-55Hz) are hypothesized to temporally coordinate stimulus encoding, enabling perception. This prediction poses a conundrum: How can gamma serve as a template when the stimulus itself drives its mediators, presumably perturbing its maintenance? Specifically, fast-spiking interneurons (FS), a key gamma generator, can be highly sensory responsive. Further, the gamma-band local field potential (LFP) shows properties inconsistent with temporal coordination. Combining tetrode recording with controlled psychophysics revealed an FS subtype ({gamma}nsFS) that was not sensory responsive, whose inter-spike intervals peaked at gamma, and that fired with higher periodicity than other FS. Successful detection was predicted by increased regularity in {gamma}nsFS spiking at gamma, persisting from before to after sensory onset. In contrast, gamma LFP power negatively predicted detection, and was negatively related to gamma band spiking by {gamma}nsFS. These results suggest that a distinct interneuron subgroup, not distracted by sensory input, mediates perceptually-relevant oscillations independent of LFP.

Genome-wide Marginal Epistatic Association Mapping in Case-Control Studies

Crawford, L. — 2018-07-23

Epistasis, commonly defined as the interaction between genetic loci, is an important contributor to the genetic architecture underlying many complex traits and common diseases. Most existing epistatic mapping methods in genome-wide association studies explicitly search over all pairwise or higher-order interactions. However, due to the potentially large search space and the resulting multiple testing burden, these conventional approaches often suffer from heavy computational cost and low statistical power. A recently proposed attractive alternative for mapping epistasis focuses instead on detecting marginal epistasis, which is defined as the combined pairwise interaction effects between a given variant and all other variants. By searching for marginal epistatic effects, one can identify genetic variants that are involved in epistasis without the need to identify the exact partners with which the variants interact -- thus, potentially alleviating much of the statistical and computational burden associated with conventional epistatic mapping procedures. However, previous marginal epistatic mapping methods are based on quantitative trait models. As we will show here, these lack statistical power in case-control studies. Here, we develop a liability threshold mixed model that extends marginal epistatic mapping to case-control studies. Our method properly accounts for case-control ascertainment and the binary nature of case-control data. We refer to this method as the liability threshold marginal epistasis test (LT-MAPIT). With simulations, we illustrate the benefits of LT-MAPIT in terms of providing effective type I error control, and being more powerful than both existing marginal epistatic mapping methods and conventional explicit search-based approaches in case-control data. We finally apply LT-MAPIT to identify both marginal and pairwise epistasis in seven complex diseases from the Wellcome Trust Case Control Consortium (WTCCC) 1 study.

Drosophila insulin-like peptide dilp1 increases lifespan and glucagon-like Akh expression epistatic to dilp2

Post, S. — 2018-07-30

Insulin/IGF signaling (IIS) regulates essential processes including development, metabolism, and aging. The Drosophila genome encodes eight insulin/IGF-like peptide (dilp) paralogs, including tandem-encoded dilp1 and dilp2. Many reports show that longevity is increased by manipulations that decrease DILP2 in adults. In contrast, dilp1 is expressed primarily in pupal stages, but also during adult reproductive diapause, although we find that dilp1 is also highly expressed in adult dilp2 mutants under non-diapause conditions. The inverse expression of dilp1 and dilp2 suggests these genes interact to regulate aging. Here, we study dilp1 and dilp2 single and double mutants to describe epistatic and synergistic interactions affecting longevity, metabolism and adipokinetic hormone (AKH), a functional homolog of glucagon. Mutants of dilp2 extend lifespan and increase Akh mRNA and protein in a dilp1-dependent manner. Loss of dilp1 alone has no impact on these traits, whereas transgene expression of dilp1 increases lifespan in dilp1-dilp2 double mutants. On the other hand, dilp1 and dilp2 redundantly interact to control circulating sugar, starvation resistance and compensatory dilp5 expression. These later interactions do not correlate with patterns for how dilp1 and dilp2 affect longevity and AKH. Thus, repression or loss of dilp2 slows aging because its depletion induces dilp1, which acts as a pro-longevity factor. Likewise, dilp2 regulates Akh through epistatic interaction with dilp1. Akh and glycogen affect aging in C. elegans and Drosophila, suggesting that dilp2 modulates lifespan via dilp1 and in part by regulating Akh. Whether DILP1 acts as an insulin receptor agonist or inhibitor remains to be resolved.

Molecular Diversity of Intrinsically Photosensitive Ganglion Cells

Berg, D. — 2018-07-31

Intrinsically photosensitive retinal ganglion cells (ipRGCs) are rare mammalian photoreceptors essential for non-image-forming vision functions, such as circadian photoentrainment and the pupillary light reflex. They comprise multiple subtypes distinguishable by morphology, physiology, projections, and levels of expression of melanopsin (Opn4), their photopigment. The molecular programs that differentiate ipRGCs from other ganglion cells and ipRGC subtypes from one another remain elusive. Here, we present comprehensive gene expression profiles of early postnatal and adult mouse ipRGCs purified from two lines of reporter mice marking different sets of ipRGC subtypes. We find dozens of novel genes highly enriched in ipRGCs. We reveal that Rasgrp1 and Tbx20 are selectively expressed in subsets of ipRGCs, though these molecularly defined groups imperfectly match established ipRGC subtypes. We demonstrate that the ipRGCs regulating circadian photoentrainment are unexpectedly diverse at the molecular level. Our findings reveal unexpected complexity in gene expression patterns across mammalian ipRGC subtypes.

Impaired expected value computations in schizophrenia are associated with a reduced ability to integrate reward probability and magnitude of recent outcomes.

Hernaus, D. — 2018-08-10

ABSTRACTO_ST_ABSBackgroundC_ST_ABSMotivational deficits in people with schizophrenia (PSZ) are associated with an inability to integrate the magnitude and probability of previous outcomes. The mechanisms that underlie probability-magnitude integration deficits, however, are poorly understood. We hypothesized that increased reliance on "value-less" stimulus-response associations, in lieu of expected value (EV)-based learning, could drive probability-magnitude integration deficits in PSZ with motivational deficits.nnMethodsHealthy volunteers (n= 38) and PSZ (n=49) completed a reinforcement learning paradigm consisting of four stimulus pairs. Reward magnitude (3/2/1/0 points) and probability (90%/80%/20%/10%) together determined each stimulus EV. Following a learning phase, new and familiar stimulus pairings were presented. Participants were asked to select stimuli with the highest reward value.nnResultsPSZ with high motivational deficits made increasingly less optimal choices as the difference in reward value (probability*magnitude) between two competing stimuli increased. Using a previously-validated computational hybrid model, PSZ relied less on EV ("Q-learning") and more on stimulus-response learning ("actor-critic"), which correlated with SANS motivational deficit severity. PSZ specifically failed to represent reward magnitude, consistent with model demonstrations showing that response tendencies in the actor-critic were preferentially driven by reward probability.nnConclusionsProbability-magnitude deficits in PSZ with motivational deficits arise from underutilization of EV in favor of reliance on value-less stimulus-response associations. Consistent with previous work and confirmed by our computational hybrid framework, probability-magnitude integration deficits were driven specifically by a failure to represent reward magnitude. This work reconfirms the importance of decreased Q-learning/increased actor-critic-type learning as an explanatory framework for a range of EV deficits in PSZ.

Hierarchical Bayesian inference for concurrent model fitting and comparison for group studies

Piray, P. — 2018-08-16

Computational modeling plays an important role in modern neuroscience research. Much previous research has relied on statistical methods, separately, to address two problems that are actually interdependent. First, given a particular computational model, Bayesian hierarchical techniques have been used to estimate individual variation in parameters over a population of subjects, leveraging their population-level distributions. Second, candidate models are themselves compared, and individual variation in the expressed model estimated, according to the fits of the models to each subject. The interdependence between these two problems arises because the relevant population for estimating parameters of a model depends on which other subjects express the model. Here, we propose a hierarchical Bayesian inference (HBI) framework for concurrent model comparison, parameter estimation and inference at the population level, combining previous approaches. We show that this framework has important advantages for both parameter estimation and model comparison theoretically and experimentally. The parameters estimated by the HBI show smaller errors compared to other methods. Model comparison by HBI is robust against outliers and is not biased towards overly simplistic models. Furthermore, the fully Bayesian approach of HBI enables researchers to quantify uncertainty in group parameter estimates, for each candidate model separately, and to perform statistical tests on parameters of a population.

Fast Estimation of Recombination Rates Using Topological Data Analysis

Humphreys, D. P. — 2018-08-20

Accurate estimation of recombination rates is critical for studying the origins and maintenance of genetic diversity. Because the inference of recombination rates under a full evolutionary model is computationally expensive, an alternative approach using topological data analysis (TDA) has been proposed. Previous TDA methods used information contained solely in the first Betti number ({beta}1)of the cloud of genomes, which relates to the number of loops that can be detected within a genealogy. While these methods are considerably less computationally intensive than current biological model-based methods, these explorations have proven difficult to connect to the theory of the underlying biological process of recombination, and consequently have unpredictable behavior under different perturbations of the data. We introduce a new topological feature with a natural connection to coalescent models, which we call{psi} . We show that{psi} and {beta}1 are differentially affected by changes to the structure of the data and use them in conjunction to provide a robust, efficient, and accurate estimator of recombination rates, TREE. Compared to previous TDA methods, TREE more closely approximates of the results of commonly used model-based methods. These characteristics make TREE well suited as a first-pass estimator of recombination rate heterogeneity or hotspots throughout the genome. In addition, we present novel arguments relating {beta}1 to population genetic models; our work justifies the use of topological statistics as summaries of distributions of genome sequences and describes a new, unintuitive relationship between topological summaries of distance and the footprint of recombination on genome sequences.

Dissecting differential signals in high-throughput data from complex tissues

Li, Z. — 2018-08-29

Samples from clinical practices are often mixtures of different cell types. The high-throughput data obtained from these samples are thus mixed signals. The cell mixture brings complications to data analysis, and will lead to biased results if not properly accounted for. We develop a method to model the high-throughput data from mixed, heterogeneous samples, and to detect differential signals. Our method allows flexible statistical inference for detecting a variety of cell-type specific changes. Extensive simulation studies and analyses of two real datasets demonstrate the favorable performance of our proposed method compared with existing ones serving similar purpose.

Genome-wide association studies in Samoans give insight into the genetic etiology of fasting serum lipid levels.

Carlson, J. — 2018-09-08

The current understanding of the genetic architecture of lipids has largely come from genome-wide association studies. To date, few studies have examined the genetic architecture of lipids in Polynesians, and none have in Samoans, whose unique population history, including many population bottlenecks, may provide insight into the biological foundations of variation in lipid levels. Here we performed a genome-wide association study of four fasting serum lipid levels: total cholesterol (TC), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and triglycerides (TG) in a sample of 2,849 Samoans, with validation genotyping for associations in a replication cohort comprising 1,798 Samoans and American Samoans. We identified multiple genome-wide significant associations (P < 5 x 10-8) previously seen in other populations - APOA1 with TG, CETP with HDL, and APOE with TC and LDL - and several suggestive associations (P < 1 x 10-5), including an association of variants downstream of MGAT1 and RAB21 with HDL. However, we observed different association signals for variants near APOE than what has been previously reported in non-Polynesian populations. The association with several known lipid loci combined with the newly-identified associations with variants near MGAT1 and RAB21 suggest that while some of the genetic architecture of lipids is shared between Samoans and other populations, part of the genetic architecture may be Polynesian-specific.

Multiple Introductions of the Mycobacterium tuberculosis Lineage 2 Beijing into Africa over centuries

Rutaihwa, L. K. — 2018-09-10

The Lineage 2-Beijing (L2-Beijing) sub-lineage of Mycobacterium tuberculosis has received much attention due to its high virulence, fast disease progression, and association with antibiotic resistance. Despite several reports of the recent emergence of L2-Beijing in Africa, no study has investigated the evolutionary history of this sub-lineage on the continent. In this study, we used whole genome sequences of 817 L2 clinical strains from 14 geographical regions globally distributed to investigate the origins and onward spread of this lineage in Africa. Our results reveal multiple introductions of L2-Beijing into Africa linked to independent bacterial populations from East-and Southeast Asia. Bayesian analyses further indicate that these introductions occurred during the past 300 years, with most of these events pre-dating the antibiotic era. Hence, the success of L2-Beijing in Africa is most likely due to its hypervirulence and high transmissibility rather than drug resistance.

CLAMP directly interacts with MSL2 to facilitate Drosophila dosage compensation

Tikhonova, E. — 2018-09-12

The binding of Drosophila male-specific lethal (MSL) dosage compensation complex exclusively to male X chromosome provides an excellent model system to understand mechanisms of selective recruitment of protein complexes to chromatin. Previous studies showed that the male-specific organizer of the complex, MSL2, and ubiquitous DNA-binding protein CLAMP are key players in the specificity of X chromosome binding. The CXC domain of MSL2 binds to genomic sites of MSL complex recruitment. Here we demonstrated that MSL2 directly interacts with the N-terminal zinc-finger domain of CLAMP. CLAMP-MSL2 and CXC-DNA interactions are cooperatively involved in recruitment of MSL complex to the X chromosome.

Over-expression of Hsp83 in grossly depleted hsrω lncRNA background causes synthetic lethality and l(2)gl phenocopy in Drosophila

Ray, M. — 2018-09-18

We examined interactions between Hsp83 and hsr{omega} lncRNAs in hsr{omega}66 Hsp90GFP homozygotes, which almost completely lack hsr{omega} lncRNAs but over-express Hsp83. All +/+; hsr{omega}66 Hsp90GFP progeny died before third instar. Rare Sp/CyO; hsr{omega}66 Hsp90GFP reached third instar stage but phenocopied l(2)gl mutants, dying after prolonged larval life, becoming progressively bulbous and transparent with enlarged brain. Additionally, ventral ganglia were elongated. However, hsr{omega}66 Hsp90GFP/TM6B heterozygotes, carrying +/+ or Sp/CyO second chromosomes, developed normally. Total RNA sequencing (+/+, +/+; hsr{omega}66/hsr{omega}66, Sp/CyO; hsr{omega}66/hsr{omega}66, +/+; Hsp90GFP/Hsp90GFP, and Sp/CyO; hsr{omega}66 Hsp90GFP/hsr{omega}66 Hsp90GFP late third instar larvae) revealed similar effects on many genes in hsr{omega}66 and Hsp90GFP homozygotes. Besides additive effect on many of them, numerous additional genes were affected in Sp/CyO; hsr{omega}66 Hsp90GFP larvae, with l(2)gl and several genes regulating CNS being highly down-regulated in surviving Sp/CyO; hsr{omega}66 Hsp90GFP larvae, but not in hsr{omega}66 or Hsp90GFP single mutants. Hsp83 binds at these gene promoters. Several omega speckle associated hnRNPs too may bind with these genes and transcripts. Hsp83-hnRNP interactions are also known. Thus, elevated Hsp83 in altered hnRNP distribution and dynamics, following absence of hsr{omega} lncRNAs and omega speckles, background can severely perturb regulatory circuits with unexpected consequences, including down-regulation of tumor suppressor gene like l(2)gl.

Drosophila insulin-like peptide 1 (DILP1) promotes organismal growth during non-feeding stages

Liao, S. — 2018-09-19

The insulin/IGF-signaling pathway is central in control of nutrient-dependent growth during development, and in adult physiology and longevity. Eight insulin-like peptides (DILP1-8) have been identified in Drosophila and several of these are known to regulate growth, metabolism, reproduction, stress responses and lifespan. However, the functional role of DILP1 is far from understood. Previous work has shown that dilp1/DILP1 is transiently expressed mainly during the non-feeding pupal stage and the first days of adult life. Here we show that mutation of dilp1 diminishes organismal weight during pupal development, whereas overexpression increases it, similar to dilp6 manipulations. No growth effects of dilp1 or dilp6 manipulations were detected during larval development. We next show that dilp1 and dilp6 increase metabolic rate in the late pupa and promote lipids as the primary source of catabolic energy. This lipid mobilization in the pupa is not correlated with transcriptional changes of adipokinetic hormone. The effects of dilp1 manipulations carry over to the adult fly. In newly eclosed flies, survival during starvation is strongly diminished in dilp1 mutants, but not in dilp2 and dilp1-dilp2 double mutants, whereas in older flies only double mutants display reduced starvation resistance. In conclusion, dilp1 and dilp6 promote growth of adult tissues during the non-feeding pupal stage, likely by utilization of stored lipids. This results in larger newly-eclosed flies with reduced stores of pupal-derived nutrients and diminished starvation tolerance and fecundity.

Detection of arterial wall abnormalities via Bayesian model selection

Larson, K. — 2018-09-21

Patient-specific modeling of hemodynamics in arterial networks has so far relied on parameter estimation for inexpensive or small-scale models. We describe here a Bayesian uncertainty quantification framework which makes two major advances: an efficient parallel implementation, allowing parameter estimation for more complex forward models, and a system for practical model selection, allowing evidence-based comparison between distinct physical models. We demonstrate the proposed methodology by generating simulated noisy flow velocity data from a branching arterial tree model in which a structural defect is introduced at an unknown location; our approach is shown to accurately locate the abnormality and estimate its physical properties even in the presence of significant observational and systemic error. As the method readily admits real data, it shows great potential in patient-specific parameter fitting for hemodynamical flow models.

Covariate Assisted Principal Regression for Covariance Matrix Outcomes

Zhao, Y. — 2018-09-23

Modeling variances in data has been an important topic in many fields, including in financial and neuroimaging analysis. We consider the problem of regressing covariance matrices on a vector covariates, collected from each observational unit. The main aim is to uncover the variation in the covariance matrices across units that are explained by the covariates. This paper introduces Covariate Assisted Principal (CAP) regression, an optimization-based method for identifying the components predicted by (generalized) linear models of the covariates. We develop computationally efficient algorithms to jointly search the projection directions and regression coefficients, and we establish the asymptotic properties. Using extensive simulation studies, our method shows higher accuracy and robustness in coefficient estimation than competing methods. Applied to a resting-state functional magnetic resonance imaging study, our approach identifies the human brain network changes associated with age and sex.

Dynamics of contrast decrement and increment responses in human visual cortex

Norcia, A. M. — 2018-09-24

The goal of the present experiments was to determine whether electrophysiological response properties of the ON and OFF visual pathways observed in animal experimental models can be observed in human. Visual Evoked Potentials (VEPs) were recorded in response to equivalent magnitude contrast increments and decrements presented using sawtooth temporal waveforms at a temporal frequency of 2.73 Hz. VEP response waveforms and response spectra for incremental and decremental stimuli were analyzed as a function of stimulus size and visual field location in 68 healthy adult participants. VEP response were larger in amplitude and shorter in latency for contrast decrements than for contrast increments. The spatial tuning was narrower for contrast decrements than for contrast increments and responses were larger for displays that were scaled for cortical magnification. VEPs recorded at the scalp differ between contrast decrements and increments of equal Weber contrast in a fashion that parallels results from the early visual system of cats and monkeys. Because our assay allows differential detection of ON and OFF pathway activity in human, the approach may be useful in future work on disease detection and treatment monitoring.

Genotypic context modulates fitness landscapes: Effects onthe speed and direction of evolution for antimicrobialresistance

Ogbunugafor, C. B. — 2018-09-25

Understanding the forces that drive the dynamics of adaptive evolution is a goal of many subfields within evolutionary biology. The fitness landscape analogy has served as a useful abstraction for addressing these topics across many systems, and recent treatments have revealed how different environments can frame the particulars of adaptive evolution by changing the topography of fitness landscapes. In this study, we examine how the larger, ambient genotypic context in which the fitness landscape being modeled is embedded affects fitness landscape topography and subsequent evolution. Using simulations on empirical fitness landscapes, we discover that genotypic context, defined by genetic variability in regions outside of the locus under study (in this case, an essential bacterial enzyme target of antibiotics), influences the speed and direction of evolution in several surprising ways. These findings have implications for how we study the evolution of drug resistance in nature, and for presumptions about how biological evolution might be expected to occur in genetically-modified organisms. More generally, the findings speak to theory surrounding how "difference can beget difference" in adaptive evolution: that small genetic differences between organisms can greatly alter the specifics of how evolution occurs, which can rapidly drive even slightly diverged populations further apart.nnAuthor summaryTechnological advances enable scientists to engineer individual mutations at specific sites within an organisms genome with increasing ease. These breakthroughs have provided scientists with tools to study how different engineered mutations affect the function of a given gene or protein, yielding useful insight into genotype-phenotype mapping and evolution. In this study, we use engineered strains of bacteria to show how the dynamics (speed and direction) of evolution of drug resistance in an enzyme depends on the species-type of that bacterial enzyme, and on the presence/absence of mutations in other genes in the bacterial genome. These findings have broad implications for public health, genetic engineering, and theories of speciation. In the context of public health and biomedicine, our results suggest that future efforts in managing antimicrobial resistance must consider genetic makeup of different pathogen populations before predicting how resistance will occur, rather than assuming that the same resistance pathways will appear in different pathogen populations. With regard to broader theory in evolutionary biology, our results show how even small genetic differences between organisms can alter how future evolution occurs, potentially causing closely-related populations to quickly diverge.

Trained-feature specific offline learning in an orientation detection task

Tamaki, M. — 2018-09-28

It has been suggested that sleep provides additional enhancement of visual perceptual learning (VPL) acquired before sleep, termed offline performance gains. A majority of the studies that found offline performance gains of VPL used discrimination tasks including the texture discrimination task (TDT). This makes it questionable whether offline performance gains on VPL are generalized to other visual tasks. The present study examined whether a Gabor orientation detection task, which is a standard task in VPL, shows offline performance gains. In Experiment 1, we investigated whether sleep leads to offline performance gains on the task. Subjects were trained with the Gabor orientation detection task, and re-tested it after a 12-hr interval that included either nightly sleep or only wakefulness. We found that performance on the task improved to a significantly greater degree after the interval that included sleep and wakefulness than the interval including wakefulness alone. In addition, offline performance gains were specific to the trained orientation. In Experiment 2, we tested whether offline performance gains occur by a nap. Also, we tested whether spontaneous sigma activity in early visual areas during non-rapid eye movement (NREM) sleep, previously implicated in offline performance gains of TDT, was associated with offline performance gains of the task. A different group of subjects had a nap with polysomnography. The subjects were trained with the task before the nap and re-tested after the nap. The performance of the task improved significantly after the nap only on the trained orientation. Sigma activity in the trained region of early visual areas during NREM sleep was significantly larger than in the untrained region, in correlation with offline performance gains. These aspects were also found with VPL of TDT. The results of the present study demonstrate that offline performance gains are not specific to a discrimination task such as TDT, and can be generalized to other forms of VPL tasks, along with trained-feature specificity. Moreover, the present results also suggest that sigma activity in the trained region of early visual areas plays an important role in offline performance gains of VPL of detection as well as discrimination tasks.

A theoretical foundation for state-transition cohort models in health decision analysis

Iskandar, R. — 2018-09-28

Following its introduction over three decades ago, the cohort model has been used extensively to model population trajectories over time in decision-analytic modeling studies. However, the stochastic process underlying cohort models has not been properly described. In this study, we explicate the stochastic process underlying a cohort model, by carefully formulating the dynamics of populations across health states and assigning probability rules on these dynamics. From this formulation, we explicate a mathematical representation of the system, which is given by the master equation. We solve the master equation by using the probability generation function method to obtain the explicit form of the probability of observing a particular realization of the system at an arbitrary time. The resulting generating function is used to derive the analytical expressions for calculating the mean and the variance of the process. Secondly, we represent the cohort model by a difference equation for the number of individuals across all states. From the difference equation, a continuous-time cohort model is recovered and takes the form of an ordinary differential equation. To show the equivalence between the derived stochastic process and the cohort model, we conduct a numerical exercise. We demonstrate that the population trajectories generated from the formulas match those from the cohort model simulation. In summary, the commonly-used cohort model represent the average of a continuous-time stochastic process on a multidimensional integer lattice governed by a master equation. Knowledge of the stochastic process underlying a cohort model provides a theoretical foundation for the modeling method.

Causal evidence for LTP-based interference in visual learning

Bang, J. W. — 2018-10-08

Training related skills in close succession results in interference but the reasons for this interference are not understood. Here we test the hypothesis that interference occurs due to competition of long-term potentiation (LTP): the LTP induced by one task impedes the LTP induced by the other. Human subjects performed two consecutive training sessions on different Gabor orientations. Immediately after the offset of the first training, we applied continuous theta burst stimulation (cTBS) to interfere with the LTP processes produced by the first training. We found that cTBS to a control site (vertex) resulted in substantial anterograde interference for the second training. Critically, cTBS to the visual cortex not only disrupted learning on the immediately preceding training, but also released the second training from the anterograde interference. These results provide strong support for the LTP-based theory of interference and suggest the possibility of directly manipulating the competition between different learning periods.

Modular modeling improves the predictions of genetic variant effects on splicing

Cheng, J. — 2018-10-10

Predicting the effects of genetic variants on splicing is highly relevant for human genetics. We describe the framework MMSplice (modular modeling of splicing) with which we built the winning model of the CAGI 2018 exon skipping prediction challenge. The MMSplice modules are neural networks scoring exon, intron, and splice sites, trained on distinct large-scale genomics datasets. These modules are combined to predict effects of variants on exon skipping, alternative donor and acceptor sites, splicing efficiency, and pathogenicity, with matched or higher performance than state-of-the-art. Our models, available in the repository Kipoi, apply to variants including indels directly from VCF files.

Synaptic circuits for irradiance coding by intrinsically photosensitive retinal ganglion cells

Sabbah, S. — 2018-10-14

We have explored the synaptic networks responsible for the unique capacity of intrinsically photosensitive retinal ganglion cells (ipRGCs) to encode overall light intensity. This luminance signal is crucial for circadian, pupillary and related reflexive responses light. By combined glutamate-sensor imaging and patch recording of postsynaptic RGCs, we show that the capacity for intensity-encoding is widespread among cone bipolar types, including OFF types.nnNonetheless, the bipolar cells that drive ipRGCs appear to carry the strongest luminance signal. By serial electron microscopic reconstruction, we show that Type 6 ON cone bipolar cells are the dominant source of such input, with more modest input from Types 7, 8 and 9 and virtually none from Types 5i, 5o, 5t or rod bipolar cells. In conventional RGCs, the excitatory drive from bipolar cells is high-pass temporally filtered more than it is in ipRGCs. Amacrine-to-bipolar cell feedback seems to contribute surprisingly little to this filtering, implicating mostly postsynaptic mechanisms. Most ipRGCs sample from all bipolar terminals costratifying with their dendrites, but M1 cells avoid all OFF bipolar input and accept only ectopic ribbon synapses from ON cone bipolar axonal shafts. These are remarkable monad synapses, equipped with as many as a dozen ribbons and only one postsynaptic process.

Human neurons from Christianson syndrome iPSCs reveal allele-specific responses to rescue strategies

Lizarraga, S. B. — 2018-10-16

Human genetic disorders provide a powerful lens to understanding the human brain. Induced pluripotent stem cells (iPSC) represent an important, new resource for mechanistic studies and therapeutic development. Christianson syndrome (CS), an X-linked neurological disorder with attenuation of brain growth postnatally (postnatal microcephaly), is caused by mutations in SLC9A6, the gene encoding endosomal Na+/H+ exchanger 6 (NHE6). We developed CS iPSC lines from patients with a mutational spectrum, as well as robust biologically-related and isogenic controls. We demonstrate that mutations in CS lead to loss of protein function by a variety of mechanisms. Regardless of mutation, all patient-derived neurons demonstrate reduced neurite growth and arborization, likely underlying diminished postnatal brain growth in patients. Additionally, phenotype rescue strategies show allele-specific responses: a gene replacement strategy shows efficacy in nonsense mutations but not in a missense mutation, whereas application of exogenous trophic factors (BDNF or IGF-1) rescues arborization phenotypes across all mutations. Our data emphasize the important principle of personalized medicine whereby success of some therapeutic strategies may be more linked to patient genotype than others.

Differential Effects of the Menstrual Cycle on Reactive and Proactive Aggression in Borderline Personality Disorder

Peters, J. R. — 2018-10-29

Borderline personality disorder (BPD) is characterized by rapidly shifting symptoms, including intense anger and aggressive behavior. Understanding how fluctuations in ovarian hormones across the menstrual cycle may contribute to symptom instability is key for accurate assessment of BPD symptoms and effective interventions. Reactive and proactive aggression, as well as anger in and out, were assessed daily in 15 physically healthy, unmedicated naturally cycling female individuals without dysmenorrhea meeting criteria for BPD across 35 days. Urine LH surge and salivary progesterone were used to confirm ovulation and verify cycle phase. Cyclical worsening of symptoms was evaluated using multilevel models to evaluate symptom differences between cycle phases. Both forms of aggressive behavior demonstrated marked cycle effects, with reactive aggression highest during perimenstrual cycle phases, co-occurring with increases in anger in and out. In contrast, highest levels of proactive aggression were observed during the follicular and ovulatory phases, when emotional symptoms and anger were otherwise at lowest levels. These findings highlight the importance of identifying the function of aggression when considering potential psychological and biological influences. Naturally cycling individuals with BPD may be at elevated risk for perimenstrual worsening of a range of interpersonally reactive symptoms, including reactive aggression, whereas proactive aggression may occur more in phases characterized by less emotional and cognitive vulnerability and greater reward sensitivity. Research on aggression in this population should consider cycle effects. Cycling individuals with BPD attempting to reduce aggressive behavior may benefit from cycle-tracking to increase awareness of these effects and to develop appropriate strategies.

Age-dependent changes in transcription factor FOXO targeting in Drosophila melanogaster

Birnbaum, A. — 2018-10-30

FOXO transcription factors have long been associated with longevity control and tissue homeostasis. Although the transcriptional regulation of FOXO have been previously characterized (especially in long-lived insulin mutants and under stress conditions), how normal aging impacts the transcriptional activity of FOXO is poorly understood. Here, we conducted a chromatin immunoprecipitation sequencing (ChIP-Seq) analysis in both young and old wild-type fruit flies, Drosophila melanogaster, to evaluate the dynamics of FOXO gene targeting during aging. Intriguingly, the number of FOXO-bound genes dramatically decreases with age (from 2617 to 224). Consistent to the reduction of FOXO binding activity, many genes targeted by FOXO in young flies are transcriptionally altered with age, either up-regulated (FOXO-repressing genes) or down-regulated (FOXO-activating genes). In addition, we show that many FOXO-bound genes in wild-type flies are unique from those in insulin receptor substrate chico mutants. Distinct from chico mutants, FOXO targets specific cellular processes (e.g., actin cytoskeleton) and signaling pathways (e.g., Hippo, MAPK) in young wild-type flies. FOXO targeting on these pathways decreases with age. Interestingly, FOXO targets in old flies are enriched in cellular processes like chromatin organization and nucleosome assembly. Furthermore, FOXO binding to core histone genes is well maintained at aged flies. Together, our findings provide new insights into dynamic FOXO targeting under normal aging and highlight the diverse and understudied regulatory mechanisms for FOXO transcriptional activity.

Inhibition of retrotransposition improves health and extends lifespan of SIRT6 knockout mice

Simon, M. — 2018-11-04

Mice deficient for SIRT6 exhibit a severely shortened lifespan, growth retardation, and highly elevated LINE1 (L1) activity. Here we report that SIRT6 deficient cells and tissues accumulate abundant cytoplasmic L1 cDNA which triggers massive type I interferon response via activation of cGAS. Remarkably, nucleoside reverse transcriptase inhibitors (NRTIs), which inhibit L1 retrotransposition, significantly improved health and lifespan of SIRT6 knockout mice and completely rescued type I interferon response. In tissue culture, inhibition of L1 with siRNA or NRTIs abrogated type I interferon response, in addition to a significant reduction of DNA damage markers. These results indicate that L1 activation contributes to the pathologies of SIRT6 knockout mice. Similarly, L1 transcription, cytoplasmic cDNA copy number and type I interferons were elevated in the wild type aged mice. As sterile inflammation is a hallmark of aging we propose that modulating L1 activity may be an important strategy for attenuating age-related pathologies.nnHighlightsO_LISIRT6 KO mice accumulate L1 cDNA triggering type I interferon response via cGAS pathwaynC_LIO_LIWild type aged mice accumulate L1 cDNA and display type I interferon responsenC_LIO_LIReverse transcriptase inhibitors rescue type I interferon response and DNA damagenC_LIO_LIReverse transcriptase inhibitors extend lifespan and improve health of SIRT6 KO micenC_LI

SIMNETS: a computationally efficient and scalable framework for identifying networks of functionally similar neurons

Hynes, J. B. — 2018-11-08

The expansion of large-scale neural recording capabilities has provided new opportunities to examine multi-scale cortical network activity at single neuron resolution. At the same time, the growing scale and complexity of these datasets introduce new conceptual and technical challenges beyond what can be addressed using traditional analysis techniques. Here, we present SIMNETS, a mathematically rigorous and efficient unsupervised relational analysis framework designed to generate intuitive, low-dimensional neuron maps that support a multi-scale view of the computational similarity (CS) relations among individual neurons. The critical innovation is the use of a novel measure of computational similarity that is based on comparing the intrinsic structure of latent spaces representing the spiking output of individual neurons. We use three publicly available neural population test datasets from the visual, motor, and hippocampal CA1 brain regions to validate the SIMNETS framework and demonstrate how it can be used to identify putative subnetworks (i.e., clusters of neurons with similar computational properties). Our analysis pipeline includes a novel statistical test designed to evaluate the likelihood of detecting spurious neuron clusters to validate network structure results. The SIMNETS framework can facilitate linking computational geometry representations across scales, from single neurons to subnetworks, within large-scale neural recording data.

Proteostasis environment shapes higher-order epistasis operating on antibiotic resistance

Guerrero, R. F. — 2018-11-16

Recent studies have shown that higher-order epistasis is ubiquitous and can have large effects on complex traits. Yet, we lack frameworks for understanding how epistatic interactions are influenced by basic aspects of cell physiology. In this study, we assess how protein quality control machinery--a critical component of cell physiology--affects epistasis for different traits related to bacterial resistance to antibiotics. Specifically, we attempt to disentangle the interactions between different protein quality control genetic backgrounds and two sets of mutations: (i) SNPs associated with resistance to antibiotics in an essential bacterial enzyme (dihydrofolate reductase, or DHFR) and (ii) differing DHFR bacterial species-specific amino acid background sequences (Escherichia coli, Listeria grayi, and Chlamydia muridarum). In doing so, we add nuance to the generic observation that non-linear genetic interactions are widespread and capricious in nature, by proposing a mechanistically-grounded analysis of how proteostasis shapes epistasis. These findings simultaneously fortify and demystify the role of environmental context in modulating higher-order epistasis, with direct implications for evolutionary theory, genetic modification technology, and efforts to manage antimicrobial resistance.

DNA methylation modules associate with incident cardiovascular disease and cumulative risk factor exposure

Westerman, K. — 2018-11-19

1 AbstractO_ST_ABSBackgroundC_ST_ABSEpigenome-wide association studies using DNA methylation have the potential to uncover novel biomarkers and mechanisms of cardiovascular disease (CVD) risk. However, the direction of causation for these associations is not always clear, and investigations to-date have generally failed to replicate at the level of individual loci. Here, we undertook module- and region-based DNA methylation analyses of incident CVD in the Womens Health Initiative (WHI) and Framingham Heart Study Offspring Cohort (FHS) in order to find more robust epigenetic biomarkers for cardiovascular risk.nnMethods and FindingsWe applied weighted gene correlation network analysis (WGCNA) and the Comb-p algorithm to find methylation modules and regions associated with incident CVD in the WHI dataset. We discovered two modules whose activation correlated with CVD risk and replicated across cohorts. One of these modules was enriched for development-related processes and overlaps strongly with epigenetic aging sites. For the other, we showed preliminary evidence for monocyte-specific effects and statistical links to cumulative exposure to traditional cardiovascular risk factors. Additionally, we found three regions (associated with the genes SLC9A1, SLC1A5, and TNRC6C) whose methylation associates with CVD risk.nnConclusionsIn sum, we present several epigenetic associations with incident CVD that reveal disease mechanisms related to development and monocyte biology. Furthermore, we show that epigenetic modules may act as a molecular readout of cumulative cardiovascular risk factor exposure, with implications for the improvement of clinical risk prediction.

Metabolism-Induced Oxidative Stress and DNA Damage Selectively Trigger Genome Instability in Polyploid Cells

Thomson, G. J. — 2018-11-29

Understanding the forces impacting genome stability is important for diverse processes such as tumorigenesis and reproductive biology. The pathogenic yeast Candida albicans displays unusual genome dynamics in which tetraploid cells, but not diploid cells, become unstable when grown on a glucose-rich pre-sporulation medium. Here, we reveal that C. albicans polyploid cells are metabolically hyperactive on this medium as evidenced by increased expression of metabolic genes as well as higher rates of fermentation and oxidative respiration. These cells also show elevated levels of reactive oxygen species (ROS), activate the ROS-responsive transcription factor Cap1, and accrue DNA double-strand breaks. Suppression of ROS levels reduced oxidative stress, DNA damage and chromosome instability. These studies reveal how metabolic flux can generate endogenous ROS, triggering DNA damage and genome instability in polyploid, but not diploid, cells. We discuss parallels with metabolism-induced instability in cancer cells and propose that ROS-induced DNA damage could have facilitated ploidy cycling in eukaryotes prior to the evolution of meiosis.

Vitamin D status and risk of incident tuberculosis disease: a systematic review and individual participant data meta-analysis

Aibana, O. — 2018-11-30

BackgroundFew studies have evaluated the association between pre-existing vitamin D deficiency (VDD) and incident TB. We assessed the impact of baseline vitamin D on TB risk. MethodsWe assessed the association between baseline vitamin D and incident TB in a prospective cohort of 6751 household contacts of TB patients in Peru. We also conducted a one-stage individual participant data meta-analysis searching PubMed and Embase for studies of vitamin D and TB until December 31, 2017. We included studies that assessed vitamin D before TB diagnosis. We defined VDD as 25-(OH)D <50 nmol/L, insufficiency as 50-75 nmol/L and sufficiency as >75nmol/L. We estimated the association between vitamin D and incident TB using conditional logistic regression in the Peru cohort and generalized linear mixed models in the meta-analysis. FindingsIn Peru, baseline VDD was associated with a statistically insignificant increase in incident TB (aOR 1{middle dot}70, 95% CI 0{middle dot}84-3{middle dot}46; p=0{middle dot}14). We identified seven studies for the meta-analysis and analyzed 3544 participants. Individuals with VDD and very low vitamin D (<25nmol/L) had increased TB risk (aOR 1{middle dot}48, 95% CI 1{middle dot}04-210;p=0{middle dot} 03 and aOR 2 08, 95% CI 0{middle dot}88-4{middle dot}92; p trend=002 respectively). Among HIV-positive patients, VDD and very low vitamin D conferred a 2-fold (aOR 2.18, 95% CI 1{middle dot} 22-3{middle dot}90; p=0{middle dot} 01) and 4-fold (aOR 4{middle dot}28, 95% CI 0{middle dot}85-21{middle dot}44; p trend=0{middle dot}01) increased risk of TB respectively. InterpretationOur findings suggest vitamin D predicts TB risk in a dose-dependent manner and vitamin D supplementation may play a role in TB prevention. FundingNational Institute of Health (NIH), National Institute of Allergy and Infectious Diseases (NIAID), National Institute on Drug Abuse (NIDA), National Institute of Mental Health (NIMH), International Maternal Pediatric Adolescent AIDS Clinical Trials Group (IMPAACT), Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institute of Dental and Craniofacial Research (NIDCR), Boehringer-Ingelheim, Bristol-Myers Squibb, Gilead Sciences, GlaxoSmithKline, Foundation, Ujala Foundation, Wyncote Foundation, NIH - Fogarty International Center Program of International Training Grants in Epidemiology Related to AIDS, NIAID Byramjee Jeejeebhoy Medical College HIV Clinical Trials Unit, NIAIDs Baltimore-Washington-India Clinical Trials Unit, National Commission on Biotechnology, the Higher Education Commission, International Research Support Initiative Program of the Higher Education Commission Government of Pakistan, the Bill and Melinda Gates Foundation, and the NIH Fogarty International Center. Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSNumerous studies have found lower serum vitamin D levels among patients with active TB disease compared to healthy controls. However, research has not clarified whether low vitamin D increases TB risk or whether TB disease leads to decreased vitamin D levels. We conducted PubMed and Medline searches for all studies available through December 31, 2017 on the association between vitamin D status and TB disease. We included the following keywords: "vitamin D," "vitamin D deficiency," "hypovitaminosis D," "25-hydroxyvitamin D," "1,25-dihydroxyvitamin D," "vitamin D2," "vitamin D3," "ergocalciferol," "cholecalciferol," and "tuberculosis." We found only seven studies had prospectively evaluated the impact of baseline vitamin D levels on risk of progression to TB disease. We report here the results of a case control study nested within a large prospective longitudinal cohort study of household contacts of TB cases and the results of an individual participant data (IPD) metaanalysis of available evidence on the association between vitamin D levels and incident TB disease. Added value of this studyWe demonstrated that low vitamin D levels predicts risk of future progression to TB disease in a dose-dependent manner. Implications of all the available evidenceThese findings suggest the possibility that vitamin D supplementation among individuals at high risk for developing TB disease might play a role in TB prevention efforts.

Novel genes for autism implicate both excitatory and inhibitory cell lineages in risk

Satterstrom, F. K. — 2018-11-30

We present the largest exome sequencing study to date focused on rare variation in autism spectrum disorder (ASD) (n=35,584). Integrating de novo and case-control variation with an enhanced Bayesian framework incorporating evolutionary constraint against mutation, we implicate 99 genes in ASD risk at a false discovery rate (FDR) [≤] 0.1. Of these 99 risk genes, 46 show higher frequencies of disruptive de novo variants in individuals ascertained for severe neurodevelopmental delay, while 50 show higher frequencies in individuals ascertained for ASD, and comparing ASD cases with disruptive mutations in the two groups shows differences in phenotypic presentation. Expressed early in brain development, most of the risk genes have roles in neuronal communication or regulation of gene expression, and 12 fall within recurrent copy number variant loci. In human cortex single-cell gene expression data, expression of the 99 risk genes is also enriched in both excitatory and inhibitory neuronal lineages, implying that disruption of these genes alters the development of both neuron types. Together, these insights broaden our understanding of the neurobiology of ASD.

Circuit dysfunction in SOD1-ALS model first detected in sensory feedback prior to motor neuron degeneration is alleviated by BMP signaling

Held, A. — 2018-12-03

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease whose origin and underlying cellular defects are still not fully understood. While motor neuron degeneration is the signature feature of ALS, it is not yet clear if motor neurons, or other cells of the motor circuit, are the site of disease initiation. To better understand the contribution of multiple cell types in ALS, we made use of a Drosophila Sod1G85R knock-in model, in which all cells harbor the disease allele. End-stage dSod1G85R animals exhibit severe motor deficits with clear degeneration of motor neurons. Interestingly, earlier in dSod1G85R larvae motor function is also compromised, but their motor neurons exhibit only subtle morphological and electrophysiological changes, that are unlikely to cause the observed decrease in locomotion. We analyzed the intact motor circuit and identified a defect in sensory feedback that likely accounts for the altered motor activity of dSod1G85R. Furthermore, we found that the cell-autonomous activation of BMP signaling in proprioceptor sensory neurons that relay the contractile status of muscles back to the central nerve cord, is able to completely rescue early stage motor defects and partially rescue late stage motor function to extend lifespan. Identifying a defect in sensory feedback as a potential initiating event in ALS motor dysfunction, coupled with the ability of modified proprioceptors to alleviate such motor deficits, underscores the critical role that non-motor neurons play in disease progression and highlights their potential as a site to identify early-stage ALS biomarkers and for therapeutic intervention.nnSignificance StatementAt diagnosis, many cellular processes are already disrupted in the ALS patient. Identifying the initiating cellular events is critical for achieving an earlier diagnosis in order to slow or prevent disease progression. Our findings indicate that neurons relaying sensory information underlie early stage motor deficits in a Drosophila knock-in model of ALS that best replicates gene dosage in familial ALS (fALS). Importantly, studies on intact motor circuits revealed defects in sensory feedback before evidence of motor neuron degeneration. These findings strengthen our understanding of how neural circuit dysfunctions lead to neurodegeneration and coupled with our demonstration that activation of BMP signaling in proprioceptors alleviates both early and late motor dysfunction, underscores the importance of considering non-motor neurons as therapeutic targets.

Drought frequency predicts life history strategies in Heliophila

Monroe, J. G. — 2018-12-13

Explaining variation in life history strategies is a long-standing goal of evolutionary biology. For plants, annual and perennial life histories are thought to reflect adaptation to environments that differ in the frequency of stress events such as drought. Here we test this hypothesis in Heliophila (Brassicaceae), a diverse genus of flowering plants native to Africa, by integrating 34 years of satellite-based drought measurements with 2192 herbaria occurrence records. Consistent with predictions from classic life history theory, we find that perennial Heliophila species occur in environments where droughts are significantly less frequent compared to annuals. These associations are predictive while controlling for phylogeny, lending support to the hypothesis that drought related natural selection has influenced the distributions of these strategies. Additionally, the collection dates of annual and perennial species indicate that annuals escape drought prone seasons during the seed phase of their life cycle. Together, these findings provide empirical support for classic hypotheses about the drivers of life history strategy in plants - that perennials out compete annuals in environments with less frequent drought and that annuals are adapted to environments with more frequent drought by escaping drought prone seasons as seeds.

Rats exhibit similar biases in foraging and intertemporal choice tasks

Kane, G. A. — 2018-12-17

Animals, including humans, consistently exhibit myopia in two different contexts: foraging, in which they harvest locally beyond what is predicted by optimal foraging theory, and intertemporal choice, in which they exhibit a preference for immediate vs. delayed rewards beyond what is predicted by rational (exponential) discounting. Despite the similarity in behavior between these two contexts, previous efforts to reconcile these observations in terms of a consistent pattern of time preferences have failed. Here, via extensive behavioral testing and quantitative modeling, we show that rats exhibit similar time preferences in both contexts: they prefer immediate vs. delayed rewards and they are sensitive to opportunity costs -- delays to future decisions. Further, a quasi-hyperbolic discounting model, a form of hyperbolic discounting with separate components for short-and long-term rewards, explains individual rats time preferences across both contexts, providing evidence for a common mechanism for myopic behavior in foraging and intertemporal choice.

A counterselectable sucrose sensitivity marker permits efficient and flexible mutagenesis in Streptococcus agalactiae

Hooven, T. A. — 2018-12-19

Streptococcus agalactiae (group B Streptococcus; GBS) is a cause of severe infections, particularly during the newborn period. While methods exist for generating chromosomal mutations in GBS, they are cumbersome and inefficient and present significant challenges if the goal is to study subtle mutations such as single base pair polymorphisms. To address this problem, we have developed an efficient and flexible GBS mutagenesis protocol based on sucrose counterselection against levansucrase (SacB) expressed from a temperature-selective shuttle vector. GBS containing the SacB expression cassette demonstrate lethal sensitivity to supplemental sucrose whether the plasmid DNA is replicating outside of the chromosome or has been integrated during a crossover event. Transmission electron microscopy shows that SacB-mediated lethal sucrose sensitivity results from accumulation of inclusion bodies that eventually lead to complete degradation of normal cellular architecture and subsequent lysis. We used this new mutagenesis technique to generate an in-frame, allelic exchange knockout of the GBS sortase gene srtA, demonstrating that >99% of colonies that emerge from our protocol had the expected knockout phenotype and that among a subset tested by sequencing, 100% had the correct genotype. We also generated barcoded nonsense mutations in the cylE gene in two GBS strains, showing that the approach can be used to make small, precise chromosomal mutations.nnImportanceThe ability to generate chromosomal mutations is fundamental to microbiology. Historically, however, GBS pathogenesis research has been made challenging by the relative genetic intractability of the organism. Generating a single knockout in GBS using traditional techniques can take many months, with highly variable success rates. Furthermore, traditional methods do not offer a straightforward way to generate single base pair polymorphisms or other subtle changes, especially to noncoding regions of the chromosome. We have developed a new sucrose counterselection-based method that permits rapid, efficient, and flexible GBS mutagenesis. Our technique requires no additional equipment beyond what is needed for traditional approaches. We believe that it will catalyze rapid advances in GBS genetics research by significantly easing the path to generating mutants.

Motivation and Cognitive Control in Depression

Grahek, I. — 2018-12-19

Depression is linked to deficits in cognitive control and a host of other cognitive impairments arise as a consequence of these deficits. Despite of their important role in depression, there are no mechanistic models of cognitive control deficits in depression. In this paper we propose how these deficits can emerge from the interaction between motivational and cognitive processes. We review depression-related impairments in key components of motivation along with new cognitive neuroscience models that focus on the role of motivation in the decision-making about cognitive control allocation. Based on this review we propose a unifying framework which connects motivational and cognitive impairments in depression. This framework is rooted in computational models of cognitive control and offers a mechanistic understanding of cognitive control deficits in depression.

The FXR2P low complexity domain drives assembly of multiple fibril types with differing ribosome association in neurons

Stackpole, E. E. — 2018-12-22

RNA binding proteins (RBPs) typically function in higher order assemblages to regulate RNA localization and translation. The Fragile X homolog FXR2P is an RBP essential for formation of Fragile X granules, which associate with axonal mRNA and ribosomes in the intact brain. Here we performed an unbiased EGFP insertional mutagenesis screen to probe for FXR2P domains important for assembly into higher order structural states in neurons. Fifteen of the 18 unique in-frame FXR2PEGFP fusions tested formed cytosolic granules. However, EGFP insertion within a 23 amino acid region of the low complexity (LC) domain induced formation of distinct FXR2PEGFP fibrils (A and B) that were found in isolation or assembled into highly ordered bundles. Type A and B complexes exhibited different developmental timelines, ultrastructure and ribosome association with ribosomes absent from bundled Type B fibrils. The formation of both fibril types was dependent on an intact RNA binding domain. We conclude that formation of these higher order FXR2P assemblages with alternative structural and compositional states in neurons requires collaboration between the LC and RNA binding domains.nnSummary StatementThe Fragile X protein FXR2P forms multiple types of fibrillar assemblages with differential ribosome associations in neurons through cooperation between its RNA binding and LC domains.

Neural ensemble dynamics in dorsal motor cortex during speech in people with paralysis

Stavisky, S. D. — 2018-12-30

Speaking is a sensorimotor behavior whose neural basis is difficult to study at the resolution of single neurons due to the scarcity of human intracortical measurements and the lack of animal models. We recorded from electrode arrays in the hand knob area of motor cortex in people with tetraplegia. Neurons in this area, which have not previously been implicated in speech, modulated during speaking and during non-speaking movement of the tongue, lips, and jaw. This challenges whether the conventional model of a motor homunculus division by major body regions extends to the single-neuron scale. Spoken words and syllables could be decoded from single trials, demonstrating the potential utility of intracortical recordings for brain-computer interfaces (BCIs) to restore speech. Two neural population dynamics features previously reported for arm movements were also present during speaking: a large initial condition-invariant signal, followed by rotatory dynamics. This suggests that common neural dynamical motifs may underlie movement of arm and speech articulators.

Ecological modeling of Long Interspersed elements reveals footprints of evolution and a role of chromatin in shaping their genome landscape in mammals.

Vitali, S. — 2019-01-07

Transposons are genome components that account for the majority of genome size in many organisms, behaving as parasitic entities and interfering with the translation mechanism. Chromatin structure influences the activity of transposons, by coordinating genome accessibility for the expression and insertion of these sequences. As a case study, we show evidences of an evolutionary response of the chromatin structure to a variation in the activity of Long Interspersed Elements (LINEs) during mammals evolution, with focus on the murine radiation and primate evolution. LINEs activity was measured using a biophysical approach for modeling LINEs as an ecosystem, where different strains of transposons might reproduce, die and compete for access to the translational machinery of the host. The model, based on the discrete stochastic processes of amplification and deactivation of LINEs copies, has been adapted to the data using Bayesian statistics to estimate its main parameters: rate of growth of transposons copy number and rate of past competition between transposons variants. This approach allows to estimate the activity of ancient LINE strains still present in the genome as deactivated components, and the possible competition among different strains. We leverage these results to highlight how the change in the chromatin structure of the murine species seems to be following an increase of LINEs activity during the appearance of the murine specific strain Lx. On the contrary, a similar response is absent in primates evolution, which follows a decrease of LINEs activity during the amplication of primate specific LIMA/LPB strains.

Hsc70 Ameliorates the Vesicle Recycling Defects Caused by Excess -Synuclein at Synapses

Banks, S. M. L. — 2019-01-10

-Synuclein overexpression and aggregation are linked to Parkinsons disease (PD), dementia with Lewy bodies (DLB), and several other neurodegenerative disorders. In addition to effects in the cell body, -synuclein accumulation occurs at presynapses where the protein is normally localized. While it is generally agreed that excess -synuclein impairs synaptic vesicle trafficking, the underlying mechanisms are unknown. We show here that acute introduction of excess human -synuclein at a classic vertebrate synapse, the lamprey reticulospinal synapse, selectively impaired the uncoating of clathrin-coated vesicles (CCVs) during synaptic vesicle recycling, leading to a severe depletion of synaptic vesicles. Furthermore, human -synuclein and lamprey {gamma}-synuclein both interact in vitro with Hsc70, the chaperone protein that uncoats CCVs at synapses. After introducing excess -synuclein to lamprey axons, Hsc70 availability was reduced at the synapses, suggesting Hsc70 sequestration as a possible mechanism underlying the synaptic vesicle trafficking defects. In support of this hypothesis, increasing the levels of exogenous Hsc70 together with -synuclein ameliorated the CCV uncoating and vesicle recycling defects. These experiments identify a reduction in Hsc70 availability at synapses, and consequently its function, as the mechanism by which -synuclein induces synaptic vesicle recycling defects. To our knowledge, this is the first report of a viable chaperone-based strategy for reversing the toxic impacts of excess -synuclein at synapses, which may be of value for ameliorating synaptic defects in PD and other synuclein-linked diseases.nnSIGNIFICANCE STATEMENTSynaptic defects caused by -synuclein overexpression are linked to cognitive deficits in PD and other diseases. However, the mechanisms by which excess -synuclein impairs synaptic vesicle trafficking are unknown. Data presented here demonstrate that acute introduction of excess -synuclein at a classical vertebrate synapse selectively inhibits CCV uncoating, leading to impaired vesicle recycling. Furthermore, increasing -synuclein reduced synaptic levels of Hsc70, the clathrin uncoating ATPase. Subsequently increasing Hsc70 restored CCV uncoating and improved vesicle recycling. This study identifies a novel molecular mechanism underlying the -synuclein-induced synaptic defects and presents one viable strategy for reversing them.

A new transgenic reporter line reveals Wnt-dependent Snail2 re-expression and cranial neural crest differentiation in Xenopus

Li, J. — 2019-01-15

During vertebrate embryogenesis, the cranial neural crest (CNC) forms at the neural plate border and subsequently migrates and differentiates into many types of cells. The transcription factor Snail2, which is induced by canonical Wnt signaling to be expressed in the early CNC, is pivotal for CNC induction and migration in Xenopus. However, snail2 expression is silenced during CNC migration, and its roles at later developmental stages remain unclear. We generated a transgenic X. tropicalis line that expresses enhanced green fluorescent protein (eGFP) driven by the snail2 promoter/enhancer, and observed eGFP expression not only in the pre-migratory and migrating CNC, but also the differentiating CNC. This transgenic line can be used directly to detect deficiencies in CNC development at various stages, including subtle perturbation of CNC differentiation. In situ hybridization and immunohistochemistry confirm that Snail2 is reexpressed in the differentiating CNC. Using a separate transgenic Wnt reporter line, we show that canonical Wnt signaling is also active in the differentiating CNC. Blocking Wnt signaling shortly after CNC migration causes reduced snail2 expression and impaired differentiation of CNC-derived head cartilage structures. These results suggest that Wnt signaling drives the reexpression of snail2 in the post-migratory CNC and regulates CNC differentiation.

Corazonin neuroendocrine pathway orchestrates stress-associated physiology in Drosophila

Zandawala, M. — 2019-01-16

Environmental factors challenge the physiological homeostasis in animals, thereby evoking stress responses. Various mechanisms have evolved to counter stress at the organism level, including regulation by neuropeptides. In recent years, much progress has been made on the mechanisms and neuropeptides that regulate responses to metabolic/nutritional stress, as well as those involved in countering osmotic and ionic stresses. Here, we identified a peptidergic pathway that links these types of regulatory functions. We uncover the neuropeptide Corazonin (Crz), previously implicated in responses to metabolic stress, as a neuroendocrine factor that inhibits the release of a diuretic hormone, CAPA, and thereby modulates the tolerance to osmotic and ionic stress. Both knockdown of Crz and acute injections of Crz peptide impact desiccation tolerance and recovery from chill-coma. Mapping of the Crz receptor (CrzR) expression identified three pairs of Capa-expressing neurons (Va neurons) in the ventral nerve cord that mediate these effects of Crz. We show that Crz acts to restore water/ion homeostasis by inhibiting release of CAPA neuropeptides via inhibition of cAMP production in Va neurons. Knockdown of CrzR in Va neurons affects CAPA signaling, and consequently increases tolerance for desiccation, ionic stress and starvation, but delays chill-coma recovery. Optogenetic activation of Va neurons stimulates excretion and simultaneous activation of Crz and CAPA-expressing neurons reduces this response, supporting the inhibitory action of Crz. Thus, Crz inhibits Va neurons to maintain osmotic and ionic homeostasis, which in turn affects stress tolerance. Earlier work demonstrated that systemic Crz signaling restores nutrient levels by promoting food search and feeding. Here we additionally propose that Crz signaling also ensures osmotic homeostasis by inhibiting release of CAPA neuropeptides and suppressing diuresis. Thus, Crz ameliorates stress-associated physiology through systemic modulation of both peptidergic neurosecretory cells and the fat body in Drosophila. Author summaryInsects are among the largest groups of animals and have adapted to inhabit almost all environments on Earth. Their success in surviving extreme conditions stems largely from their ability to withstand environmental stress, such as desiccation and cold. However, the neural mechanisms that are responsible for coordinating responses to counter these stresses are largely unknown. To address this, we delineate a neuroendocrine axis utilizing the neuropeptides Corazonin (Crz) and CAPA, that coordinate responses to metabolic and osmotic stress. We show that Crz inhibits the release of a diuretic peptide, CAPA from a set of neurosecretory cells. CAPA in turn influences osmotic and ionic balance via actions on the Malpighian tubules (the insect analogs of the kidney) and the intestine. Taken together with earlier work, our data suggest that Crz acts to restore metabolic homeostasis at starvation and osmotic homeostasis during desiccation by inhibiting release of the diuretic hormone CAPA. Hence, this work provides a mechanistic understanding of the neuroendocrine mitigation of metabolic and osmotic stress by two peptide systems.

Waterborne, abiotic and other indirectly transmitted (W.A.I.T.) infections are defined by the dynamics of free-living pathogens and environmental reservoirs

Miller-Dickson, M. — 2019-01-20

While the ecology of infectious disease is a rich field with decades worth of empirical evidence and theory, there are aspects that remain relatively under-examined. One example is the importance of the free-living survival stage of certain pathogens, and especially is cases where they are transmitted indirectly between hosts through an environmental reservoir intermediate. In this study, we develop an integrated, broadly applicable mathematical method to examine diseases fitting this description--the waterborne, abiotic and other indirectly transmitted (W.A.I.T.) infection framework. To demonstrate its utility, we construct realistic models of two very different epidemic scenarios: cholera in a densely populated setting with limited access to clean drinking water and hepatitis C virus in an urban setting of injection-drug users. Using these two exemplars, we find that the W.A.I.T. model fortifies the centrality of reservoir dynamics in the "sit and wait" infection strategy, and provides a way to simulate a diverse set of intervention strategies.

Inferring tumor evolution from longitudinal samples

Myers, M. A. — 2019-01-22

Background: Determining the clonal composition and somatic evolution of a tumor greatly aids in accurate prognosis and effective treatment for cancer. In order to understand how a tumor evolves over time and/or in response to treatment, multiple recent studies have performed longitudinal DNA sequencing of tumor samples from the same patient at several different time points. However, none of the existing algorithms that infer clonal composition and phylogeny using several bulk tumor samples from the same patient integrate the information that these samples were obtained from longitudinal observations. Results: We introduce a model for a longitudinally-observed phylogeny and derive constraints that longitudinal samples impose on the reconstruction of a phylogeny from bulk samples. These constraints form the basis for a new algorithm, Cancer Analysis of Longitudinal Data through Evolutionary Reconstruction (CALDER), which infers phylogenetic trees from longitudinal bulk DNA sequencing data. We show on simulated data that constraints from longitudinal sampling can substantially reduce ambiguity when deriving a phylogeny from multiple bulk tumor samples, each a mixture of tumor clones. On real data, where there is often considerable uncertainty in the clonal composition of a sample, longitudinal constraints yield more parsimonious phylogenies with fewer tumor clones per sample. We demonstrate that CALDER reconstructs more plausible phylogenies than existing methods on two longitudinal DNA sequencing datasets from chronic lymphocytic leukemia patients. These findings show the advantages of directly incorporating temporal information from longitudinal sampling into tumor evolution studies. Availability: CALDER is available at https://github.com/raphael-group.

Intracortical neural activity distal to seizure-onset-areas predicts human focal seizures

Proix, T. — 2019-01-24

The apparent unpredictability of epileptic seizures has a major impact in the quality of life of people with pharmacologically resistant seizures. Here, we present initial results and a proof-of-concept of how focal seizures can be predicted early in advance based on intracortical signals recorded from small neocortical patches away from identified seizure onset areas. We show that machine learning algorithms can discriminate between interictal and preictal periods based on multiunit activity (i.e. thresholded action potential counts) and multi-frequency band local field potentials recorded via 4 X 4 mm2 microelectrode arrays. Microelectrode arrays were implanted in 5 patients undergoing neuromonitoring for resective surgery. Post-implant analysis revealed arrays were outside the seizure onset areas. Preictal periods were defined as the 1-hour period leading to a seizure. A 5-minute gap between the preictal period and the putative seizure onset was enforced to account for potential errors in the determination of actual seizure onset times. We used extreme gradient boosting and long short-term memory networks for prediction. Prediction accuracy based on the area under the receiver operating characteristic curves reached 90% for at least one feature type in each patient. Importantly, successful prediction could be achieved based exclusively on multiunit activity. This result indicates that preictal activity in the recorded neocortical patches involved not only subthreshold postsynaptic potentials, perhaps driven by the distal seizure onset areas, but also neuronal spiking in distal recurrent neocortical networks. Beyond the commonly identified seizure onset areas, our findings point to the engagement of large-scale neuronal networks in the neural dynamics building up toward a seizure. Our initial results obtained on currently available human intracortical recordings warrant new studies on larger datasets, and open new perspectives for seizure prediction and control by emphasizing the contribution of multiscale neural signals in large-scale neuronal networks.

The hidden costs of dietary restriction: implications for its evolutionary and mechanistic origins

McCracken, A. W. — 2019-01-29

Dietary restriction (DR) consistently and universally extends health-and lifespan across taxa. Despite efforts to uncover the mechanisms underpinning DR - and ultimately translate its beneficial outcomes to humans - precise and universal mechanisms have not been identified. In biomedical science, the effects of DR are interpreted as regulating pro-longevity molecular pathways. This reasoning is guided by the conviction that DR evolved as an adaptive, pro-longevity physiological response to restricted food intake. Current evolutionary theory states that organisms should invest in their soma more heavily during periods of DR, and, when their resource availability improves, should outcompete age-matched rich-fed controls in survival and/or reproduction. Here we present a formal test of these key predictions utilising a large-scale demographic approach detailing mortality and fecundity in Drosophila melanogaster fed alternating dietary regimes (N > 66,000 flies across 11 genetic lines). Our experiments reveal surprising and substantial mortality costs when returning to a rich diet after periods of DR. These results suggest the effects of DR are not necessarily intrinsically pro-longevity and could be considered an escape from costs incurred under nutrient-rich conditions, in addition to novel, discrete costs associated with restricting dietary protein. These insights question the relevance of current evolutionary explanations of DR in interpreting the mechanistic basis of dietary restriction.

Sensorimotor adaptation compensates for distortions of 3D shape information

Cesanek, E. — 2019-02-04

Visual perception often fails to recover the veridical 3D shape of objects in the environment due to ambiguity and variability in the available depth cues. However, we rely heavily on 3D shape estimates when planning movements, for example reaching to pick up an object from a slanted surface. Given the wide variety of distortions that can affect 3D perception, how do our actions remain accurate across different environments? One hypothesis is that the visuomotor system performs selective filtering of 3D information to minimize distortions. Indeed, some studies have found that actions appear to preferentially target stereo information when it is put in conflict with texture information. However, since these studies analyze averages over multiple trials, this apparent preference could be produced by sensorimotor adaptation. In Experiment 1, we create a set of cue-conflict stimuli where one available depth cue is affected by a constant bias. Sensory feedback rapidly aligns the motor output with physical reality in just a few trials, which can make it seem as if action planning selectively relies on the reinforced cue--yet no change in the relative influences of the cues is necessary to eliminate the constant errors. In contrast, when one depth cue becomes less correlated with physical reality, variable movement errors will occur, causing canonical adaptation to fail as the opposite error corrections cancel out. As a result, canonical adaptation cannot explain the preference for stereo found in studies with variable errors. However, Experiment 2 shows that persistent errors can produce a novel form of adaptation that gradually reduces the relative influence of an unreliable depth cue. These findings show that grasp control processes are continuously modified based on sensory feedback to compensate for both biases and noise in 3D visual processing, rather than having a hardwired preference for one type of depth information.

Cue-selective adaptation operates on a separable encoding of stereo and texture information

Cesanek, E. — 2019-02-05

To perform accurate movements, the sensorimotor system must maintain a delicate calibration of the mapping between visual inputs and motor outputs. Previous work has focused on the mapping between visual inputs and individual locations in egocentric space, but little attention has been paid to the mappings that support interactions with 3D objects. In this study, we investigated sensorimotor adaptation of grasping movements targeting the depth dimension of 3D paraboloid objects. Object depth was specified by separately manipulating binocular disparity (stereo) and texture gradients. At the end of each movement, the fingers closed down on a physical object consistent with one of the two cues, depending on the condition (haptic-for-texture or haptic-for-stereo). Unlike traditional adaptation paradigms, where relevant spatial properties are determined by a single dimension of visual information, this method enabled us to investigate whether adaptation processes can selectively adjust the influence of different sources of visual information depending on their relationship to physical depth. In two experiments, we found short-term changes in grasp performance consistent with a process of cue-selective adaptation: the slope of the grip aperture with respect to a reliable cue (correlated with physical reality) increased, whereas the slope with respect to the unreliable cue (uncorrelated with physical reality) decreased. In contrast, slope changes did not occur during exposure to a set of stimuli where both cues remained correlated with physical reality, but one was rendered with a constant bias of 10 mm; the grip aperture simply became uniformly larger or smaller, as in standard adaptation paradigms. Overall, these experiments support a model of cue-selective adaptation driven by correlations between error signals and input values (i.e., supervised learning), rather than mismatched haptic and visual signals.

Characterization of prevalence and health consequences of uniparental disomy in four million individuals from the general population

Nakka, P. — 2019-02-05

Meiotic nondisjunction and resulting aneuploidy can lead to severe health consequences in humans. Aneuploidy rescue can restore euploidy but may result in uniparental disomy (UPD), the inheritance of both homologs of a chromosome from one parent with no representative copy from the other. Current understanding of UPD is limited to ~3,300 cases for which UPD was associated with clinical presentation due to imprinting disorders or recessive diseases. Thus, the prevalence of UPD and its phenotypic consequences in the general population are unknown. We searched for instances of UPD in over four million consented research participants from the personal genetics company 23andMe, Inc., and 431,094 UK Biobank participants. Using computationally detected DNA segments identical-by-descent (IBD) and runs of homozygosity (ROH), we identified 675 instances of UPD across both databases. Here we present the first characterization of UPD prevalence in the general population, a machine-learning framework to detect UPD using ROH, and a novel association between autism and UPD of chromosome 22.

netNMF-sc: Leveraging gene-gene interactions for imputation and dimensionality reduction in single-cell expression analysis

Elyanow, R. — 2019-02-08

MotivationSingle-cell RNA-sequencing (scRNA-seq) enables high throughput measurement of RNA expression in individual cells. Due to technical limitations, scRNA-seq data often contain zero counts for many transcripts in individual cells. These zero counts, or dropout events, complicate the analysis of scRNA-seq data using standard analysis methods developed for bulk RNA-seq data. Current scRNA-seq analysis methods typically overcome dropout by combining information across cells, leveraging the observation that cells generally occupy a small number of RNA expression states. ResultsWe introduce netNMF-sc, an algorithm for scRNA-seq analysis that leverages information across both cells and genes. netNMF-sc combines network-regularized non-negative matrix factorization with a procedure for handling zero inflation in transcript count matrices. The matrix factorization results in a low-dimensional representation of the transcript count matrix, which imputes gene abundance for both zero and non-zero entries and can be used to cluster cells. The network regularization leverages prior knowledge of gene-gene interactions, encouraging pairs of genes with known interactions to be close in the low-dimensional representation. We show that netNMF-sc outperforms existing methods on simulated and real scRNA-seq data, with increasing advantage at higher dropout rates (e.g. above 60%). Furthermore, we show that the results from netNMF-sc - including estimation of gene-gene covariance - are robust to choice of network, with more representative networks leading to greater performance gains. AvailabilitynetNMF-sc is available at github.com/raphael-group/netNMF-sc. Contactbraphael@princeton.edu

Generalizing to generalize: when (and when not) to be compositional in task structure learning

Franklin, N. T. — 2019-02-12

Humans routinely face novel environments in which they have to generalize in order toact adaptively. However, doing so involves the non-trivial challenge of deciding which aspects of a task domain to generalize. While it is sometimes appropriate to simply re-use a learned behavior, often adaptive generalization entails recombining distinct components of knowledge acquired across multiple contexts. Theoretical work has suggested a computational trade-off in which it can be more or less useful to learn and generalize aspects of task structure jointly or compositionally, depending on previous task statistics, but empirical studies are lacking. Here we develop a series of navigation tasks which manipulate the statistics of goal values ("what to do") and state transitions ("how to do it") across contexts, and assess whether human subjects generalize these task components separately or conjunctively. We find that human generalization is sensitive to the statistics of the previously experienced task domain, favoring compositional or conjunctive generalization when the task statistics are indicative of such structures, and a mixture of the two when they are more ambiguous. These results support the predictions of a normative "meta-generalization learning" agent that does not only generalize previous knowledge but also generalizes the statistical structure most likely to support generalization. Author NoteThis work was supported in part by the National Science Foundation Proposal 1460604 "How Prefrontal Cortex Augments Reinforcement Learning" to MJF. We thank Mark Ho for providing code used in the behavioral task. We thank Matt Nassar for helpful discussions. Correspondence should be addressed to Nicholas T. Franklin (nfranklin@fas.harvard.edu) or Michael J. Frank (michael_frank@brown.edu).

The human non-visual opsin OPN3 regulates pigmentation of epidermal melanocytes through interaction with MC1R

Ozdeslik, R. N. — 2019-02-18

Opsins form a family of light-activated, retinal-dependent G-protein coupled receptors (GPCRs) that serve a multitude of visual and non-visual functions. Opsin3 (OPN3 or encephalopsin), initially identified in the brain, remains one of the few members of the mammalian opsin family with unknown function and ambiguous light-absorption properties. We recently discovered that OPN3 is highly expressed in human epidermal melanocytes--the skin cells that produce melanin. The melanin pigment is a critical defense against ultraviolet radiation and its production is mediated by the Gs-coupled melanocortin-1 receptor (MC1R). The physiological function and light-sensitivity of OPN3 in melanocytes is yet to be determined. Here we show that in human epidermal melanocytes OPN3 acts as a negative regulator of melanin production by interacting with MC1R and modulating its cAMP signaling. OPN3 negatively regulates the cAMP response evoked by MC1R via activation of the Gi subunit of G-proteins, thus decreasing cellular melanin levels. In addition to their functional relationship, OPN3 and MC1R colocalize at both the plasma membrane and in intracellular structures and form a physical complex. Remarkably, OPN3 can bind retinal, but does not mediate light-induced signaling in melanocytes. Our results identify a novel function for OPN3 in the regulation of the melanogenic pathway in epidermal melanocytes. Our results reveal a light-independent function for the poorly characterized OPN3 and a novel pathway that greatly expands our understanding of melanocyte and skin physiology. SignificanceOur data reveals a novel function for the non-visual opsin OPN3 in regulating the pigmentation of human melanocytes by interacting with and modulating the activity of MC1R.

Notch signalling mediates secondary senescence

Teo, Y. V. — 2019-02-19

Oncogene induced senescence (OIS) is a tumour suppressive response to oncogene activation that can be transmitted to neighbouring cells through secreted factors of the senescence associated secretory phenotype (SASP). Using single-cell transcriptomics we observed two distinct endpoints, a primary marked by Ras and a secondary by Notch. We find that secondary senescence in vitro and in vivo requires Notch, rather than SASP alone as previously thought. Currently, primary and secondary senescent cells are not thought of as functionally distinct endpoints. A blunted SASP response and the induction of fibrillar collagens in secondary senescence compared to OIS point towards a functional diversification. One Sentence SummaryNotch signalling is an essential driver of secondary senescence with primary and secondary senescence being distinct molecular endpoints.

A Machine Learning Approach for Long-Term Prognosis of Bladder Cancer based on Clinical and Molecular Features

Song, Q. — 2019-02-21

Improving the consistency and reproducibility of bladder cancer prognoses necessitates the development of accurate, predictive prognostic models. Current methods of determining the prognosis of bladder cancer patients rely on manual decision-making, including factors with high intra- and inter-observer variability, such as tumor grade. To advance the long-term prediction of bladder cancer prognoses, we developed and tested a computational model to predict the 10-year overall survival outcome using population-based bladder cancer data, without considering tumor grade classification. The resulted predictive model demonstrated promising performance using a combination of clinical and molecular features, and was also strongly related to patient overall survival in Cox models. Our study suggests that machine learning methods can provide reliable long-term prognoses for bladder cancer patients, without relying on the less consistent tumor grade. If validated in clinical trials, this automated approach could guide and improve personalized management and treatment for bladder cancer patients.

Sequencing of 53,831 diverse genomes from the NHLBI TOPMed Program

Taliun, D. — 2019-03-06

Summary paragraphThe Trans-Omics for Precision Medicine (TOPMed) program seeks to elucidate the genetic architecture and disease biology of heart, lung, blood, and sleep disorders, with the ultimate goal of improving diagnosis, treatment, and prevention. The initial phases of the program focus on whole genome sequencing of individuals with rich phenotypic data and diverse backgrounds. Here, we describe TOPMed goals and design as well as resources and early insights from the sequence data. The resources include a variant browser, a genotype imputation panel, and sharing of genomic and phenotypic data via dbGaP. In 53,581 TOPMed samples, >400 million single-nucleotide and insertion/deletion variants were detected by alignment with the reference genome. Additional novel variants are detectable through assembly of unmapped reads and customized analysis in highly variable loci. Among the >400 million variants detected, 97% have frequency <1% and 46% are singletons. These rare variants provide insights into mutational processes and recent human evolutionary history. The nearly complete catalog of genetic variation in TOPMed studies provides unique opportunities for exploring the contributions of rare and non-coding sequence variants to phenotypic variation. Furthermore, combining TOPMed haplotypes with modern imputation methods improves the power and extends the reach of nearly all genome-wide association studies to include variants down to ~0.01% in frequency.

Hierarchical clustering of gene-level association statistics reveals shared and differential genetic architecture among traits in the UK Biobank

McGuirl, M. R. — 2019-03-04

Genome-wide association (GWA) studies have generally focused on a single phenotype of interest. Emerging biobanks that pair genotype data from thousands of individuals with phenotype data using medical records or surveys enable testing for genetic associations in each phenotype assayed. However, methods for characterizing shared genetic architecture among multiple traits are lagging behind. Here, we present a new method, Ward clustering to identify Internal Node branch length outliers using Gene Scores (WINGS), for characterizing shared and divergent genetic architecture among multiple phenotypes. The objective of WINGS (freely available at https://github.com/ramachandran-lab/PEGASUS-WINGS) is to identify groups of phenotypes, or "clusters", that share a core set of genes enriched for mutations in cases. We show in simulations that WINGS can reliably detect phenotype clusters across a range of percent shared architecture and number of phenotypes included. We then use the gene-level association test PEGASUS with WINGS to characterize shared genetic architecture among 87 case-control and seven quantitative phenotypes in 349,468 unrelated European-ancestry individuals from the UK Biobank. We identify 10 significant phenotype clusters that contain two to eight phenotypes. One significant cluster of seven immunological phenotypes is driven by seven genes; these genes have each been associated with two or more of those same phenotypes in past publications. WINGS offers a precise and efficient new application of Ward hierarchical clustering to generate hypotheses regarding shared genetic architecture among phenotypes in the biobank era.

Systematic evaluation and validation of reference and library selection methods for deconvolution of cord blood DNA methylation data

Gervin, K. — 2019-03-07

BackgroundUmbilical cord blood (UCB) is commonly used in epigenome-wide association studies of prenatal exposures. Accounting for cell type composition is critical in such studies as it reduces confounding due to the cell specificity of DNA methylation (DNAm). In the absence of cell sorting information, statistical methods can be applied to deconvolve heterogeneous cell mixtures. Among these methods, reference-based approaches leverage age appropriate cell-specific DNA-methylation profiles to estimate cellular composition. In UCB, four reference datasets comprising DNAm signatures profiled in purified cell populations have been published using the Illumina 450K and 850K EPIC arrays. These datasets are biologically and technically different, and currently there is no consensus on how to best apply them. Here, we systematically evaluate and compare these datasets and provide recommendations for reference-based UCB deconvolution.nnResultsWe first evaluated the four reference datasets to ascertain both the purity of the samples and the potential cell cross-contamination. We filtered samples and combined datasets to obtain a joint UCB reference. We selected deconvolution libraries using two different approaches: automatic selection using the top differentially methylated probes from the function pickCompProbes in minfi and a standardized library selected using the IDOL (Identifying Optimal Libraries) iterative algorithm. We compared the performance of each reference separately and in combination, using the two approaches for reference library selection, and validated the results in an independent cohort (Generation R Study, n=191) with matched FACS measured cell counts. Strict filtering and combination of the references significantly improved the accuracy and efficiency of cell type estimates. Ultimately, the IDOL library outperformed the library from the automatic selection method implemented in pickCompProbes.nnConclusionThese results have important implications for epigenetic studies in UCB as implementing this method will optimally reduce confounding due to cellular heterogeneity. This work provides guidelines for future reference-based UCB deconvolution and establishes a framework for combining reference datasets in other tissues.

Mortality among adults living with HIV treated for tuberculosis based on positive, negative, or no bacteriologic test results for tuberculosis: the IeDEA consortium

Humphrey, J. M. — 2019-03-08

BackgroundIn resource-constrained settings, people living with HIV (PLWH) treated for tuberculosis (TB) despite negative bacteriologic tests have a higher mortality than those treated with positive tests. Many PLWH are treated without bacteriologic testing; their mortality compared to those with bacteriologic testing is uncertain.nnMethodsWe conducted an observational cohort study among PLWH [≥] 15 years of age who initiated TB treatment at clinical sites affiliated with four regions of the International epidemiology Databases to Evaluate AIDS (IeDEA) consortium from 2012-2014: Caribbean, Central and South America, and Central, East, and West Africa. The primary exposure of interest was the TB bacteriologic test status at TB treatment initiation: positive, negative, or no test result. The hazard for death in the 12 months following TB treatment initiation was estimated using the Cox proportional hazard model, adjusted for patient- and site-level factors. Missing covariates were multiply imputed.nnResultsAmong 2,091 PLWH included, the median age at TB treatment initiation was 36 years, 44% were female, 53% had CD4 counts [≤] 200 cells/mm3, and 52% were on antiretroviral treatment (ART). Compared to patients with positive bacteriologic tests, the adjusted hazard for death was higher among patients with no test results (HR 1.56, 95% CI 1.08-2.26) but not different than those with negative tests (HR 1.28, 95% CI 0.91-1.81). Older age was also associated with a higher hazard for death, while being on ART, having a higher CD4 count, West Africa region, and tertiary facility level were associated with lower hazards for death.nnConclusionPLWH treated for TB with no bacteriologic test results were more likely to die than those treated with positive tests, underscoring the importance of TB bacteriologic diagnosis in resource-constrained settings. Research is needed to understand the causes of death among PLWH treated for TB in the absence of positive bacteriologic tests.

Circuits that encode and predict alcohol associated preference.

Scaplen, K. M. — 2019-03-15

Substance use disorders are chronic relapsing disorders often impelled by enduring memories and persistent cravings. Alcohol, as well as other addictive substances, remolds neural circuits important for memory to establish obstinate preference despite aversive consequences. How pertinent circuits are selected and shaped to result in these unchanging, inflexible memories is unclear. Using neurogenetic tools available in Drosophila melanogaster we define how circuits required for alcohol associated preference shift from population level dopaminergic activation to select dopamine neurons that predict behavioral choice. During memory expression, these dopamine neurons directly, and indirectly via the mushroom body (MB), modulate the activity of interconnected glutamatergic and cholinergic output neurons. Transsynaptic tracing of these output neurons revealed at least two regions of convergence: 1) a center of memory consolidation within the MB implicated in arousal, and 2) a structure outside the MB implicated in integration of naive and learned responses. These findings provide a circuit framework through which dopamine neuron activation shifts from reward delivery to cue onset, and provides insight into the inflexible, maladaptive nature of alcohol associated memories.

Timing and Duration of Gbx2 Expression Delineates Thalamocortical and Dopaminergic Medial Forebrain Bundle Circuitry

Normand, E. A. — 2019-03-15

Gene expression is a dynamic process, which is highly coordinated during development to ensure the proper allocation and identity of neuronal cell types within the brain. Equally important during neurodevelopment is how cohorts of neurons establish axonal projections that innervate terminal target sites. We sought to bridge the temporal dynamics of gene expression, within a specific genetic lineage, to the establishment of neuronal circuits derived from cohorts of the lineage-specific progenitors. A central goal was to be able to accomplish genetic inducible circuit mapping non-invasively and with commonly available CreER/loxP technology. Specifically, we genetically marked thalamic neuron progenitors that expressed the transcription factor Gbx2 at an early embryonic stage and tracked the formation of lineage-derived thalamocortical axons during embryogenesis. We then assessed the neural circuitry at an early postnatal stage. We show that the temporal specificity of lineage marking provides a high degree of clarity for following neural circuit development. We also determined that the onset and duration of gene expression can delineate subsets of neural circuits derived from a common lineage. For example, we uncovered a novel contribution of Gbx2-expressing progenitors to midbrain dopamine neurons and dopaminergic axons of the medial forebrain bundle. We anticipate that this system can be instructive in elucidating changes in neural circuit development in both normal development and in mutant mice in which neural circuit formation is altered.

The interaction of compliance and activation on the force-length operating range and force generating capacity of skeletal muscle

Cox, S. M. — 2019-03-16

Muscle performance is influenced by where it operates on its force-length curve. Here we explore how activation and tendon compliance interact to influence muscle operating lengths and force-generating capacity. To study this, we built a musculoskeletal model of the lower limb of the guinea fowl and simulated the force-length operating range during fixed-end fixed-posture contractions for 39 actuators under thousands of combinations of activation and posture using three different muscle models: Muscles with non-compliant tendons, muscles with compliant tendons but no activation dependent shift in optimal fiber length (L0), and muscles with both compliant tendons and activation-dependent shifts in L0. We found that activation dependent effects altered muscle fiber lengths up to 40% and increased or decreased force capacity by up to 50% during fixed-end contractions. Typically, activation-compliance effects reduce muscle force and are dominated by the effects of tendon compliance at high activations. At low activation, however, activation-dependent shifts in L0 are equally important and can result in relative force changes for low compliance muscles of up to 60%. There are regions of the force-length curve in which muscles are most sensitive to compliance and there are troughs of influence where these factors have little effect. These regions are hard to predict, though, because the magnitude and location of these areas of high and low sensitivity shift with compliance level. Here we provide a map for when these effects will meaningfully influence force capacity and an example of their contributions to force production during a static task, namely standing.

Statistical context dictates the relationship between feedback-related EEG signals and learning

Nassar, M. — 2019-03-18

Successful decision-making requires learning expectations based on experienced outcomes. This learning should be calibrated according to the surprise associated with an outcome, but also to the statistical context dictating the most likely source of surprise. For example, when occasional changepoints are expected, surprising outcomes should be weighted heavily, demanding increased learning. In contrast, when signal corruption is expected to occur occasionally, surprising outcomes can suggest a corrupt signal that should be ignored by learning systems. Here we dissociate surprising outcomes from the degree to which they demand learning using a predictive inference task and computational modeling. We show that the updating P300, a stimulus-locked electrophysiological response previously associated with adjustments in learning behavior, does so conditionally on the source of surprise. Larger P300 signals predicted greater learning in a changing context, but predicted less learning in a context where surprise was indicative of a one-off outlier (oddball). The conditional predictive relationship between the P300 and learning behavior was persistent even after adjusting for known sources of learning rate variability. Our results suggest that the P300 provides a surprise signal that is interpreted by downstream learning processes differentially according to statistical context in order to appropriately calibrate learning across complex environments.

Dissecting the cognitive phenotype of post-stroke fatigue using drift diffusion modeling of sustained attention

Ulrichsen, K. M. — 2019-03-20

Post-stroke fatigue (PSF) is a prevalent symptom among stroke patients. Its symptom burden is pervasive, persistent and associated with poor rehabilitation outcomes, though its mechanisms are poorly understood. Many patients with PSF experience cognitive difficulties, but studies aiming to identify cognitive correlates of PSF have been largely inconclusive. In contrast to conventional neuropsychological assessment, computational modeling of behavioral data allows for a dissection of specific cognitive processes associated with group or individual differences in fatigue. With the aim to zero in on the cognitive phenotype of PSF, we fitted a hierarchical drift diffusion model (hDDM) to response time data from Attention Network Test (ANT) obtained from 53 chronic stroke patients. The computational model accurately reconstructed the individual level response time distributions in the different ANT conditions, and hDDM regressions identified an interaction between trial number and fatigue symptoms on non-decision time, intuitively indicating that the cognitive phenotype of fatigue entails an increased vulnerability to sustained attentional effort. These novel results demonstrate the significance of considering the sustained nature of cognitive effort when defining the cognitive phenotype of post-stroke fatigue, and suggest that the use of computational approaches offers a further characterization of the specific processes underlying observed behavioral differences.

Epigenome-wide meta-analysis of PTSD across 10 military and civilian cohorts identifies novel methylation loci

Smith, A. K. — 2019-03-21

Differences in susceptibility to posttraumatic stress disorder (PTSD) may be related to epigenetic differences between PTSD cases and trauma-exposed controls. Such epigenetic differences may provide insight into the biological processes underlying the disorder. Here we describe the results of the largest DNA methylation meta-analysis of PTSD to date with data from the Psychiatric Genomics Consortium (PGC) PTSD Epigenetics Workgroup. Ten cohorts, military and civilian, contributed blood-derived DNA methylation data (HumanMethylation450 BeadChip) from 1,896 PTSD cases (42%) and trauma-exposed controls (58%). Utilizing a common QC and analysis strategy, we identified ten CpG sites associated with PTSD (9.61E-07

Modeling mechanisms of tremor reduction for essential tremor using symmetric biphasic DBS

Lee, S. — 2019-03-21

Essential tremor (ET) is the most common movement disorder, in which the primary symptom is a prominent, involuntary 4-10 Hz movement. For severe, medication refractory cases, deep brain stimulation (DBS) targeting the ventral intermediate nucleus of the thalamus (VIM) can be an effective treatment for cessation of tremor and is thought to work in part by disrupting tremor frequency oscillations (TFOs) in VIM. However, DBS is not universally effective and may be further disrupting cerebellar-mediated activity in the VIM. Here, we applied biophysically detailed computational modeling to investigate whether the efficacy of DBS is affected by the mechanism of generation of TFOs or by the pattern of stimulation. We simulated the effects of DBS using standard, asymmetric pulses as well as biphasic, symmetric pulses to understand biophysical mechanisms of how DBS disrupts TFOs generated either extrinsically or intrinsically. The model results suggested that the efficacy of DBS in the VIM is affected by the mechanism of generation of TFOs. Symmetric biphasic DBS reduced TFOs more than standard DBS in both networks, and these effects were stronger in the intrinsic network. For intrinsic tremor frequency activity, symmetric biphasic DBS was more effective at reducing TFOs. Simulated non-tremor signals were also transmitted during symmetric biphasic DBS, suggesting that this type of DBS may help to reduce side effects caused by disruption of the cerebellothalamocortical pathway. Biophysical details in the model provided a mechanistic interpretation of the cellular and network dynamics contributing to these effects that can be empirically tested in future studies.nnSignificance StatementEssential tremor (ET) is a common movement disorder, whose primary symptom is an involuntary rhythmic movement of the limbs or head. An area of the human tha-lamus demonstrates electrical activity that oscillates at the frequencies of tremor, and deep brain stimulation (DBS) in this area can reduce tremor. It is not fully understood how DBS affects tremor frequency activity in the thalamus, and studying different patterns of DBS stimulation may help to clarify these mechanisms. We created a computational model of different shapes of DBS and studied how they reduce different hypothesized generators of tremor frequency activity. A greater understanding of how DBS affects the thalamus may lead to improved treatments to reduce tremor and alleviate side effects in patients with ET.

Germline features associated with immune infiltration in solid tumors

Shahamatdar, S. — 2019-03-24

Given the clinical success of immune checkpoint blockade (ICB) across a diverse set of solid tumors, and the emerging role for different immune infiltrates in contributing to response to ICB, a comprehensive assessment of the properties that dictate immune infiltrations may reveal new biological insights and inform the development of new effective therapies. Multiple studies have examined somatic and functional immune properties associated with different tumor infiltrates; however, germline features that associate with specific immune infiltrates in cancers have been incompletely characterized. Here, we analyzed over 7 million autosomal germline variants in the TCGA cohort (5788 European-ancestry samples across 30 cancer types) and tested for pan-cancer association with established immune-related phenotypes that describe the tumor immune microenvironment. We identified: one SNP associated with the fraction of follicular helper T cells in bulk tumor; 77 unique candidate genes, some of which are involved in cytokine-mediated signaling (e.g. CNTF and TRIM34) and cancer pathogenesis (e.g. ATR and AKAP9); and subnetworks with genes that are part of DNA repair (RAD51 and XPC) and transcription elongation (CCNT2) pathways. We found a positive association between polygenic risk for rheumatoid arthritis and absolute fraction of infiltrating CD8 T cells. Overall, we identified multiple germline genetic features associated with specific tumor-immune phenotypes across cancer, and developed a framework for probing inherited features that contribute to variation in immune infiltration.

Recovery of trait heritability from whole genome sequence data

Wainschtein, P. — 2019-03-25

Heritability, the proportion of phenotypic variance explained by genetic factors, can be estimated from pedigree data 1, but such estimates are uninformative with respect to the underlying genetic architecture. Analyses of data from genome-wide association studies (GWAS) on unrelated individuals have shown that for human traits and disease, approximately one-third to two-thirds of heritability is captured by common SNPs 2-5. It is not known whether the remaining heritability is due to the imperfect tagging of causal variants by common SNPs, in particular if the causal variants are rare, or other reasons such as overestimation of heritability from pedigree data. Here we show that pedigree heritability for height and body mass index (BMI) appears to be largely recovered from whole-genome sequence (WGS) data on 25,465 unrelated individuals of European ancestry. We assigned 33.7 million genetic variants to groups based upon their minor allele frequencies (MAF) and linkage disequilibrium (LD) with variants nearby, and estimated and partitioned genetic variance accordingly. The estimated heritability was 0.68 (SE 0.10) for height and 0.30 (SE 0.10) for BMI, with a range of ~0.60 - 0.71 for height and ~0.25 - 0.35 for BMI, depending on quality control and analysis strategies. Low-MAF variants in low LD with neighbouring variants were enriched for heritability, to a greater extent for protein-altering variants, consistent with negative selection thereon. Cumulatively variants with 0.0001 < MAF < 0.1 explained 0.47 (SE 0.07) and 0.30 (SE 0.10) of heritability for height and BMI, respectively. Our results imply that rare variants, in particular those in regions of low LD, is a major source of the still missing heritability of complex traits and disease.

Proteinarium: Multi-Sample Protein-Protein Interaction Visualization and Analysis Tool

Armanious, D. — 2019-03-26

BackgroundData analysis has become crucial in the post genomic era where the accumulation of genomic information is mounting exponentially. Analyzing protein-protein interactions in the context of the interactome is a powerful approach to understanding disease phenotypes. ResultsWe describe Proteinarium, a multi-sample protein-protein interaction network analysis and visualization tool. Proteinarium can be used to analyze data for samples with dichotomous phenotypes, multiple samples from a single phenotype or a single sample. Then, by similarity clustering, the network-based relations of samples are identified and clusters of related samples are presented as a dendrogram. Each branch of the dendrogram is built based on network similarities of the samples. The protein-protein interaction networks can be analyzed and visualized on any branch of the dendrogram. Proteinariums input can be derived from transcriptome analysis, whole exome sequencing data or any high-throughput screening approach. Its strength lies in use of gene lists for each sample as a distinct input which are further analyzed through protein interaction analyses. Proteinarium output includes the gene lists of visualized networks and PPI interaction files where users can analyze the network(s) on other platforms such as Cytoscape. In addition, since the dendrogram is written in Newick tree format, users can visualize it in other software platforms like Dendroscope, ITOL. ConclusionsProteinarium, through the analysis and visualization of PPI networks, allows researchers to make important observations on high throughput data for a variety of research questions. Proteinarium identifies significant clusters of patients based on their shared network similarity for the disease of interest and the associated genes. Proteinarium is a command-line tool written in Java with no external dependencies and it is freely available at https://github.com/Armanious/Proteinarium.

VIVA (VIsualization of VAriants): A VCF file visualization tool

Tollefson, G. A. — 2019-03-28

The volume and pace of data accumulation from high-throughput sequencing studies have been amplified by recent rapid technological advances in biological sciences. Visualization of genomic data is essential for quality control, exploration, and interpretation. Here, we describe a user-friendly visualization tool for variant call format (VCF) files with which the users can interactively evaluate and share genomic data, as well as create publication quality graphics.

Epsilon-Genic Effects Bridge the Gap Between Polygenic and Omnigenic Complex Traits

Cheng, W. — 2019-04-02

Traditional univariate genome-wide association studies generate false positives and negatives due to difficulties distinguishing associated variants from variants with spurious nonzero effects that do not directly influence the trait. Recent efforts have been directed at identifying genes or signaling pathways enriched for mutations in quantitative traits or case-control studies, but these can be computationally costly and hampered by strict model assumptions. Here, we present gene-{varepsilon}, a new approach for identifying statistical associations between sets of variants and quantitative traits. Our key insight is that enrichment studies on the gene-level are improved when we reformulate the genome-wide SNP-level null hypothesis to identify spurious small-to-intermediate SNP effects and classify them as non-causal. gene-{varepsilon} efficiently identifies enriched genes under a variety of simulated genetic architectures, achieving greater than a 90% true positive rate at 1% false positive rate for polygenic traits. Lastly, we apply gene-{varepsilon} to summary statistics derived from six quantitative traits using European-ancestry individuals in the UK Biobank, and identify enriched genes that are in biologically relevant pathways. Author SummaryEnrichment tests augment the standard univariate genome-wide association (GWA) framework by identifying groups of biologically interacting mutations that are enriched for associations with a trait of interest, beyond what is expected by chance. These analyses model local linkage disequilibrium (LD), allow many different mutations to be disease-causing across patients, and generate biologically interpretable hypotheses for disease mechanisms. However, existing enrichment analyses are hampered by high computational costs, and rely on GWA summary statistics despite the high false positive rate of the standard univariate GWA framework. Here, we present the gene-level association framework gene-{varepsilon} (pronounced "genie"), an empirical Bayesian approach for identifying statistical associations between sets of mutations and quantitative traits. The central innovation of gene-{varepsilon} is reformulating the GWA null model to distinguish between (i) mutations that are statistically associated with the disease but are unlikely to directly influence it, and (ii) mutations that are most strongly associated with a disease of interest. We find that, with our reformulated SNP-level null hypothesis, our gene-level enrichment model outperforms existing enrichment methods in simulation studies and scales well for application to emerging biobank datasets. We apply gene-{varepsilon} to six quantitative traits in the UK Biobank and recover novel and functionally validated gene-level associations.

Quantifying the mechanics of locomotion of the schistosome pathogen with respect to changes in its physical environment

Zhang, S. — 2019-04-05

Schistosomiasis is a chronic and morbid disease of poverty affecting approximately 200 million people worldwide. Mature schistosome flatworms wander in the hosts hepatic portal and mesenteric venous system where they encounter a range of blood flow conditions and geometrical confinement. However, the mechanisms that support schistosome locomotion and underlie the pathogens adaptation to its physical environment are largely unknown. By combining microfabrication and traction force microscopy, we developed various in vitro assays to quantify the mechanics of locomotion of adult male S. mansoni in different physiologically relevant conditions. We show that in unconfined settings, the parasite undergoes two-anchor marching mediated by the coordinated action of its oral and ventral suckers. This mode of locomotion is maintained when the worm faces an external flow, to which it responds by adjusting the strength of its suckers. In geometrically confined conditions, S. mansoni switches to a different crawling modality by generating retrograde peristaltic waves along its body, a mechanism shared with terrestrial and marine worms. But while the surface of most worms has backward-pointing bristles that rectify peristaltic waves and facilitate forward locomotion, S mansoni has isotropically oriented tubercles. This requires tight coordination between muscle contraction and substrate friction but confers S. mansoni the ability to reverse its direction of locomotion without turning its body, which is likely advantageous to maneuver in narrow bore vessels. We show that the parasite can also coordinate the action of its suckers with its peristaltic body contractions to increase crawling speed. Throughout this study, we report on a number of biomechanical parameters to quantify the motility of adult schistosomes (e.g. sucker grabbing strength, rate of detachment under flow, peristaltic wave properties and traction stresses). The new series of in vitro assays make it possible to quantify key phenotypical aspects of S. mansoni motility that could guide the discovery of new drugs to treat schistosomiasis.

Goal congruency dominates reward value in accounting for behavioral and neural correlates of value-based decision-making

Froemer, R. — 2019-04-05

How we engage with a set of options (e.g., items on a menu) is affected both by the rewards they promise and our goal in choosing between them. Typically, our goal is to maximize potential reward and minimize potential punishment. Previous research on value-based decision-making has characterized how people make decisions with this goal in mind, identifying consistent behavioral and neural signatures associated with how rewarding a set of choice options are, overall and relative to one another. However, these studies suffer from a common confound: in each case, more rewarding options were also more congruent with ones goal of choosing the best option. Previous findings can therefore be accounted for by the reward value or the goal congruency of ones options. To compare these accounts directly, we had participants make choices while their goal varied between choosing the best or the worst option, resulting in situations where either high-reward or low-reward options were most goal-congruent. We found that goal congruency uniquely accounts for past observations that decision speed varies with the overall value of ones options. Within brain regions commonly associated with choice value, fMRI activity was associated with both relative and overall goal congruency. These goal congruency correlates were dissociable from separate correlates of the overall reward associated with a choice set (independent of ones goal). Our findings call for a reinterpretation of previous research on value-based choice, and offer an approach moving forward for disentangling the roles of rewards and goals in how we evaluate our options. Significance StatementWhether it is between restaurants or career paths, to make adaptive decisions we must evaluate our options and identify those that are most conducive to our current goal. Dysfunctional decision-making can therefore result from aberrant reward processing (e.g., impulse disorders) or from aberrant goal processing (e.g., OCD, ADHD). By focusing only on how people choose their favorite option in a choice set (when rewards and goals are aligned), past research has been unable to distinguish the degree to which behavior and neural activity are determined by reward versus goal processing. We disentangle these processes and show that behavior and fMRI activity are differentially influenced by the promised rewards versus the degree to which those rewards align with ones current goal.

Bayesian Estimation of Population Size Changes by Sampling Tajima’s Trees

Palacios, J. A. — 2019-04-11

The large state space of gene genealogies is a major hurdle for inference methods based on Kingmans coalescent. Here, we present a new Bayesian approach for inferring past population sizes which relies on a lower resolution coalescent process we refer to as "Tajimas coalescent". Tajimas coalescent has a drastically smaller state space, and hence it is a computationally more efficient model, than the standard Kingman coalescent. We provide a new algorithm for efficient and exact likelihood calculations for data without recombination, which exploits a directed acyclic graph and a correspondingly tailored Markov Chain Monte Carlo method. We compare the performance of our Bayesian Estimation of population size changes by Sampling Tajimas Trees (BESTT) with a popular implementation of coalescent-based inference in BEAST using simulated data and human data. We empirically demonstrate that BESTT can accurately infer effective population sizes, and it further provides an efficient alternative to the Kingmans coalescent. The algorithms described here are implemented in the R package phylodyn, which is available for download at https://github.com/JuliaPalacios/phylodyn.

Age-Related Changes in the Neural Dynamics of Bottom-Up and Top-Down Processing During Visual Object Recognition: An Electrophysiological Investigation

Lai, L. Y. — 2019-04-13

When recognizing objects in our environments, we rely on both what we see and what we know. While elderly adults have been found to display increased sensitivity to top-down influences of contextual information during object recognition, the locus of this increased sensitivity remains unresolved. To address this issue, we examined the effects of aging on the neural dynamics of bottom-up and top-down visual processing during rapid object recognition. Specific EEG ERP components indexing bottom-up and top-down processes along the visual processing stream were assessed while systematically manipulating the degree of object ambiguity and scene context congruity. An increase in early attentional feedback mechanisms (as indexed by N1) as well as a functional reallocation of executive attentional resources (as indexed by P200) prior to object identification were observed in elderly adults, while post-perceptual semantic integration (as indexed by N400) remained intact. These findings suggest that compromised bottom-up perceptual processing of visual input in healthy aging leads to an increased involvement of top-down processes to resolve greater perceptual ambiguity during object recognition.

Spatiotemporally distinct neural mechanisms underlie our reactions to and comparison between value-based options

Froemer, R. — 2019-04-16

Previous research suggests that people evaluate options in at least two ways: (1) appraising their overall value and (2) choosing between them. Here we test whether these processes are temporally dissociable, with appraisal-related processes tied to the time ones options appear and choice-related processes tied to the time a decision is made. We recorded EEG while participants individually rated and subsequently made choices between consumer goods. As predicted, we found appraisal-related neural activity locked to the onset of the stimuli and choice-related activity locked to (and preceding) the response. Patterns of appraisal- and choice-related activity were further associated with distinct topographical profiles. Using a novel neural index of ones certainty about a given items value, we also provide evidence that choices and choice-related activity were further modulated by this option-specific value certainty. Taken together, our results support the hypothesis that spatiotemporally distinct mechanisms underlie appraisal and choice, suggesting that commonly observed neural correlates of choice value may reflect either or both of these processes.

Decoding Complex Sounds Using Broadband Population Recordings from Secondary Auditory Cortex of Macaques

Heelan, C. — 2019-04-20

Direct electronic communication with sensory areas of the neocortex is a challenging ambition for brain-computer interfaces. Here, we report the first successful neural decoding of English words with high intelligibility from intracortical spike-based neural population activity recorded from the secondary auditory cortex of macaques. We acquired 96-channel full-broadband population recordings using intracortical microelectrode arrays in the rostral and caudal parabelt regions of the superior temporal gyrus (STG). We leveraged a new neural processing toolkit to investigate the choice of decoding algorithm, neural preprocessing, audio representation, channel count, and array location on neural decoding performance. The results illuminated a view of the auditory cortex as a spatially distributed network and a general purpose processor of complex sounds. The presented spike-based machine learning neural decoding approach may further be useful in informing future encoding strategies to deliver direct auditory percepts to the brain as specific patterns of microstimulation.

A methodology for morphological feature extraction and unsupervised cell classification

Bhaskar, D. — 2019-04-30

Cell morphology is an important indicator of cell state, function, stage of development, and fate in both normal and pathological conditions. Cell shape is among key indicators used by pathologists to identify abnormalities or malignancies. With rapid advancements in the speed and amount of biological data acquisition, including images and movies of cells, computer-assisted identification and analysis of images becomes essential. Here, we report on techniques for recognition of cells in microscopic images and automated cell shape classification. We illustrate how our unsupervised machine-learning-based approach can be used to classify distinct cell shapes from a large number of microscopic images.nnTechnical AbstractWe develop a methodology to segment cells from microscopy images and compute quantitative descriptors that characterize their morphology. Using unsupervised techniques for dimensionality reduction and density-based clustering, we perform label-free cell shape classification. Cells are identified with minimal user input using mathematical morphology and region-growing segmentation methods. Physical quantities describing cell shape and size (including area, perimeter, Feret diameters, etc.) are computed along with other features including shape factors and Hus image moments.nnCorrelated features are combined to obtain a low-dimensional (2-D or 3-D) embedding of data points corresponding to individual segmented cell shapes. Finally, a hierarchical density-based clustering algorithm (HDBSCAN) is used to classify cells. We compare cell classification results obtained from different combinations of features to identify a feature set that delivers optimum classification performance for our test data consisting of phase-contrast microscopy images of a pancreatic-cancer cell line, MIA PaCa-2.

Encoding information in synthetic metabolomes

Kennedy, E. — 2019-05-05

Biomolecular information systems offer numerous potential advantages over conventional semiconductor technologies. Downstream from DNA, the metabolome is an information-rich molecular system with diverse chemical dimensions which could be harnessed for information storage and processing. As a proof of principle of postgenomic data storage, here we demonstrate a workflow for representing abstract data in synthetic metabolomes. Our approach leverages robotic liquid handling for writing digital information into chemical mixtures, and mass spectrometry for extracting the data. We present several kilobyte-scale image datasets stored in synthetic metabolomes, which are decoded with accuracy exceeding 98-99% using multi-mass logistic regression. Cumulatively, >100,000 bits of digital image data was written into metabolomes. These early demonstrations provide insight into the benefits and limitations of postgenomic chemical information systems.

Adding noise to Markov cohort models

Iskandar, R. — 2019-05-17

Following its introduction over thirty years ago, the Markov state-transition cohort model has been used extensively to model population trajectories over time in decision modeling and cost-effectiveness studies. We recently showed that a cohort model represents the average of a continuous-time stochastic process on a multidimensional integer lattice governed by a master equation (ME), which represents the time-evolution of the probability function of a integer-valued random vector. From this theoretical connection, this study introduces an alternative modeling method, stochastic differential equation (SDE), which captures not only the mean behavior but also the variance. We first derive the continuous approximation to the master equation by relaxing integrality constraint of the state space in the form of Fokker Planck equation (FPE), which represents the time-evolution of the probability function of a real-valued random vector. Instead of working with the FPE, the SDE method constitutes time-evolution of the random vector of population counts. We derive the SDE from first principles and describe an algorithm to construct an SDE and solve the SDE via simulation for use in practice. We show the applications of SDE in two case studies. The first example demonstrates that the population trajectories, the mean and the variance, from the SDE and other commonly-used methods match. The second examples shows that users can readily apply the SDE method in their existing works without the need for additional inputs. In addition, in both examples, the SDE is superior to microsimulation in terms of computational speed. In summary, the SDE provides an alternative modeling framework and is less computationally expensive that microsimulation for a typical modeling problem in decision analyses.

Decrease in within-trial variability contributes to a decrease in across-trial variability of neural firing in the primate cortex during neural computations

Sendhilnathan, N. — 2019-05-12

The conventional approach to understanding neural responses underlying complex computations is to study across-trial averages of repeatedly performed computations from single neurons. When a brain region performs complex computations, such as processing stimulus related information or motor planning, it has been repeatedly shown through measures such as the Fano factor (FF) that neural variability across trials in the network decreases. However, in reality, multiple neurons contribute to a common computation on a single trial, rather than a single neuron contributing to a computation on multiple trials. Therefore, on individual trials the concept of FF loses significance. In this study, we extended previous work using measures of variability that are confined to a single trial and found that neurons perform a common computation, they briefly fire with increased regularity in spike timings, with similar inter-spike interval durations. We propose that this decrease in within-trial variability in neural spiking contributes to a decrease in across-trial variability in neural firing rates during network level computations. We confirmed our hypothesis by testing it on the activity of frontal eye field neurons recorded as two monkeys performed a memory-guided saccade task, and also on simulated spike trains. Furthermore, this phenomenon also has important behavioral correlates: the reaction time of the animal was faster when the within-trial variability was lower. We show that a decrease in within-trial variability is linked to a decrease in across-trial variability in neural response and indicates stationarity of neural network variability across time.nnNew & NoteworthyDuring computations, neural variability across trials decreases. In reality, multiple neurons contribute to a common computation on a single trial, rather than a single neuron contributing to a computation on multiple trials. We found that when a network of neurons performs a common computation, they briefly fire with increased regularity in spike timings. We propose that this decrease in within-trial variability in neural spiking contributes to a decrease in the observed across-trial variability.

Bayesian multivariate reanalysis of large genetic studies identifies many new associations

Turchin, M. C. — 2019-05-16

Genome-wide association studies (GWAS) have now been conducted for hundreds of phenotypes of relevance to human health. Many such GWAS involve multiple closely-related phenotypes collected on the same samples. However, the vast majority of these GWAS have been analyzed using simple univariate analyses, which consider one phenotype at a time. This is de-spite the fact that, at least in simulation experiments, multivariate analyses have been shown to be more powerful at detecting associations. Here, we conduct multivariate association analyses on 13 different publicly-available GWAS datasets that involve multiple closely-related phenotypes. These data include large studies of anthropometric traits (GIANT), plasma lipid traits (GlobalLipids), and red blood cell traits (HaemgenRBC). Our analyses identify many new associations (433 in total across the 13 studies), many of which replicate when follow-up samples are available. Overall, our results demonstrate that multivariate analyses can help make more effective use of data from both existing and future GWAS.nn1 Author SummaryGenome-wide association studies (GWAS) have become a common and powerful tool for identifying significant correlations between markers of genetic variation and physical traits of interest. Often these studies are conducted by comparing genetic variation against single traits one at a time ( univariate); however, it has previously been shown that it is possible to increase your power to detect significant associations by comparing genetic variation against multiple traits simultaneously ( multivariate). Despite this apparent increase in power though, researchers still rarely conduct multivariate GWAS, even when studies have multiple traits readily available. Here, we reanalyze 13 previously published GWAS using a multivariate method and find >400 additional associations. Our method makes use of univariate GWAS summary statistics and is available as a software package, thus making it accessible to other researchers interested in conducting the same analyses. We also show, using studies that have multiple releases, that our new associations have high rates of replication. Overall, we argue multivariate approaches in GWAS should no longer be overlooked and how, often, there is low-hanging fruit in the form of new associations by running these methods on data already collected.

Lexical landscapes as large in silico data forexamining advanced properties of fitness landscapes

Meszaros, V. A. — 2019-05-17

In silico approaches have served a central role in the development of evolutionary theory for generations. This especially applies to the concept of the fitness landscape, one of the most important abstractions in evolutionary genetics, and one which has benefited from the presence of large empirical data sets only in the last decade or so. In this study, we propose a method that allows us to generate enormous data sets that walk the line between in silico and empirical: word usage frequencies as catalogued by the Google ngram corpora. These data can be codified or analogized in terms of a multidimensional empirical fitness landscape towards the examination of advanced concepts--adaptive landscape by environment interactions, clonal competition, higher-order epistasis and countless others. We argue that the greater Lexical Landscapes approach can serve as a platform that offers an astronomical number of fitness landscapes for exploration (at least) or theoretical formalism (potentially) in evolutionary biology.

TDP-43 α-helical structure tunes liquid-liquid phase separation and function

Conicella, A. E. — 2019-05-18

Liquid-liquid phase separation (LLPS) is involved in the formation of membraneless organelles (MLOs) associated with RNA processing. Present in several MLOs, TDP-43 undergoes LLPS and is linked to the pathogenesis of amyotrophic lateral sclerosis (ALS). While some disease variants of TDP-43 disrupt self-interaction and function, here we show that designed single mutations can enhance TDP-43 assembly and function via modulating helical structure. Using molecular simulation and NMR spectroscopy, we observe large structural changes in a dimeric TDP-43. Two conserved glycine residues (G335 and G338) are potent inhibitors of helical extension and helix-helix interaction, which are removed in part by variants including the ALS-associated G335D. Substitution to helix-enhancing alanine at either of these positions dramatically enhances phase separation in vitro and decreases fluidity of phase separated TDP-43 reporter compartments in cells. Furthermore, G335A increases TDP-43 splicing function in a mini-gene assay. Therefore, TDP-43 helical region serves as a short but uniquely tunable module that shows promise as for controlling assembly and function in cellular and synthetic biology applications of LLPS.

The calcium channel subunit α2δ-3 organizes synapses via a novel activity-dependent, autocrine BMP signaling pathway

Hoover, K. M. — 2019-05-16

Synapses are highly specialized for neurotransmitter signaling, yet activity-dependent growth factor release also plays critical roles at synapses. While efficient neurotransmitter signaling is known to rely on precise apposition of release sites and neurotransmitter receptors, molecular mechanisms enabling high-fidelity growth factor signaling within the synaptic microenvironment remain obscure. Here we show that the auxiliary calcium channel subunit 2{delta}-3 promotes the function of a novel activity-dependent autocrine BMP signaling pathway at the Drosophila NMJ. 2{delta} proteins have conserved synaptogenic activity, although how they execute this function has remained elusive. We find that 2{delta}-3 provides an extracellular scaffold for autocrine BMP signaling, suggesting a new mechanistic framework for understanding 2{delta}s conserved role in synapse organization. We further establish a transcriptional requirement for activity-dependent, autocrine BMP signaling in determining synapse density, structure, and function. We propose that activity-dependent, autocrine signals provide neurons with continuous feedback on their activity state and are thus well poised to modulate synapse structure and function.

Morphology and development of the Portuguese man of war, Physalia physalis

Munro, C. — 2019-05-27

The Portuguese man of war, Physalia physalis, is a siphonophore that uses a gas-filled float as a sail to catch the wind. It is one of the most conspicuous, but poorly understood members of the pleuston, a community of organisms that occupy a habitat at the sea-air interface. The development, morphology, and colony organization of P. physalis is very different from all other siphonophores. Here, we propose a framework for homologizing the axes with other siphonophores, and also suggest that the tentacle bearing zooids should be called tentacular palpons. We also look at live and fixed larval and non-reproductively mature juvenile specimens, and use optical projection tomography to build on existing knowledge about the morphology and development of this species. Previous descriptions of P. physalis larvae, especially descriptions of budding order, were often framed with the mature colony in mind. However, we use the simpler organization of larvae and the juvenile specimens to inform our understanding of the morphology, budding order, and colony organization in the mature specimen. Finally, we review what is known about the ecology and lifecyle of P. physalis.

Time-dependent Canonical Correlation Analysis for Multilevel Time Series

Cao, X. — 2019-05-24

Canonical Correlation Analysis is a technique in multivariate data analysis for finding linear projections that maximize the correlation between two groups of variables. The correlations are typically defined without accounting for the serial correlations between observations, a typical setting for time series data. To understand the coupling dynamics and temporal variations between the two time-varying sources, we introduce the time-dependent canonical correlation analysis (TDCCA), a method for inferring time-dependent canonical vectors from multilevel time series data. A convex formulation of the problem is proposed, which leverages the singular value decomposition (SVD) characterization of all solutions of the CCA problem. We use simulated datasets to validate the proposed algorithm. Moreover, we propose a novel measure, canonical correlation variation as another way to assess the dynamic pattern of brain connections and we apply it to a real resting state fMRI dataset to study the aging effects on brain connectivity. Additionally, we explore our proposed method in a task-related fMRI to detect the temporal dynamics due to different motor tasks. We show that, compared to extant methods, the TDCCA-based approach not only detect temporal changes but also improves feature extraction. Together, this paper contributes broadly to new computational methodologies in understanding multilevel time series.

Extra-cellular matrix induced by steroids through a G-protein coupled receptor in a Drosophila model of renal fibrosis

Zheng, W. — 2019-05-29

Aldosterone is produced by the mammalian adrenal cortex to modulate blood pressure and fluid balance, however excessive, prolonged aldosterone production promotes fibrosis and kidney failure. How aldosterone triggers disease may involve actions that are independent of its canonical mineralocorticoid receptor. Here we present a Drosophila model of renal pathology caused by excess extra-cellular matrix formation, stimulated by exogenous aldosterone and insect ecdysone steroids. Chronic administration of aldosterone or ecdysone induces expression and accumulation of collagen-like pericardin at adult nephrocytes - podocyte-like cells that filter circulating hemolymph. Excess pericardin deposition disrupts nephrocyte (glomerular) filtration and causes proteinuria in Drosophila, hallmarks of mammalian kidney failure. Steroid-induced pericardin arises from cardiomyocytes associated with nephrocytes, reflecting an analogous role of mammalian myofibroblasts in fibrotic disease. Remarkably, the canonical ecdysteroid nuclear hormone receptor, ecdysone receptor EcR, is not required for aldosterone or ecdysone to stimulate pericardin production or associated renal pathology. Instead, these hormones require a cardiomyocyte-associated G-protein coupled receptor, dopamine-EcR (dopEcR), a membrane-associated receptor previously characterized in the fly brain as affecting behavior. This Drosophila renal disease model reveals a novel signaling pathway through which steroids may potentially modulate human fibrosis through proposed orthologs of dopEcR.nnSignificance StatementAldosterone regulates salt and fluid homeostasis, yet excess aldosterone contributes to renal fibrosis. Aldosterone acts through a nuclear hormone receptor, but an elusive, G-protein coupled receptor (GPCR) is thought to also mediate the hormones pathology. Here we introduce a Drosophila model of renal fibrosis. Flies treated with human aldosterone produce excess extra-cellular matrix and that causes kidney pathology. Flies treated with the insect steroid ecdysone produce similar pathology, and from this analogous response we identify an alternative receptor through which steroids mediate renal fibrosis -- the GPCR dopamine-Ecdysone Receptor (dopEcR). dopEcR functions in heart muscle cells associated with nephrocytes, analogous to the role of myofibroblasts in human fibrosis. This finding opens avenues to identify mammalian GPCR homologs of dopEcR through which aldosterone mediates renal fibrosis.

Reward Predictive Representations Generalize Across Tasks in Reinforcement Learning

Lehnert, L. — 2019-05-30

In computer science, reinforcement learning is a powerful framework with which artificial agents can learn to maximize their performance for any given Markov decision process (MDP). Advances over the last decade, in combination with deep neural networks, have enjoyed performance advantages over humans in many difficult task settings. However, such frameworks perform far less favorably when evaluated in their ability to generalize or transfer representations across different tasks. Existing algorithms that facilitate transfer typically are limited to cases in which the transition function or the optimal policy is portable to new contexts, but achieving "deep transfer" characteristic of human behavior has been elusive. Such transfer typically requires discovery of abstractions that permit analogical reuse of previously learned representations to superficially distinct tasks. Here, we demonstrate that abstractions that minimize error in predictions of reward outcomes generalize across tasks with different transition and reward functions. Such reward-predictive representations compress the state space of a task into a lower dimensional representation by combining states that are equivalent in terms of both the transition and reward functions. Because only state equivalences are considered, the resulting state representation is not tied to the transition and reward functions themselves and thus generalizes across tasks with different reward and transition functions. These results contrast with those using abstractions that myopically maximize reward in any given MDP and motivate further experiments in humans and animals to investigate if neural and cognitive systems involved in state representation perform abstractions that facilitate such equivalence relations. Author summaryHumans are capable of transferring abstract knowledge from one task to another. For example, in a right-hand-drive country, a driver has to use the right arm to operate the shifter. A driver who learned how to drive in a right-hand-drive country can adapt to operating a left-hand-drive car and use the other arm for shifting instead of re-learning how to drive. Despite the fact that both tasks require different coordination of motor skills, both tasks are the same in an abstract sense: In both tasks, a car is operated and there is the same progression from 1st to 2nd gear and so on. We study distinct algorithms by which a reinforcement learning agent can discover state representations that encode knowledge about a particular task, and evaluate how well they can generalize. Through a sequence of simulation results, we show that state abstractions that minimize errors in prediction about future reward outcomes generalize across tasks, even those that superficially differ in both the goals (rewards) and the transitions from one state to the next. This work motivates biological studies to determine if distinct circuits are adapted to maximize reward vs. to discover useful state representations.

Relating anthropometric indicators to brain structure in 2-month-old Bangladeshi infants growing up in poverty: a pilot study

Turesky, T. K. — 2019-05-31

Anthropometric indicators, including stunting, underweight, and wasting, have previously been associated with poor neurocognitive outcomes. This link may exist because malnutrition and infection, which are known to affect height and weight, also impact brain structure according to animal models. However, a relationship between anthropometric indicators and brain structural measures has not been tested yet, perhaps because stunting, underweight, and wasting are uncommon in higher-resource settings. Further, with diminished anthropomorphic growth prevalent in low-resource settings, where biological and psychosocial hazards are most severe, one might expect additional links between measures of poverty, anthropometry, and brain structure. To begin to examine these relationships, we conducted an MRI study in 2-3-month-old infants growing up in the extremely impoverished urban setting of Dhaka, Bangladesh. The sample size was relatively small because the challenges of investigating infant brain structure in a low-resource setting needed to be realized and resolved before introducing a larger cohort. Initially, fifty-four infants underwent T1 sequences using 3T MRI, and structural images were segmented into gray and white matter maps, which were carefully evaluated for accurate tissue labeling by a pediatric neuroradiologist. Gray and white matter volumes from 29 infants (79 {+/-} 10 days-of-age; F/M = 12/17), whose segmentations were of relatively high quality, were submitted to semi-partial correlation analyses with stunting, underweight, and wasting, which were measured using height-for-age (HAZ), weight-for-age (WAZ), and weight-for-height (WHZ) scores. Positive semi-partial correlations (after adjusting for chronological age and sex and correcting for multiple comparisons) were observed between white matter volume and HAZ and WAZ; however, WHZ was not correlated with any measure of brain volume. In examining the role of poverty, no associations were observed between income-to-needs or maternal education and brain volumetric measures, suggesting that risk factors previously linked with poverty were not associated with total brain tissue volume pre- or peri-natally in this sample. Overall, these results provide the first link between diminished anthropomorphic growth and white matter volume in infancy. Challenges of conducting a developmental neuroimaging study in a low-resource country are described.

The Impact of Antimalarial Resistance on the Genetic Structure of Plasmodium falciparum in the DRC

Verity, R. J. — 2019-05-31

The Democratic Republic of the Congo (DRC) harbors 11% of global malaria cases, yet little is known about the spatial and genetic structure of the parasite population in that country. We sequenced 2537 Plasmodium falciparum infections, including a nationally representative population sample from DRC and samples from surrounding countries, using molecular inversion probes - a novel high-throughput genotyping tool. We identified an east-west divide in haplotypes known to confer resistance to chloroquine and sulfadoxine-pyrimethamine. Furthermore, we identified highly related parasites over large geographic distances, indicative of gene flow and migration. Our results were consistent with a background of isolation by distance combined with the effects of selection for antimalarial drug resistance. This study provides a high-resolution view of parasite genetic structure across a large country in Africa and provides a baseline to study how implementation programs may impact parasite populations.

Layer 6 ensembles can selectively regulate the behavioral impact and layer-specific representation of sensory deviants

Voigts, J. — 2019-06-03

Predictive models can enhance the salience of unanticipated input, and the neocortical laminar architecture is believed to be central to this computation. Here, we examined the role of a key potential node in model formation, layer (L) 6, using behavioral, electrophysiological and imaging methods in mouse somatosensory cortex. To test the contribution of L6, we applied weak optogenetic drive that changed which L6 neurons were sensory-responsive, without affecting overall firing rates in L6 or L2/3. This stimulation suppressed L2/3 deviance encoding, but maintained other stimulus encoding. The stimulation also selectively suppressed behavioral sensitivity to deviant stimuli without impacting baseline performance. In contrast, stronger L6 drive inhibited firing and suppressed overall sensory function. These findings indicate that, despite their sparse activity, specific ensembles of stimulus-driven L6 neurons are required to form neocortical predictions, and for their behavioral benefit.

Allosteric Motions of the CRISPR-Cas9 HNH Nuclease Probed by NMR and Molecular Dynamics

East, K. W. — 2019-06-05

CRISPR-Cas9 is a widely employed genome-editing tool with functionality reliant on the ability of the Cas9 endonuclease to introduce site-specific breaks in double-stranded DNA. In this system, an intriguing allosteric communication has been suggested to control its DNA cleavage activity through flexibility of the catalytic HNH domain. Here, solution NMR experiments and a novel Gaussian accelerated Molecular Dynamics (GaMD) simulation method are used to capture the structural and dynamic determinants of allosteric signaling within the HNH domain. We reveal the existence of a millisecond timescale dynamic pathway that spans HNH from the region interfacing the adjacent RuvC nuclease and propagates up to the DNA recognition lobe in full-length CRISPR-Cas9. These findings reveal a potential route of signal transduction within the CRISPR-Cas9 HNH nuclease, advancing our understanding of the allosteric pathway of activation. Further, considering the role of allosteric signaling in the specificity of CRISPR-Cas9, this work poses the mechanistic basis for novel engineering efforts aimed at improving its genome editing capability.nnnnO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/660613v4_ufig1.gif" ALT="Figure 1">nView larger version (64K):norg.highwire.dtl.DTLVardef@13429c5org.highwire.dtl.DTLVardef@1a2493borg.highwire.dtl.DTLVardef@175e120org.highwire.dtl.DTLVardef@f1852_HPS_FORMAT_FIGEXP M_FIG C_FIG

Anatomical meniscus construct with zone specific biochemical composition and structural organization

Bahcecioglu, G. — 2019-06-10

A PCL/hydrogel construct that would mimic the structural organization, biochemistry and anatomy of meniscus was engineered. The compressive (380 {+/-} 40 kPa) and tensile modulus (18.2 {+/-} 0.9 MPa) of the PCL scaffolds were increased significantly when constructs were printed with a shifted design and circumferential strands mimicking the collagen organization in native tissue (p<0.05). Presence of circumferentially aligned PCL strands also led to elongation and alignment of the human fibrochondrocytes. Gene expression of the cells in agarose (Ag), gelatin methacrylate (GelMA), and GelMA-Ag hydrogels was significantly higher than that of cells on the PCL scaffolds after a 21-day culture. GelMA exhibited the highest level of collagen type I (COL1A2) mRNA expression, while GelMA-Ag exhibited the highest level of aggrecan (AGG) expression (p<0.001, compared to PCL). GelMA and GelMA-Ag exhibited a high level of collagen type II (COL2A1) expression (p<0.05, compared to PCL). Anatomical scaffolds with circumferential PCL strands were impregnated with cell-loaded GelMA in the periphery and GelMA-Ag in the inner region. GelMA and GelMA-Ag hydrogels enhanced the production of COL 1 and COL 2 proteins after a 6-week culture (p<0.05). COL 1 expression increased gradually towards the outer periphery, while COL 2 expression decreased. We were thus able to engineer an anatomical meniscus with a cartilage-like inner region and fibrocartilage-like outer region.

Epithelium intrinsic vitamin A signaling co-ordinates pathogen clearance in the gut via IL-18

Iyer, N. — 2019-06-12

Intestinal epithelial cells (IECs) are at the forefront of host-pathogen interactions, coordinating a cascade of immune responses to confer protection against pathogens. Here we show that IEC-intrinsic vitamin A signaling restricts pathogen invasion early in the infection and subsequently activates immune cells to promote pathogen clearance. Mice blocked for retinoic acid receptor (RAR) signaling selectively in IECs (stop{Delta}IEC) showed significantly higher Salmonella burden in colonic tissues early in the infection associated with higher luminal and systemic loads of the pathogen at later stages. Higher susceptibility of stop{Delta}IEC mice correlated with attenuated mucosal interferon gamma (IFN{gamma}) production by underlying immune cells. We found that, at homeostasis, the intestinal epithelium of stop{Delta}IEC mice produced significantly lower amounts of interleukin 18 (IL-18), a potent inducer of IFN{gamma}. Regulation of IL-18 by vitamin A was also observed in a dietary model of vitamin A supplementation. IL-18 reconstitution in stop{Delta}IEC mice restored resistance to Salmonella by promoting epithelial cell shedding to eliminate infected cells and limit pathogen invasion early in infection. Further, IL-18 augmented IFN{gamma} production by underlying immune cells to restrict pathogen burden and systemic spread. Our work uncovers a critical role for vitamin A in coordinating a biphasic immune response to Salmonella infection by regulating IL-18 production by IECs.nnAuthor SummaryEpithelial cells line the intestinal lumen, forming a barrier between the body and dietary and microbial contents in the lumen. Apart from absorbing nutrients from diet, these epithelial cells help mediate a stable, symbiotic relationship between microbes in the gut and the immune cells. During infection, they help co-ordinate the immune response to counter the infection. How dietary micronutrients, such as vitamin A, inform epithelial cell function during infection is poorly understood. Using a model where epithelial cells in the gut cannot respond to vitamin A signals, we find that epithelial vitamin A signaling promotes resistance to Salmonella infection. We show that, vitamin A increases the production of a key cytokine, interleukin 18, by epithelial cells. IL-18 promotes shedding of infected epithelial cells to reduce the pathogen invasion while also inducing the production of interferon gamma by immune cells to mediate pathogen clearance. Thus, epithelial cells dynamically respond to dietary vitamin A to regulate interleukin 18 production and potentiate resistance to infection.

Night watch during REM sleep for the first-night effect

Tamaki, M. — 2019-06-12

We experience disturbed sleep in a new place, and this effect is known as the first-night effect (FNE) in sleep research. We previously demonstrated that the FNE was associated with a protective night-watch system during NREM sleep in one hemisphere, which is shown as interhemispheric asymmetry in sleep depth in the default-mode network (DMN), and interhemispheric asymmetry in increased vigilance to monitor external stimuli. The present study investigated whether rapid eye movement (REM) sleep exhibited a form similar to a night-watch system during NREM sleep. First, we tested whether theta activity, which is an index of the depth of REM sleep, showed interhemispheric asymmetry in association with the FNE, by source-localizing to the DMN. However, interhemispheric asymmetry in theta activity during REM sleep was not found in association with the FNE. Next, we tested whether vigilance, as measured by evoked brain responses to deviant sounds, was increased in one hemisphere and showed interhemispheric asymmetry in association with the FNE during REM sleep. Because vigilance is different between the phasic period where rapid eye movements occur and the tonic period where rapid eye movements do not occur during REM sleep, REM sleep was split into phasic and tonic periods for measurements of evoked brain responses. While the evoked brain responses are generally small during the phasic period without the FNE, we found that the evoked brain response was significantly augmented by the FNE during the phasic period. In contrast, the evoked brain response during the tonic period did not differ by the presence of the FNE. Interhemispheric asymmetry in brain responses was not found during the phasic or tonic periods. These results suggest that a night-watch system for the FNE appears as interhemispheric asymmetry in sleep depth and vigilance during NREM sleep, but it appears as increased vigilance in both hemispheres during the phasic period, when vigilance to external stimuli is generally reduced without the FNE, during REM sleep. Therefore, a night-watch system associated with the FNE may be subserved by different neural mechanisms during NREM and REM sleep.

Targeting of the dosage-compensated male X-chromosome during early Drosophila development

Rieder, L. E. — 2019-06-14

The essential process of dosage compensation, which corrects for the imbalance in X-linked gene expression between XX females and XY males, represents a key model for how genes are targeted for coordinated regulation. However, the mechanism by which dosage compensation complexes identify the X-chromosome during early development remained unknown because of the difficulty of sexing embryos prior to zygotic transcription. We used meiotic drive to sex Drosophila embryos prior to zygotic transcription and ChIP-seq to measure dynamics of dosage compensation factor targeting. The Drosophila Male-Specific Lethal dosage compensation complex (MSLc) requires the ubiquitous zinc-finger protein Chromatin-Linked Adaptor for MSL Proteins (CLAMP) to identify the X-chromosome. We observe a multi-stage process in which MSLc first identifies CLAMP binding sites throughout the genome followed by concentration at the strongest X-linked MSLc sites. We provide insight into the dynamic mechanism by which a large transcription complex identifies its binding sites during early development.

A Bayesian framework for identifying consistent patterns of microbial abundance between body sites

Meier, R. — 2019-06-18

Recent studies have found that the microbiome in both gut and mouth are associated with diseases of the gut, including cancer. If resident microbes could be found to exhibit consistent patterns between the mouth and gut, disease status could potentially be assessed non-invasively through profiling of oral samples. Currently, there exists no generally applicable method to test for such associations. Here we present a Bayesian framework to identify microbes that exhibit consistent patterns between body sites, with respect to a phenotypic variable. For a given operational taxonomic unit (OTU), a Bayesian regression model is used to obtain Markov-Chain Monte Carlo estimates of abundance among strata, calculate a correlation statistic, and conduct a formal test based on its posterior distribution. Extensive simulation studies demonstrate overall viability of the approach, and provide information on what factors affect its performance. Applying our method to a dataset containing oral and gut microbiome samples from 77 pancreatic cancer patients revealed several OTUs exhibiting consistent patterns between gut and mouth with respect to disease subtype. Our method is well powered for modest sample sizes and moderate strength of association and can be flexibly extended to other research settings using any currently established Bayesian analysis programs.

In situ analysis of mucus residing bacterial community reveals an ecological niche key for gut microbiome stability

Duncan, K. — 2019-06-19

Mucus associated bacterial communities are critical for determining disease pathology and promoting colonization resistance. Yet the key ecological properties of mucus resident communities remain poorly defined. Using an approach that combines in situ hybridization, laser microdissection and 16s rRNA sequencing of spatially distinct regions of the mouse gut lumen, we discovered that a dense microbial community resembling a biofilm is embedded in the mucus layer. The mucus associated biofilm-like community excluded bacteria belonging to phylum Proteobacteria. Additionally, it was significantly more diverse and consisted of bacterial species that were unique to it. By employing germ-free mice deficient in T and B lymphocytes we found that formation of biofilm-like structure was independent of adaptive immunity. Instead the integrity of biofilm-like community depended on Gram-positive commensals such as Clostridia. Additionally, biofilm-like community in the mucus lost fewer Clostridia and showed smaller bloom of Proteobacteria compared to the lumen upon antibiotic treatment. When subjected to time restricted feeding biofilm like structure significantly enhanced in size and showed enrichment of Clostridia. Taken together our work discloses that mucus associated biofilm-like community represent a specialized community that is structurally and compositionally distinct that excludes aerobic bacteria while enriching for anaerobic bacteria such as Clostridia, exhibits enhanced stability to antibiotic treatment and that can be modulated by dietary changes.

NMRdock: Lightweight and Modular NMR Processing

East, K. W. — 2019-06-22

NMR is a widely employed tool in chemistry, biology, and physics for the study of molecular structure and dynamics. Advances in computation have produced scores of software programs necessary for the processing and analysis of NMR data. However, the production of NMR software has been largely overseen by academic labs, each with their own preferred OS, environment, and dependencies. This lack of broader standardization and the complexity of installing and maintaining NMR-related software creates a barrier of entry into the field. To further complicate matters, as computation evolves, many aging software packages become deprecated. To reduce the barrier for newcomers and to prevent deprecation of aging software, we have created the NMRdock container. NMRdock utilizes containerization to package NMR processing and analysis programs into a single, easy-to-install Docker image that can be run on any modern OS. The current image contains two bedrock NMR data processing programs (NMRPipe and NMRFAM Sparky). However, future development of NMRdock aims to add modules for additional analysis programs to build a library of tools in a standardized and easy-to-implement manner. NMRdock is open source and free to download at https://compbiocore.github.io/nmrdock/.

Epstein Barr virus genomes reveal population structure and type 1 association with endemic Burkitt lymphoma

Kaymaz, Y. — 2019-07-02

Endemic Burkitt lymphoma (eBL), the most prevalent pediatric cancer in sub-Saharan Africa, is associated with malaria and Epstein Barr virus (EBV). In order to better understand the role of EBV in eBL, we improved viral DNA enrichment methods and generated a total of 98 new EBV genomes from both eBL cases (N=58) and healthy controls (N=40) residing in the same geographic region in Kenya. Comparing cases and controls, we found that EBV type 1 was significantly associated with eBL with 74.5% of patients (41/55) versus 47.5% of healthy children (19/40) carrying type 1 (OR=3.24, 95% CI=1.36 - 7.71, P=0.007). Controlling for EBV type, we also performed a genome-wide association study identifying 6 nonsynonymous variants in the genes EBNA1, EBNA2, BcLF1, and BARF1 that were enriched in eBL patients. Additionally, we observed that viruses isolated from plasma of eBL patients were identical to their tumor counterpart consistent with circulating viral DNA originating from the tumor. We also detected three intertypic recombinants carrying type 1 EBNA2 and type 2 EBNA3 regions as well as one novel genome with a 20 kb deletion resulting in the loss of multiple lytic and virion genes. Comparing EBV types, genes show differential variation rates as type 1 appears to be more divergent. Besides, type 2 demonstrates novel substructures. Overall, our findings address the complexities of EBV population structure and provide new insight into viral variation, which has the potential to influence eBL oncogenesis.nnKey PointsO_LIEBV type 1 is more prevalent in eBL patients compared to the geographically matched healthy control group.nC_LIO_LIGenome-wide association analysis between cases and controls identifies 6 eBL-associated nonsynonymous variants in EBNA1, EBNA2, BcLF1, and BARF1 genes.nC_LIO_LIAnalysis of population structure reveals that EBV type 2 exists as two genomic sub groups.nC_LI

The Bric-a-Brac transcription factors are necessary for formation of functional germline stem cell niches through control of dpp expression in the Drosophila melanogaster ovary

Miscopein Saler, L. — 2019-07-03

Many studies have focused on the mechanisms of stem cell maintenance via their interaction with a particular niche or microenvironment in adult tissues, but how formation of a functional niche is initiated, including how stem cells within a niche are established, is less well understood. Adult Drosophila melanogaster ovary Germline Stem Cell (GSC) niches are comprised of somatic cells forming a stack called a Terminal Filament (TF) and underlying Cap Cells (CCs) and Escort Cells (ECs), which are in direct contact with GSCs. In the adult, the Engrailed (En) transcription factor is specifically expressed in niche cells where it directly controls expression of the decapentaplegic gene (dpp) encoding a member of the Bone Morphogenetic Protein (BMP) family of secreted signaling molecules, which are key factors for GSC maintenance. In late third instar larval ovaries, in response to BMP signaling from newly-formed niches, adjacent primordial germ cells become GSCs. The bric-a-brac paralogs (bab1 and bab2) encode BTB/POZ-domain containing transcription factors, that are also expressed in developing GSCs niches where they are required for TF formation. Here, we demonstrate that Bab1 and Bab2 display redundant cell autonomous function for TF morphogenesis and we identify a new function for these genes in GSC establishment. Moreover, we show that Bab proteins control dpp expression in otherwise correctly specified CCs, independently of En and its paralog Invected (Inv). In fact, our results also indicate that en/inv function in larval stages are neither essential for TF formation, nor GSC establishment. Finally, when bab2 was overexpressed in ovarian somatic cells outside of the niche, where en/inv were not expressed, ectopic BMP signaling activation was induced in adjacent germ cells of adult ovaries, which formed GSC-like tumors. Together, these results indicate that Bab transcription factors are positive regulators of BMP signaling for acquisition of GSC status.

A missense variant in CREBRF is associated with taller stature in Samoans

Carlson, J. C. — 2019-07-02

ObjectivesStudies have demonstrated that rs373863828, a missense mutation in CREBRF, is associated with a number of anthropometric traits including body mass index (BMI), obesity, percent body fat, hip circumference, and abdominal circumference. Given the biological relationship between height and adiposity, we hypothesized that the effect of this variant on BMI might be due in part to a previously untested association of this variant with height.nnMethodsWe tested the hypothesis that minor allele of rs373863828 is associated with height in a Samoan population in two adult cohorts and in a separate cohort of children (age 5 - 18 years old) using linear mixed modeling.nnResultsWe found evidence of a strong relationship between rs373863828 and greater mean height in Samoan adults (0.77 cm greater average height for each copy of the minor allele) with the same direction of effect in Samoan children.nnConclusionsThese results suggest that the missense variant rs373863828 in CREBRF, first identified through an association with larger BMI, may be related to an underlying biological mechanism affecting overall body size including stature.

Automated data extraction from historical city directories the rise and fall of mid-century gas stations in Providence, RI

Bell, S. — 2019-07-12

The location of defunct environmentally hazardous businesses like gas stations has many implications for modern American cities. To track down these locations, we present the directoreadr code (github.com/brown-ccv/directoreadr). Using scans of Polk city directories from Providence, RI, directoreadr extracts and parses business location data with a high degree of accuracy. The image processing pipeline ran without any human input for 94.4% of the pages we examined. For the remaining 5.6%, we processed them with some human input. Through hand-checking a sample of three years, we estimate that ~94.6% of historical gas stations are correctly identified and located, with historical street changes and non-standard address formats being the main drivers of errors. As an example use, we look at gas stations, finding that gas stations were most common early in the study period in 1936, beginning a sharp and steady decline around 1950. We are making the dataset produced by directoreadr publicly available. We hope it will be used to explore a range of important questions about socioeconomic patterns in Providence and cities like it during the transformations of the mid-1900s.

SINATRA: A Sub-Image Analysis Pipeline for Selecting Features that Differentiate Classes of 3D Shapes

Wang, B. — 2019-07-14

The recent curation of large-scale databases with 3D surface scans of shapes has motivated the development of tools that better detect global patterns in morphological variation. Studies which focus on identifying differences between shapes have been limited to simple pairwise comparisons and rely on pre-specified landmarks (that are often known). We present SINATRA: the first statistical pipeline for analyzing collections of shapes without requiring any correspondences. Our novel algorithm takes in two classes of shapes and highlights the physical features that best describe the variation between them. We use a rigorous simulation framework to assess our approach. Lastly, as a case study, we use SINATRA to analyze mandibular molars from four different suborders of primates and demonstrate its ability recover known morphometric variation across phylogenies.

The BioLuminescent-OptoGenetic in vivo Response to Coelenterazine is Proportional, Sensitive and Specific in Neocortex

Gomez-Ramirez, M. — 2019-07-22

BioLuminescent (BL) light production can modulate neural activity and behavior through coexpressed OptoGenetic (OG) elements, an approach termed BL-OG. Yet, the relationship between BL-OG effects and bioluminescent photon emission has not been characterized in vivo. Further, the degree to which BL-OG effects strictly depend on optogenetic mechanisms driven by bioluminescent photons is unknown. Crucial to every neuromodulation method is whether the activator shows a dynamic concentration range driving robust, selective, and non-toxic effects. We systematically tested the effects of four key components of the BL-OG mechanism (luciferin, oxidized luciferin, luciferin vehicle, and bioluminescence), and compared these against effects induced by the Luminopsin-3 (LMO3) BL-OG molecule, a fusion of slow burn Gaussia luciferase (sbGLuc) and Volvox ChannelRhodopsin-1 (VChR1). We performed combined bioluminescence imaging and electrophysiological recordings while injecting specific doses of Coelenterazine (substrate for sbGluc), Coelenteramide (CTM, the oxidized product of CTZ), or CTZ vehicle. CTZ robustly drove activity in mice expressing LMO3, with photon production proportional to firing rate. In contrast, low and moderate doses of CTZ, CTM, or vehicle did not modulate activity in mice that did not express LMO3. We also failed to find bioluminescence effects on neural activity in mice expressing an optogenetically non-sensitive LMO3 variant. We observed weak responses to the highest dose of CTZ in control mice, but these effects were significantly smaller than those observed in the LMO3 group. These results show that in neocortex in vivo, there is a large CTZ range wherein BL-OG effects are specific to its active chemogenetic mechanism.

Neural systems for memory-based value judgment and decision-making

Vaidya, A. R. — 2019-07-24

Real life choices often require that we draw inferences about the value of options based on structured, schematic knowledge about their utility for our current goals. Other times, value information may be retrieved directly from a specific prior experience with an option. In a functional magnetic resonance imaging (fMRI) experiment, we investigated the neural systems involved in retrieving and assessing information from different memory sources to support value-based choice. Participants completed a task in which items could be conferred positive or negative value based on schematic associations (i.e. schema value), or learned directly from experience via deterministic feedback (i.e. experienced value). We found that ventromedial prefrontal cortex (vmPFC) activity correlated with the influence of both experience- and schema-based values on participants decisions. Connectivity between vmPFC and middle temporal cortex also tracked the inferred value of items based on schematic associations on the first presentation of ingredients, prior to any feedback. In contrast, the striatum responded to participants willingness to bet on ingredients as a function of the unsigned strength of their memory for those options values. These results argue that striatum and vmPFC play distinct roles in memory-based value judgment and decision-making. Specifically, the vmPFC assesses the value of options based on information inferred from schematic knowledge and retrieved from prior direct experience, while the striatum controls a decision to act on options based on memory strength.

Genomic Influences on Self-Reported Childhood Maltreatment

Dalvie, S. — 2019-07-28

Childhood maltreatment is highly prevalent and serves as a risk factor for mental and physical disorders. Self-reported childhood maltreatment appears heritable, but the specific genetic influences on this phenotype are largely unknown. The aims of this study were to 1) identify genetic variation associated with reported childhood maltreatment, 2) calculate the relevant SNP-based heritability estimates, and 3) quantify the genetic overlap of reported childhood maltreatment with mental and physical health-related phenotypes. Genome-wide association analysis for childhood maltreatment was undertaken, using a discovery sample from the UK Biobank (UKBB) (n=124,000) and a replication sample from the Psychiatric Genomics Consortium-posttraumatic stress disorder working group (PGC-PTSD) (n=26,290). Heritability estimations for childhood maltreatment and genetic correlations with mental/physical health traits were calculated using linkage disequilibrium score regression (LDSR). Two genome-wide significant loci associated with childhood maltreatment, located on chromosomes 3p13 (rs142346759, beta=0.015, p=4.35x10-8, FOXP1) and 7q31.1 (rs10262462, beta=-0.016, p=3.24x10-8, FOXP2), were identified in the discovery dataset but were not replicated in the PGC-PTSD sample. SNP-based heritability for childhood maltreatment was estimated to be [~]6%. Childhood maltreatment was most significantly genetically correlated with depressive symptoms (rg=0.70, p=4.65x10-40). This is the first large-scale genetic study to identify specific variants associated with self-reported childhood maltreatment. FOXP genes could influence traits such as depression and thereby be relevant to childhood maltreatment. Alternatively, these variants may be associated with a greater likelihood of reporting maltreatment. A clearer understanding of the genetic relationships of childhood maltreatment, including particular abuse subtypes, with various psychiatric disorders, may ultimately be useful in in developing targeted treatment and prevention strategies.

Age-dependent effects of reduced mTor signalling on life expectancy through distinct physiology

Simons, M. J. P. — 2019-07-30

Research on the mechanisms of ageing has identified ways via which lifespan can be extended in model organisms, increasing the potential for translation of these findings to our own species. However, the large majority of research on animal models involves dietary, genetic or pharmacological treatments throughout life - limiting translational potential and ignoring age-dependent effects. Previously, we have suggested using demographic meta-analysis that reduced mTor signalling has the potential to instantly rejuvenate. We have now tested this prediction experimentally using large-scale demographic data (N > 10,000) combined with conditional knockdown of mTor in Drosophila melanogaster. Indeed, reduced mTor decreased mortality rate when applied during old age. Interestingly, we found that transient treatment during early adult life had long-lasting benefits. Age-dependent deep-RNAseq indicated that these effects arose from distinct physiology and implicate alternative splicing as a potential mechanism for the long-lasting benefits of transient mTor reduction. These findings suggest that reducing mTor short term or during old age could be used to combat ageing. In addition, our findings suggest that the results from experimental research on mTor signalling, and potentially other mechanisms of ageing, that employ life-long interventions are likely to be a complex composite of age-dependent effects that counteract or enhance each other.

Small molecules for modulating protein driven liquid-liquid phase separation in treating neurodegenerative disease

Wheeler, R. J. — 2019-08-05

Neurodegenerave diseases such as amyotrophic lateral sclerosis (ALS) are oten associated with mutaons in proteins that are associated with stress granules. Stress granules are condensates formed by liquid-liquid phase separaon which, when aberrant, can lead to altered condensaon behaviours and disease phenotypes. Here, we identified lipoamide, a small molecule which specifically prevents cytoplasmic condensaon of stress granule proteins. Thermal proteome profiling showed that lipoamide preferentially stabilises intrinsically disordered domain-containing proteins. These include SRSF1 and SFPQ, stress granule proteins necessary for lipoamide activity. The redox state of SFPQ correlates with its condensate-dissolving behaviour, in concordance with the importance of the dithiolane ring for lipoamide activity. In animals, lipoamide ameliorates aging-associated aggregaon of a stress granule reporter, improves neuronal morphology, and recovers motor defects caused by expression of ALS-associated FUS and TDP-43 mutants. In conclusion, lipoamide is a well-tolerated small molecule modulator of stress granule condensaon and dissection of its molecular mechanism identified a cellular pathway for redox regulaon of stress granule formaon.

Individual differences in model-based planning are linked to the ability to infer latent structure

Rmus, M. — 2019-08-02

Humans appear to represent many forms of knowledge in associative networks whose nodes are multiply connected, including sensory, spatial, and semantic. Recent work has shown that explicitly augmenting artificial agents with such graph-structured representations endows them with more human-like capabilities of compositionality and transfer learning. An open question is how humans acquire these representations. Previously, it has been shown that humans can learn to navigate graph-structured conceptual spaces on the basis of direct experience with trajectories that intentionally draw the network contours (Schapiro et al., 2012;2016), or through direct experience with rewards that covary with the underlying associative distance (Wu et al., 2018). Here, we provide initial evidence that this capability is more general, extending to learning to reason about shortest-path distances across a graph structure acquired across disjoint experiences with randomized edges of the graph - a form of latent learning. In other words, we show that humans can infer graph structures, assembling them from disordered experiences. We further show that the degree to which individuals learn to reason correctly and with reference to the structure of the graph corresponds to their propensity, in a separate task, to use model-based reinforcement learning to achieve rewards. This connection suggests that the correct acquisition of graph-structured relationships is a central ability underlying forward planning and reasoning, and may be a core computation across the many domains in which graph-based reasoning is advantageous.

Curated Multiple Sequence Alignment for the Adenomatous Polyposis Coli (APC) Gene and Accuracy of In Silico Pathogenicity Predictions

Karabachev, A. D. — 2019-08-02

Computational algorithms are often used to assess pathogenicity of Variants of Uncertain Significance (VUS) that are found in disease-associated genes. Most computational methods include analysis of protein multiple sequence alignments (PMSA), assessing interspecies variation. Careful validation of PMSA-based methods has been done for relatively few genes, partially because creation of curated PMSAs is labor-intensive. We assessed how PMSA-based computational tools predict the effects of the missense changes in the APC gene, in which pathogenic variants cause Familial Adenomatous Polyposis. Most Pathogenic or Likely Pathogenic APC variants are protein-truncating changes. However, public databases now contain thousands of variants reported as missense. We created a curated APC PMSA that contained >3 substitutions/site, which is large enough for statistically robust in silico analysis. The creation of the PMSA was not easily automated, requiring significant querying and computational analysis of protein and genome sequences. Of 1924 missense APC variants in the NCBI ClinVar database, 1800 (93.5%) are reported as VUS. All but two missense variants listed as P/LP occur at canonical splice or Exonic Splice Enhancer sites. Pathogenicity predictions by five computational tools (Align-GVGD, SIFT, PolyPhen2, MAPP, REVEL) differed widely in their predictions of Pathogenic/Likely Pathogenic (range 17.5-75.0%) and Benign/Likely Benign (range 25.0-82.5%) for APC missense variants in ClinVar. When applied to 21 missense variants reported in ClinVar as Benign, the five methods ranged in accuracy from 76.2-100%. Computational PMSA-based methods can be an excellent classifier for variants of some hereditary cancer genes. However, there may be characteristics of the APC gene and protein that confound the results of in silico algorithms. A systematic study of these features could greatly improve the automation of alignment-based techniques and the use of predictive algorithms in hereditary cancer genes.nnAuthor SummaryA critical problem in clinical genetics today is interpreting whether a genetic variant is benign or causes disease (pathogenic). Some of the hardest variants to interpret are those that change one amino acid for another in a protein sequence (a "missense variant"). Various computer programs are often used to predict whether mutations in disease-associated genes likely cause disease. Most computer programs involve studying how the gene has changed during evolution, comparing the protein sequences of different species by aligning them with each other. Variants in amino acids that have not tolerated mutation during evolution are usually predicted to be pathogenic, and variants in amino acids that have tolerated variation are usually predicted to be benign. High quality alignments are necessary to make accurate predictions. However, creating high quality alignments is difficult, not easily automated, and requires significant manual curation. Results from computer-generated predictions are used in current published guidelines as one tool for evaluating whether variants will disrupt the protein function and cause disease. These guidelines may be applied to genes in which single amino acid substitutions do not commonly cause disease. One such example is the APC gene, which is responsible for Familial Adenomatous Polyposis (FAP). Missense APC changes are not a common cause of FAP. Our analysis of APC demonstrated the difficulty of generating an accurate protein sequence alignment and the tendency of computer tools to overestimate the damaging effects of amino acid substitutions. Our results suggest that the rules for using computer-based tools to predict whether a variant causes disease should be modified when applied to genes in which missense variants rarely cause disease.

Biological Pathways and Gene Networks Link Inflammation and Vascular Remodeling to Both Heart Failure with Preserved and Reduced Ejection Fraction in Women across Ethnicities

Liu, Q. — 2019-08-06

IntroductionHeart failure (HF) is understudied among women; especially, genomic evidence implicating shared or unique mechanisms of HF with respect to reduced or preserved ejection fraction (HFrEF, HFpEF) is lacking across ethnic populations of women. Prior genome-wide association studies (GWAS) have identified approximately 30 suggestive genetic variants for HF, although none have been specifically linked to HFrEF or HFpEF.nnObjectivesWe aimed to define, replicate, and annotate genetic variants to HFrEF, HFpEF, or both, as well as to investigate potential biological mechanisms underlying HFrEF and HFpEF among African American (AA) and European American (EA) women in three well-characterized, high-quality prospective cohorts, the Womens Health Initiative (WHI) study, the Jackson Heart Study (JHS), and the Framingham Heart Study (FHS).nnMethodsGWAS analysis on HFrEF and HFpEF were first performed among 7,982 AA and 4,133 EA in the WHI, followed by pathway analysis employing two independent methodological platforms (GSA-SNP and Mergeomics) curating KEGG, Reactome, and BioCarta pathway databases. GWAS signals and biological pathways identified using the WHI were replicated in the JHS and FHS. For all replicated pathways, we performed cross-phenotype and cross-ethnicity validation analyses to examine shared pathways between HFrEF and HFpEF, and phenotype-specific pathways, across ethnicities. We further prioritized key driver genes for HF according to specific pathways identified.nnResultsWe validated one previously reported genetic locus and identified six new ones, among which one locus was allocated to HFrEF and five to HFpEF. Additionally, we defined five biological pathways shared between HFrEF and HFpEF and discovered six HFpEF-specific pathways. These pathways overlapped in two main domains for molecular signaling: 1) inflammation and 2) vascular remodeling (including angiogenesis and vascular patterning), involving key driver genes from collagen and HLA gene families.nnConclusionsOur network analysis of three large prospective cohorts of women in the United States defined several novel loci for HF and its subtypes. In particular, several key driver genes reinforce the mechanistic role of inflammation and vascular remodeling in the development of HF, especially HFpEF. Given that therapeutic strategies developed for left ventricular dysfunction have had limited success for HFpEF, several new targets and pathways identified and validated in this study should be further assessed in risk stratification as well as the design of potential new HF interventions.

Dopamine waves as a mechanism for spatiotemporal credit assignment

Hamid, A. A. — 2019-08-13

Significant evidence supports the view that dopamine shapes reward-learning by encoding prediction errors. However, it is unknown whether dopamine decision-signals are tailored to the functional specialization of target regions. Here, we report a novel set of wave-like spatiotemporal activity-patterns in dopamine axons across the dorsal striatum. These waves switch between different activational motifs and organize dopamine transients into localized clusters within functionally related striatal subregions. These specific motifs are associated with distinct task contexts: At reward delivery, dopamine signals rapidly resynchronize into propagating waves with opponent directions depending on instrumental task contingencies. Moreover, dopamine dynamics during reward pursuit signal the extent to which mice have instrumental control and interact with reward waves to predict future behavioral adjustments. Our results are consistent with a computational architecture in which striatal dopamine signals are sculpted by inference about instrumental controllability and provide evidence for a spatiotemporally "vectorized" role of dopamine in credit assignment.

Automated and rapid self-report of nociception in transgenic mice.

Black, C. J. — 2019-08-12

A time-resolved, conscious report of detected nociceptive stimuli in mice offers an opportunity to examine the relationship between higher order neural circuits and pain perception. We have developed a detection behavior in transgenic mice that leverages temporally precise and cell-specific stimulation to elicit self-reports of nociception. Conscious reporting of peripheral nociceptive input may help identify neural mechanisms that generate pain perception.

Selection on mutators is not frequency-dependent

Raynes, Y. — 2019-08-15

The evolutionary fate of mutator mutations - i.e., genetic variants that raise the genome-wide mutation rate - in asexual populations is often described as being frequency (or number) dependent. This common intuition suggests that mutators can invade a population by hitchhiking with a sweeping beneficial mutation, but only when sufficiently frequent to produce such a mutation before non-mutators do. Here, we use stochastic, agent-based simulations to show that neither the strength nor the sign of selection on mutators depend on their initial frequency, and while the overall probability of hitchhiking increases predictably with frequency, the per-capita probability of fixation remains unchanged.

Opponent neurochemical and functional processing in NREM and REM sleep in visual learning

Tamaki, M. — 2019-08-19

Sleep is beneficial for learning. However, whether NREM or REM sleep facilitates learning, whether the learning facilitation results from plasticity increases or stabilization and whether the facilitation results from learning-specific processing are all controversial. Here, after training on a visual task we measured the excitatory and inhibitory neurochemical (E/I) balance, an index of plasticity in human visual areas, for the first time, while subjects slept. Off-line performance gains of presleep learning were associated with the E/I balance increase during NREM sleep, which also occurred without presleep training. In contrast, increased stabilization was associated with decreased E/I balance during REM sleep only after presleep training. These indicate that the above-mentioned issues are not matters of controversy but reflect opposite neurochemical processing for different roles in learning during different sleep stages: NREM sleep increases plasticity leading to performance gains independently of learning, while REM sleep decreases plasticity to stabilize learning in a learning-specific manner.

Reward does not facilitate visual perceptual learning until sleep occurs

Tamaki, M. — 2019-08-19

A growing body of evidence indicates that visual perceptual learning (VPL) is enhanced by reward provided during training. Another line of studies has shown that sleep following training also plays a role in facilitating VPL, an effect known as the offline performance gain of VPL. However, whether the effects of reward and sleep interact on VPL remains unclear. Here, we show that reward interacts with sleep to facilitate offline performance gains of VPL. First, we demonstrated a significantly larger offline performance gain over a 12-h interval including sleep in a reward group than that in a No-reward group. However, the offline performance gains over the 12-h interval without sleep were not significantly different with or without reward during training, indicating a crucial interaction between reward and sleep in VPL. Next, we tested whether neural activations during posttraining sleep were modulated after reward was provided during training. Reward provided during training enhanced REM sleep time, increased oscillatory activities for reward processing in the prefrontal region during REM sleep, and inhibited neural activation in the untrained region in early visual areas in NREM and REM sleep. The offline performance gains were significantly correlated with oscillatory activities of visual processing during NREM sleep and reward processing during REM sleep in the reward group but not in the No-reward group. These results suggest that reward provided during training becomes effective during sleep, with excited reward processing sending inhibitory signals to suppress noise in visual processing, resulting in larger offline performance gains over sleep.nnSignificance statementIndependent lines of research have shown that visual perceptual learning (VPL) is improved by reward or sleep. Here, we show that reward provided during training increased offline performance gains of VPL over sleep. Moreover, during posttraining sleep, reward was associated with longer REM sleep, increased activity in reward processing in the prefrontal region during REM sleep, and decreased activity in the untrained region of early visual areas during NREM and REM sleep. Offline performance gains were correlated with modulated oscillatory activity in reward processing during REM sleep and visual processing during NREM sleep. These results suggest that reward provided during training becomes effective on VPL through the interaction between reward and visual processing during sleep after training.

Human Neocortical Neurosolver (HNN): A new software tool for interpreting the cellular and network origin of human MEG/EEG data

Neymotin, S. A. — 2019-08-20

Magneto- and electro-encephalography (MEG/EEG) non-invasively record human brain activity with millisecond resolution providing reliable markers of healthy and disease states. Relating these macroscopic signals to underlying cellular- and circuit-level generators is a limitation that constrains using MEG/EEG to reveal novel principles of information processing or to translate findings into new therapies for neuropathology. To address this problem, we built Human Neocortical Neurosolver (HNN, https://hnn.brown.edu) software. HNN has a graphical user interface designed to help researchers and clinicians interpret the neural origins of MEG/EEG. HNNs core is a neocortical circuit model that accounts for biophysical origins of electrical currents generating MEG/EEG. Data can be directly compared to simulated signals and parameters easily manipulated to develop/test hypotheses on a signals origin. Tutorials teach users to simulate commonly measured signals, including event related potentials and brain rhythms. HNNs ability to associate signals across scales makes it a unique tool for translational neuroscience research.

Skeletal muscles that actuate sexual displays are specialized for de novo androgen synthesis

Schuppe, E. — 2019-08-20

The gonads (testes) act as the primary organ where androgenic hormones are made to regulate reproductive behavior in male vertebrates. Yet many endocrinologists have also long suspected that other tissues in the body can autonomously synthesize their own androgens to support behavioral output. We examine this idea here by studying whether avian skeletal muscles that actuate elaborate socio-sexual displays are specialized to maintain the molecular machinery otherwise needed for de novo androgen synthesis. Our results show that the vocal organ, or syrinx (SYR), of two songbirds species does in fact express all transporters and enzymes necessary to create androgenic hormones from scratch. This includes genes that encode proteins to mediate rate-limiting steps of steroidogenesis, which are seldom found outside of the gonads. We also show that expression levels of these genes are far greater in the SYR than non-display muscles, matching expression levels found in another extra-gonadal site of steroidogenesis--the brain. Furthermore, we uncover a nearly identical gene expression signature in a woodpecker neck muscle, the longus colli ventralis (LC). This tissue actuates the birds social drum display, which serves the same functions as song. This same study also demonstrates that the elevated expression of steroidogenic genes persists in this neck muscle year-round, suggesting that the LCs capacity to make androgens is a constitutive trait. Altogether, our findings suggest that muscles involved in sexual display is uniquely specialized to locally make steroid hormones, likely supporting their own role in behavioral production.

Quantifying the cost of cognitive stability and flexibility

Papadopetraki, D. — 2019-08-29

Exerting cognitive control is known to carry a subjective effort cost and people are generally biased to avoid it. Recent theorizing suggests that the cost of cognitive effort serves as a motivational signal to bias people away from excessive focusing and towards more cognitive flexibility. We asked whether the effort cost of stable distractor resistance is higher than that of flexible updating of working memory representations. We tested this prediction by using (i) a delayed response paradigm in which we manipulate demands for distractor resistance and flexible updating, as well as (ii) a subsequent cognitive effort discounting paradigm that allows us to quantify subjective effort costs. We demonstrate, in two different samples (28 and 62 participants) that participants discount tasks both high in distractor resistance and flexible updating when comparing with taking a break. As predicted, when directly contrasting distractor resistance and flexible updating the subjective cost of performing a task requiring distractor resistance is higher than that requiring flexible updating.

Novel genetic determinants of telomere length from a multi-ethnic analysis of 75,000 whole genome sequences in TOPMed

Taub, M. A. — 2019-09-04

Telomeres shorten in replicating somatic cells, and telomere length (TL) is associated with age-related diseases 1,2. To date, 17 genome-wide association studies (GWAS) have identified 25 loci for leukocyte TL 3-19, but were limited to European and Asian ancestry individuals and relied on laboratory assays of TL. In this study from the NHLBI Trans-Omics for Precision Medicine (TOPMed) program, we used whole genome sequencing (WGS) of whole blood for variant genotype calling and the bioinformatic estimation of TL in n=109,122 trans-ethnic (European, African, Asian and Hispanic/Latino) individuals. We identified 59 sentinel variants (p-value <5x10-9) from 36 loci (20 novel, 13 replicated in external datasets). There was little evidence of effect heterogeneity across populations, and 10 loci had >1 independent signal. Fine-mapping at OBFC1 indicated the independent signals colocalized with cell-type specific eQTLs for OBFC1 (STN1). We further identified two novel genes, DCLRE1B (SNM1B) and PARN, using a multi-variant gene-based approach.

A supragranular nexus for the effects of neocortical beta events on human tactile perception

Law, R. G. — 2019-09-02

Transient neocortical events with high spectral power in the 15-29Hz beta band are among the most reliable predictors of sensory perception: High prestimulus beta event rates in primary somatosensory lead to sensory suppression, most effective at 100-300ms prestimulus latency. However, the synaptic and neuronal mechanisms inducing betas perceptual effects have not been completely localized. We combined human MEG with neural modeling designed to account for these macroscale signals to interpret the cellular and circuit mechanisms that underlie the influence of beta on tactile detection. Extending prior studies, we modeled the hypothesis that higher-order thalamic bursts, sufficient for beta event generation in cortex, recruit supragranular GABAB inhibition acting on a 300ms time scale to suppress sensory information. Consistency between model and MEG data supported this hypothesis and led to a further prediction, validated in our data, that stimuli are perceived when beta events occur simultaneously with tactile stimulation. The post-event suppressive mechanism explains an array of studies that associate beta with decreased processing, while the during-event mechanism may demand a reinterpretation of the role of beta events in the context of coincident timing.nnSignificance statementSomatosensory beta events - transient 15-29Hz oscillations in electromagnetic recordings - are thought to be generated when "top-down" bursts of spikes presumably originating in higher-order thalamus arrive in upper layers of somatosensory cortex. Physiological evidence had shown that the immediate action of these top-down projections should be excitatory; however, after a beta event, sensory perception is noticeably inhibited for approximately 300ms. The source of this post-event sensory suppression, in particular, had been unresolved. Using a detailed computational model of somatosensory cortex, we find evidence for the hypothesis that these bursts couple indirectly to GABAB inhibition in upper layers of cortex, and that beta events first briefly disinhibit sensory relay before a longer period of inhibition.

Perceptual learning of lesions in mammograms induced by response feedback during training

Frank, S. M. — 2019-09-02

Detecting subtle lesions in mammograms indicative of early breast cancer usually requires years of experience. Well-designed training paradigms could be a strong tool for promoting perceptual learning (PL) with rapid and long-lasting improvement in detectability of these subtle mammographic lesions. Given that PL occurs without feedback about the accuracy of subjects responses, the role of feedback has been completely ignored in clinical applications of PL. However, in this study, we found that the contents of the feedback profoundly and differentially influence the formation and retention of PL to detect calcification and architectural distortion lesions, two types of mammographic lesions that are frequently missed in mammographic screenings. We trained subjects to detect one type of lesion in a mammogram and manipulated the content of the response feedback during training for 3 groups (no feedback, correctness only, and both correctness and location of the lesion). We found that PL occurred for both lesions when both correctness and location feedback were provided. PL also occurred for calcifications but not for distortions when only correctness was provided. No learning occurred without feedback for either lesion. A retest conducted six months later showed that PL was retained only in the group with both correctness and location feedback for both types of lesions. In contrast to the general consensus of basic PL studies, our results demonstrate that the content of the response feedback is a determining factor in forming and retaining PL to detect mammographic lesions.

Alcohol causes lasting differential transcription in Drosophila mushroom body neurons

Petruccelli, E. — 2019-08-30

Repeated alcohol experiences can produce long-lasting memories for sensory cues associated with intoxication. These memories can ultimately trigger relapse in individuals recovering from alcohol use disorder (AUD). The molecular mechanisms by which alcohol changes memories to become long-lasting and inflexible remain unclear. New methods to analyze gene expression within precise neuronal cell-types can provide further insight towards AUD prevention and treatment. Here, we employed genetic tools in Drosophila melanogaster to investigate the lasting consequences of ethanol on transcription in memory-encoding neurons. Drosophila rely on mushroom body (MB) neurons to make associative memories, including memories of ethanol-associated sensory cues. Differential expression analyses found that distinct transcripts, but not genes, in the MB were associated with experiencing ethanol alone compared to forming a memory of an odor cue associated with ethanol. These findings reveal the dynamic and highly context-specific regulation of splicing associated with encoding behavioral experiences. Our data thus demonstrate that alcohol can have lasting effects on transcription and RNA processing during memory formation, and identify new transcript targets for future AUD and addiction investigation.

Bacterial Hyperswarming as a Protective Response to Intestinal Stress

Chen, W. — 2019-09-08

Bacterial swarming, a collective movement on a surface, has rarely been associated with human pathophysiology. Here, we report for the first time that bacterial swarmers are associated with protection against intestinal inflammation. We show that bacterial swarmers are highly predictive of intestinal stress in mice and humans. We isolated a novel Enterobacter swarming strain, SM3, from mouse feces. SM3 and other known commensal swarmers contrast to their respective swarming-deficient, but swimming-competent isogenic strains abrogated intestinal inflammation in mice. Treatment of colitic mice with SM3, but not its mutants, enriched beneficial fecal anaerobes belonging to the family, Bacteroidales S24-7. We observed SM3 swarming associated pathways in the in vivo fecal metatranscriptomes. In vitro growth of S24-7 was enriched in presence of SM3 or its mutants conjecturing that bacterial swarming in vivo might influence SM3s access to S24-7 in the intestines. Overall, our work identifies a new paradigm in which intestinal stress allows for the emergence of swarming bacteria, which can counterintuitively heal intestinal inflammation.

Subjective value, not a grid-like code, describes neural activity in ventromedial prefrontal cortex during value-based decision-making

Lee, S. — 2019-09-08

Across many studies, ventromedial prefrontal cortex (vmPFC) activity has been found to correlate with subjective value during value-based decision-making. Recently, however, vmPFC has also been shown to reflect a hexagonal gridlike code during navigation through physical and conceptual space. This raises the possibility that the subjective value correlates previously observed in vmPFC may have actually been a misconstrued gridlike signal. Here, we first show that, in theory, a hexagonal gridlike code of two-dimensional attribute space could mimic vmPFC activity previously attributed to subjective value. However, using fMRI data from a large number of subjects performing an intertemporal choice task, we show clear and unambiguous evidence that subjective value is a better description of vmPFC activity than a hexagonal gridlike code. In fact, we find no significant evidence at all for a hexagonal gridlike code in vmPFC activity during intertemporal choice. This result limits the generality of gridlike modulation as description of vmPFC activity. We suggest that vmPFC may flexibly switch representational schemes so as to encode the most relevant information for the current task.

Reward evokes visual perceptual learning following reinforcement rules

Wang, Z. — 2019-09-08

Visual perceptual learning (VPL) is defined as a long-term performance enhancement as a result of visual experiences. A number of studies have demonstrated that reward can evoke VPL. However, the mechanisms of how reward evoke VPL remain unknown. One possible hypothesis is that VPL is obtained through reward related reinforcement processing. If this hypothesis is true, learning can only occur when reward follows the stimulus presentation. Another interpretation is that VPL is acquired through an enhancement of alertness in association with reward. If the alertness hypothesis is true, learning should occur when reward precedes the stimulus presentation. In our study, we tested the plausibility of the two hypotheses by manipulating the order of reward and stimulus presentation. In Experiment 1, we separated participants into two groups. During training, the Before group received water reward 400ms prior to the onset of trained orientation stimulus while the After group received water reward 400ms subsequent to the onset of trained orientation stimulus. Both groups were trained using the Continuous Flash Suppression paradigm to render the stimulus imperceptible to the participants by the presentation of dynamic noise in the untrained eye. We found training only in the After group indicating that reward may evoke learning through reinforcement-like processing. In Experiment 2, we excluded the possibility that alertness may not be sufficient to elicit learning when presented before stimulus. We presented beep sound prior to the onset of stimulus to increase alertness. Our finding demonstrated that alertness is sufficient enough to evoke learning. In conclusion, our study provided evidence that reward can evoke VPL through reinforcement process.

The \"Locus of Learning\" Problem: Effects of Stimulus and Task Structure on Temporal Perceptual Learning

Xu, R. — 2019-09-08

The ability to discriminate sub-second intervals can be improved with practice, a process known as temporal perceptual learning (TPL). A central question in TPL is whether training improves the low-level sensory representation of a temporal interval or optimizes a set of task-specific response strategies. Here, we trained three groups of participants over five days on a single-interval temporal discrimination task using either fixed intervals (FI) or random intervals (RI). Before and after training, discrimination thresholds were also obtained on an untrained task. Our results revealed that only the FI group showed improvements with five days of training, but this learning did not generalize from the trained task to the untrained task in any group. These results highlight task-specificity in TPL and suggest that training-dependent improvements in timing ability might reflect an active reweighting of decision units, in addition to refinements in the sensory representation of a learned interval.

Dopamine Promotes Cognitive Effort by Biasing the Benefits Versus Costs of Cognitive Work

Westbrook, A. — 2019-09-23

Stimulants like methylphenidate are increasingly used for cognitive enhancement, but precise mechanisms are unknown. We found that methylphenidate boosts willingness to expend cognitive effort by altering the benefit-to-cost ratio of cognitive work. Willingness to expend effort was greater for participants with higher striatal dopamine synthesis capacity, while methylphenidate and sulpiride - a selective D2 receptor antagonist - increased cognitive motivation more for participants with lower synthesis capacity. A sequential sampling model informed by momentary gaze revealed that decisions to expend effort are related to amplification of benefit-versus-cost information attended early in the decision process, while the effect of benefits is strengthened with higher synthesis capacity and by methylphenidate. These findings demonstrate that methylphenidate boosts the perceived benefits-versus-costs of cognitive effort by modulating striatal dopamine signaling. One Sentence SummaryStriatal dopamine increases cognitive effort by respectively amplifying and attenuating the subjective benefits and costs of cognitive control.

Consumption of a Western-style diet modulates the response of the murine gut microbiome to ciprofloxacin

Cabral, D. J. — 2019-09-24

Dietary composition and antibiotic use are known to have major impacts on the structure and function of the gut microbiome, often resulting in dysbiosis. Despite this, little research has been done to explore the role of host diet as a determinant of antibiotic-induced microbiome disruption. Here, we utilize a multi-omic approach to characterize the impact of Western-style diet consumption on ciprofloxacin-induced changes to gut microbiome community structure and transcriptional activity. We found that mice consuming a Western-style diet experienced a greater expansion of Firmicutes following ciprofloxacin treatment than those eating a control diet. At the transcriptional level, we found that ciprofloxacin induced a reduction in the abundance of TCA cycle transcripts on both diets, suggesting that carbon metabolism plays a key role in the response of the gut microbiome to this antibiotic. Despite this shared response, we observed extensive differences in the response of the microbiota to ciprofloxacin on each diet. In particular, at the whole-community level we detected an increase in starch degradation, glycolysis, and pyruvate fermentation following antibiotic treatment in mice on the Western diet, which we did not observe in mice on the control diet. Similarly, we observed diet-specific changes in the transcriptional activity of two important commensal bacteria, Akkermansia muciniphila and Bacteroides thetaiotaomicron, involving diverse cellular processes such as nutrient acquisition, stress responses, and capsular polysaccharide (CPS) biosynthesis. These findings demonstrate that host diet plays a key role in determining the extent of disruption of microbiome composition and function induced by antibiotic treatment. ImportanceWhile both diet and antibiotics are individually known to have profound impacts on gut microbiome composition, little work has been done to examine the effect of these two factors combined. A number of negative health outcomes, including diabetes and obesity, are associated with diets high in simple sugars in fats but low in host-indigestible fiber, and some of these outcomes may be mediated by the gut microbiome. Likewise, treatment with broad-spectrum antibiotics and the resulting dysbiosis is associated with many of the same detrimental side effects. Previous work has shown that nutrient availability, as influenced by host diet, plays an important role in determining the extent of antibiotic-induced disruption to the gut microbiome. Due to the growing incidence of disorders related to antibiotic-induced dysbiosis, it is essential to determine how the prevalence of high fat and sugar "Western"-style diets impacts the response of the microbiome to antibiotics.

United States wildlife and wildlife product imports from 2000-2014

Eskew, E. A. — 2019-09-24

The global wildlife trade network is a massive system that has been shown to threaten biodiversity conservation, introduce non-native species and pathogens, and cause chronic animal welfare concerns. Despite its scale and impact, comprehensive characterization of the global wildlife trade is hampered by data that are limited in their temporal or taxonomic scope and detail. To help fill this gap, we present data on 15 years of the importation of wildlife and their derived products into the United States (2000-2014), originally collected by the United States Fish and Wildlife Service. We curated and cleaned the data and added taxonomic information to improve data usability. These data include > 2 million wildlife or wildlife product shipments, representing > 60 biological classes and > 3.2 billion live organisms. These data will be broadly useful to both scientists and policymakers seeking to better understand the volume, sources, biological composition, and potential risks of the global wildlife trade.

Early developmental exposure to Fluoxetine and Citalopram results in different neurodevelopmental outcomes

Liu, K. — 2019-09-24

Although selective serotonin reuptake inhibitors (SSRIs) are commonly prescribed for prenatal depression, there exists controversy over the adverse effects of SSRI use on fetal development. Few studies have adequately isolated outcomes due to SSRI exposure and those due to maternal psychiatric conditions. Here, we directly investigated the outcomes of exposure to widely-used SSRIs fluoxetine and citalopram on the developing nervous system of Xenopus laevis tadpoles, using an integrative experimental approach. We exposed tadpoles to low doses of citalopram and fluoxetine during a critical developmental period and found that the different groups of tadpoles displayed opposing behavioral effects. While both groups showed reduced schooling behavior, the fluoxetine group showed increased seizure susceptibility and reduced startle habituation. In contrast, the citalopram treated tadpoles had decreased seizure susceptibility and increased habituation. Both groups had abnormal dendritic morphology in the optic tectum, a brain area important for all three behaviors tested. Whole-cell electrophysiological recordings of tectal neurons showed no differences in synaptic function across groups; however, tectal cells from fluoxetine-treated tadpoles had decreased voltage gated K+ currents while cells in the citalopram group had increased K+ currents. Both the behavior and electrophysiological findings indicate that cells and circuits in the fluoxetine treated optic tecta are hyperexcitable, while the citalopram group exhibits decreased excitability. Taken all together, these results show that early developmental exposure to SSRIs is sufficient to induce neurodevelopmental effects, however these effects can be complex and vary depending on the SSRI used. This may explain some of the discrepancies across human studies, and further underscores the importance of serotonergic signaling for the developing nervous system.

Platinum chemotherapy induces lymphangiogenesis to prime tissues for tumor metastasis

Harris, A. R. — 2019-10-07

Chemotherapy has been used to inhibit cancer growth for decades, but emerging evidence shows it can affect the tumor stroma unintentionally promoting cancer malignancy. After treatment of primary tumors, remaining drugs drain via lymphatics. Though all drugs interact with the lymphatics, we know little of their impact on them. Here, we show a previously unknown effect of platinums, a widely used class of chemotherapeutics, to directly induce systemic lymphangiogenesis and activation. These changes are dose-dependent, long-lasting, and occur in healthy and cancerous tissue in multiple mouse models of breast cancer. We saw similar effects in human ovarian and breast cancer patients whose treatment regimens included platinums. Carboplatin treatment of healthy mice prior to mammary tumor inoculation increases cancer metastasis as compared to no pre-treatment. These platinum-induced phenomena could be blocked by VEGFR3 inhibition. These findings have implications for cancer patients receiving platinums and may support the inclusion of anti-VEGFR3 therapy into treatment regimens or differential design of treatment regimens to alter these potential effects. SummaryPlatinum chemotherapy induces VEGFR3-dependent lymphangiogenesis, priming tissues for metastasis of breast cancer. Inhibition of VEGFR3 via antibody blockade can reverse these effects.

Inherited Causes of Clonal Hematopoiesis of Indeterminate Potential in TOPMed Whole Genomes

Bick, A. G. — 2019-09-27

Age is the dominant risk factor for most chronic human diseases; yet the mechanisms by which aging confers this risk are largely unknown.1 Recently, the age-related acquisition of somatic mutations in regenerating hematopoietic stem cell populations was associated with both hematologic cancer incidence2-4 and coronary heart disease prevalence.5 Somatic mutations with leukemogenic potential may confer selective cellular advantages leading to clonal expansion, a phenomenon termed Clonal Hematopoiesis of Indeterminate Potential (CHIP).6 Simultaneous germline and somatic whole genome sequence analysis now provides the opportunity to identify root causes of CHIP. Here, we analyze high-coverage whole genome sequences from 97,691 participants of diverse ancestries in the NHLBI TOPMed program and identify 4,229 individuals with CHIP. We identify associations with blood cell, lipid, and inflammatory traits specific to different CHIP genes. Association of a genome-wide set of germline genetic variants identified three genetic loci associated with CHIP status, including one locus at TET2 that was African ancestry specific. In silico-informed in vitro evaluation of the TET2 germline locus identified a causal variant that disrupts a TET2 distal enhancer. Aggregates of rare germline loss-of-function variants in CHEK2, a DNA damage repair gene, predisposed to CHIP acquisition. Overall, we observe that germline genetic variation altering hematopoietic stem cell function and the fidelity of DNA-damage repair increase the likelihood of somatic mutations leading to CHIP.

Temporal discounting correlates with directed exploration but not with random exploration

Sadeghiyeh, H. — 2019-09-26

The explore-exploit dilemma describes the trade off that occurs any time we must choose between exploring unknown options and exploiting options we know well. Implicit in this trade off is how we value future rewards -- exploiting is usually better in the short term, but in the longer term the benefits of exploration can be huge. Thus, in theory there should be a tight connection between how much people value future rewards, i.e. how much they discount future rewards relative to immediate rewards, and how likely they are to explore, with less temporal discounting associated with more exploration. By measuring individual differences in temporal discounting and correlating them with explore-exploit behavior, we tested whether this theoretical prediction holds in practice. We used the 27-item Delay-Discounting Questionnaire to estimate temporal discounting and the Horizon Task to quantify two strategies of explore-exploit behavior: directed exploration, where information drives exploration by choice, and random exploration, where behavioral variability drives exploration by chance. We find a clear correlation between temporal discounting and directed exploration, with more temporal discounting leading to less directed exploration. Conversely, we find no relationship between temporal discounting and random exploration. Unexpectedly, we find that the relationship with directed exploration appears to be driven by a correlation between temporal discounting and uncertainty seeking at short time horizons, rather than information seeking at long horizons. Taken together our results suggest a nuanced relationship between temporal discounting and explore-exploit behavior that may be mediated by multiple factors.

Antimicrobial resistance gene levels are independent of levofloxacin administration but related to abundance of specific pathobionts in dementia patients

Rowan-Nash, A. D. — 2019-10-05

BackgroundThe issue of antimicrobial resistance continues to grow worldwide, and long-term care facilities are significant reservoirs of antimicrobial-resistant organisms, in part due to high frequency of antimicrobial use. Patients with advanced dementia are particularly vulnerable to multidrug-resistant organism acquisition and antimicrobial overuse, which has negative consequences for the gut microbiome and can contribute to the selection and propagation of antimicrobial resistance genes. In this study, we longitudinally examined a group of advanced dementia patients treated with the fluoroquinolone antimicrobial levofloxacin, finding a correlation between abundance of pathogens and antimicrobial resistance genes, which we confirmed in a larger cohort of subjects with advanced dementia.nnResultsWe observed significant inter- and intra-subject heterogeneity in the composition of the microbiota of the longitudinal levofloxacin cohort, suggesting temporal instability. Within this dataset, we did not find significant impacts of levofloxacin on the diversity, composition, function, or resistome of the gut microbiota of this population. However, we were able to link the antimicrobial resistance gene burden in a sample with the relative abundance of several pathobionts - particularly Escherichia coli, Proteus mirabilis, and Enterococcus faecalis, as well as less-prevalent species including Providencia stuartii and Staphylococcus haemolyticus. Furthermore, we used metagenomic assembly and binning to demonstrate that these species had higher genomic resistance gene levels than common gut commensals, and we were able to predict antimicrobial resistance gene burden from the relative abundances of these species in a separate, larger cohort from the same population.nnConclusionsWe found that the relative abundances of several pathobionts were correlated with and were even predictive of the level of antimicrobial resistance genes in corresponding samples, and that these species carried high levels of resistances genes in their assembled genomes. In order to test this observation, we utilized a larger metagenomics dataset from a similar population and confirmed the association between pathobiont abundance and antimicrobial resistance genes. Given the high frequency with which these species were found at high levels in this population and the underlying vulnerability to infection with multidrug resistant organisms of advanced dementia patients, attention to microbial blooms of these species may be warranted. Additionally, in this study, we were able to utilize genomic assembly from metagenomic data to more definitively associate antimicrobial resistance gene levels with specific assembled species; as this technology continues to develop, assembly could prove to be a valuable method to monitor both specific resistance genes and blooms of multidrug-resistant organisms.

Cathepsin G Degrades Synovial Fluid Lubricin: Relevance For Osteoarthritis Pathogenesis

Huang, S. — 2019-10-15

Lubricin (PRG4) is a mucin type protein that plays an important role in maintaining normal joint function by providing lubrication and chondroprotection. Improper lubricin modification and degradation has been observed in idiopathic osteoarthritis (OA), while the detailed mechanism still remains unknown. We hypothesized that the protease cathepsin G (CG) may participate in degrading lubricin in synovial fluid (SF). The presence of endogenous CG in SF was confirmed in 16 patients with knee OA. Recombinant human lubricin (rhPRG4) and native lubricin purified from the SF of patients were incubated with exogenous CG and lubricin degradation was monitored using western blot, staining by Coomassie or Periodic Acid-Schiff in gels, and with proteomics. Full length lubricin ([~]300 kDa), was efficiently digested with CG generating a 25-kDa protein fragment, originating from the densely glycosylated mucin domain ([~]250 kDa). The 25-kDa fragment was present in the SF from OA patients, and the amount was increased after incubation with CG. A CG digest of rhPRG4 revealed 135 peptides and 72 glycopeptides, and confirmed that the protease could cleave in different domains of lubricin. Our results suggest that synovial CG may take part in the degradation of lubricin, which could affect the lubrication of OA joints.

Disrupted Autophagy and Neuronal Dysfunction in C. elegans Knockin Models of FUS Amyotrophic Lateral Sclerosis

Baskoylu, S. — 2019-10-11

It remains unclear how mutations in FUS, a ribonucleoprotein, lead to neuronal dysfunction in Amyotrophic Lateral Sclerosis (ALS) patients. To examine mechanisms underlying ALS FUS dysfunction, we generated the first C. elegans knock-in models using CRISPR/Cas9-mediated genome editing, creating R524S and P525L ALS FUS models. Although FUS inclusions were not detected, ALS FUS animals showed defective neuromuscular function, as well as stress-induced locomotion defects. Unlike C. elegans lacking the endogenous FUS ortholog, ALS FUS animals had impaired neuronal autophagy and increased SQST-1 accumulation in ALS FUS motor neurons. Loss of sqst-1, the C. elegans ortholog for ALS-linked, autophagy adaptor protein SQSTM1/p62, suppressed both neuromuscular and stress-induced locomotion defects in ALS FUS animals, but did not suppress neuronal autophagy defects. Therefore, autophagy dysfunction is upstream of, and not dependent on, SQSTM1 function in ALS FUS pathogenesis. Combined, our findings demonstrate that autophagy dysfunction likely contributes to protein homeostasis and neuromuscular defects in ALS FUS knock-in animals.

Functional brain network reconfiguration during learning in a dynamic environment

Kao, C.-H. — 2019-10-10

When learning about dynamic and uncertain environments, people should update their beliefs most strongly when new evidence is most informative, such as when the environment undergoes a surprising change or existing beliefs are highly uncertain. Here we show that modulations of surprise and uncertainty are encoded in a particular, temporally dynamic pattern of whole-brain functional connectivity, and this encoding is enhanced in individuals that adapt their learning dynamics more appropriately in response to these factors. The key feature of this whole-brain pattern of functional connectivity is stronger connectivity, or functional integration, between the fronto-parietal and other functional systems. Our results provide new insights regarding the association between dynamic adjustments in learning and dynamic, large-scale changes in functional connectivity across the brain.

DNA Break-Induced Epigenetic Drift as a Cause of Mammalian Aging

Hayano, M. — 2019-10-21

There are numerous hallmarks of aging in mammals, but no unifying cause has been identified. In budding yeast, aging is associated with a loss of epigenetic information that occurs in response to genome instability, particularly DNA double-strand breaks (DSBs). Mammals also undergo predictable epigenetic changes with age, including alterations to DNA methylation patterns that serve as epigenetic "age" clocks, but what drives these changes is not known. Using a transgenic mouse system called "ICE" (for inducible changes to the epigenome), we show that a tissues response to non-mutagenic DSBs reorganizes the epigenome and accelerates physiological, cognitive, and molecular changes normally seen in older mice, including advancement of the epigenetic clock. These findings implicate DSB-induced epigenetic drift as a conserved cause of aging from yeast to mammals.nnOne Sentence SummaryDNA breaks induce epigenomic changes that accelerate the aging clock in mammals

Mechanophenotyping of 3D Multicellular Clusters using Displacement Arrays of Rendered Tractions

Leggett, S. E. — 2019-10-21

Epithelial tissues mechanically deform the surrounding extracellular matrix during embryonic development, wound repair, and tumor invasion. Ex vivo measurements of such multicellular tractions within three-dimensional (3D) biomaterials could elucidate collective dissemination during disease progression, and enable preclinical testing of targeted anti-migration therapies. However, past 3D traction measurements have been low throughput due to the challenges of imaging and analyzing information-rich 3D material deformations. Here, we demonstrate a method to profile multicellular clusters in a 96-well plate format based on spatially heterogeneous contractile, protrusive, and circumferential tractions. As a case study, we profile multicellular clusters across varying states of the epithelial-mesenchymal transition, revealing a successive loss of protrusive and circumferential tractions, as well as the formation of localized contractile tractions with elongated cluster morphologies. These cluster phenotypes were biochemically perturbed using drugs, biasing towards traction signatures of different epithelial or mesenchymal states. This higher-throughput analysis is promising to systematically interrogate and perturb aberrant mechanobiology, which could be utilized with human patient samples to guide personalized therapies.

Analysis of unusual and signature APOBEC-mutations in HIV-1 pol next-generation sequences

Tzou, P. L. — 2019-11-05

IntroductionAt low mutation-detection thresholds, next generation sequencing (NGS) for HIV-1 genotypic resistance testing is susceptible to artifactual detection of mutations arising from PCR error and APOBEC-mediated G-to-A hypermutation.nnMethodsWe analyzed published HIV-1 pol Illumina NGS data to characterize the distribution of mutations at eight NGS thresholds: 20%, 10%, 5%, 2%, 1%, 0.5%, 0.2%, and 0.1%. At each threshold, we determined the proportion of amino acid mutations that were unusual (defined as having a prevalence <0.01% in HIV-1 group M sequences) or were signature APOBEC mutations.nnResultsEight studies, containing 855 samples, in the NCBI Sequence Read Archive were analyzed. As detection thresholds were lowered, there was a progressive increase in the proportion of positions with both usual and unusual mutations and in the proportion of all mutations that were unusual. The median proportion of positions with an unusual mutation increased gradually from 0% at the 20% threshold to 0.3% at the 1% threshold and then exponentially to 1.3% (0.5% threshold), 6.9% (0.2% threshold), and 23.2% (0.1% threshold). In two of three studies with available plasma HIV-1 RNA levels, the proportion of positions with unusual mutations was negatively associated with virus levels. Although the complete set of signature APOBEC mutations was much smaller than that of unusual mutations, the former outnumbered the latter in one-sixth of the samples at the 0.5%, 1%, and 2% thresholds.nnConclusionsThe marked increase in the proportion of positions with unusual mutations at thresholds below 1% and in samples with lower virus loads suggests that, at low thresholds, many unusual mutations are artifactual, reflecting PCR error or G-to-A hypermutation. Profiling the numbers of unusual and signature APOBEC pol mutations at different NGS thresholds may be useful to avoid selecting a threshold that is too low and poses an unacceptable risk of identifying artifactual mutations.

Single-cell tumor phylogeny inference with copy-number constrained mutation losses

Satas, G. — 2019-11-26

MotivationSingle-cell DNA sequencing enables the measurement of somatic mutations in individual tumor cells, and provides data to reconstruct the evolutionary history of the tumor. Nearly all existing methods to construct phylogenetic trees from single-cell sequencing data use single-nucleotide variants (SNVs) as markers. However, most solid tumors contain copy-number aberrations (CNAs) which can overlap loci containing SNVs. Particularly problematic are CNAs that delete an SNV, thus returning the SNV locus to the unmutated state. Such mutation losses are allowed in some models of SNV evolution, but these models are generally too permissive, allowing mutation losses without evidence of a CNA overlapping the locus. ResultsWe introduce a novel loss-supported evolutionary model, a generalization of the infinite sites and Dollo models, that constrains mutation losses to loci with evidence of a decrease in copy number. We design a new algorithm, Single-Cell Algorithm for Reconstructing the Loss-supported Evolution of Tumors (SO_SCPLOWCARLETC_SCPLOW), that infers phylogenies from single-cell tumor sequencing data using the loss-supported model and a probabilistic model of sequencing errors and allele dropout. On simulated data, we show that SO_SCPLOWCARLETC_SCPLOW outperforms current single-cell phylogeny methods, recovering more accurate trees and correcting errors in SNV data. On single-cell sequencing data from a metastatic colorectal cancer patient, SO_SCPLOWCARLETC_SCPLOW constructs a phylogeny that is both more consistent with the observed copy-number data and also reveals a simpler monooclonal seeding of the metastasis, contrasting with published reports of polyclonal seeding in this patient. SO_SCPLOWCARLETC_SCPLOW substantially improves single-cell phylogeny inference in tumors with CNAs, yielding new insights into the analysis of tumor evolution. AvailabilitySoftware is available at github.com/raphael-group/scarlet Contactbraphael@princeton.edu

WormCat: an online tool for annotation and visualization of Caenorhabditis elegans genome-scale data

Holdorf, A. D. — 2019-11-15

The emergence of large gene expression datasets has revealed the need for improved tools to identify enriched gene categories and visualize enrichment patterns. While Gene Ontogeny (GO) provides a valuable tool for gene set enrichment analysis, it has several limitations. First, it is difficult to graphically compare multiple GO analyses. Second, genes from some model systems are not well represented. For example, around 30% of Caenorhabditis elegans genes are missing from analysis in commonly used databases. To allow categorization and visualization of enriched C. elegans gene sets in different types of genome-scale data, we developed WormCat, a web-based tool that uses a near-complete annotation of the C. elegans genome to identify co-expressed gene sets and scaled heat map for enrichment visualization. We tested the performance of WormCat using a variety of published transcriptomic datasets and show that it reproduces major categories identified by GO. Importantly, we also found previously unidentified categories that are informative for interpreting phenotypes or predicting biological function. For example, we analyzed published RNA-seq data from C. elegans treated with combinations of lifespan-extending drugs where one combination paradoxically shortened lifespan. Using WormCat, we identified sterol metabolism as a category that was not enriched in the single or double combinations but emerged in a triple combination along with the lifespan shortening. Thus, WormCat identified a gene set with potential phenotypic relevance that was not uncovered with previous GO analysis. In conclusion, WormCat provides a powerful tool for the analysis and visualization of gene set enrichment in different types of C. elegans datasets.

Breast tumor stiffness instructs bone metastasis via mechanical memory

Watson, A. W. — 2019-11-20

The mechanical microenvironment of primary breast tumors plays a substantial role in promoting tumor progression. While the transitory response of cancer cells to pathological stiffness in their native microenvironment has been well described, it is still unclear how mechanical stimuli in the primary tumor influence distant, late-stage metastatic phenotypes across time and space in absentia. Here, we show that primary tumor stiffness promotes stable, non-genetically heritable phenotypes in breast cancer cells. This mechanical memory instructs cancer cells to adopt and maintain increased cytoskeletal dynamics, traction force, and 3D invasion in vitro, in addition to promoting osteolytic bone metastasis in vivo. Furthermore, we established a mechanical conditioning (MeCo) score comprised of mechanically-regulated genes as a global gene expression measurement of tumor stiffness response. Clinically, we show that a high MeCo score is strongly associated with bone metastasis in patients. Using a discovery approach, we mechanistically traced mechanical memory in part to ERK-mediated mechanotransductive activation of RUNX2, an osteogenic gene bookmarker and bone metastasis driver. The combination of these RUNX2 traits permits the stable transactivation of osteolytic target genes that remain upregulated after cancer cells disseminate from their activating microenvironment in order to modify a distant microenvironment. Using genetic, epigenetic, and functional approaches, we were able to simulate, repress, select and extend RUNX2-mediated mechanical memory and alter cancer cell behavior accordingly. In concert with previous studies detailing the influence of biochemical properties of the primary tumor stroma on distinct metastatic phenotypes, our findings detailing the influence of biomechanical properties support a generalized model of cancer progression in which the integrated properties of the primary tumor microenvironment govern the secondary tumor microenvironment, i.e., soil instructs soil.

Adolescent substance use and functional connectivity between the ventral striatum and hippocampus

Huntley, E. D. — 2019-11-20

Neurodevelopmental explanations for adolescent substance use have focused on heightened sensitivity of the brains reward system, centered around the ventral striatum (VS). Recent evidence demonstrates increased functional connectivity between the VS and hippocampus in adolescents relative to adults, suggesting that the adolescent brain may learn from subsequent exposure to risks/rewards. However, a link between VS-hippocampal circuitry and adolescent substance use has yet to be established. Two separate longitudinal studies were conducted to evaluate whether variation in VS-hippocampal resting-state functional connectivity (rs-FC) predicts subsequent adolescent substance use. Study 1 consisted of 19 youth recruited from a high sociodemographic risk population (N = 19; 14 female; 47% Black Non-Hispanic, 32% White Non-Hispanic). To replicate results of Study 1, Study 2 utilized data from the National Consortium on Adolescent Neurodevelopment and Alcohol, an ongoing multi-site imaging study (N= 644; 339 female; 11% Black Non-Hispanic, 11% Hispanic/Latino, 66% White Non-Hispanic). Resting-state fMRI data were collected at a baseline time point and lifetime and past year self-reported substance use was collected at a follow up visit. Regression models tested whether baseline VS-hippocampal rs-FC predicted substance use at follow up. Across both studies, higher VS-hippocampal rs-FC at baseline predicted greater substance use at follow up. These data provide the first evidence linking increased VS-hippocampal connectivity with greater adolescent substance use. Results fit with the emerging idea that adolescent substance use is driven by not only a heightened sensitivity to reward, but also a stronger link between reinforcement learning and episodic memory for rewarding outcomes.

Context-dependent role of vinculin in neutrophil adhesion, motility and trafficking

Wilson, Z. S. — 2019-11-20

Neutrophils are innate immune effector cells that traffic from the peripheral blood to extravascular sites of inflammation. {beta}2 integrins are involved during multiple phases of neutrophil recruitment, including the transition from rolling to arrest, firm attachment and motility within the vasculature. Following neutrophil arrest, adhesion stabilization occurs as the neutrophil interacts with the endothelial surface and crawls into a favorable position for extravasation. The cytoskeletal protein vinculin has been implicated in other cell types as a regulator of adhesion strength by promoting focal adhesion maturation and as a sensor of the mechanical properties of the microenvironment. Neutrophils express vinculin but do not form mature focal adhesions. Here, we characterize the role of vinculin in {beta}2 integrin-dependent neutrophil adhesion, motility, mechanosensing, and recruitment. We observe that knockout of vinculin attenuates, but does not completely abrogate, neutrophil adhesion, spreading, and crawling under static conditions. In the presence of forces from fluid flow, vinculin was not required for neutrophil adhesion or migration. Vinculin deficiency only mildly attenuated neutrophil traction stresses and spreading on stiff, but not soft, polyacrylamide gels indicating a minor role for vinculin in the mechanosensing of the neutrophil as compared to slower moving mesenchymal cells that form mature focal adhesions. Consistent with these findings, we observe in vivo neutrophil recruitment into the inflamed peritoneum of mice remains intact in the absence of vinculin. Together, these data suggest that while vinculin regulates some aspects of neutrophil adhesion and spreading, it may be dispensable for neutrophil recruitment and motility in vivo.

Falciparum malaria from coastal Tanzania and Zanzibar remains highly connected despite effective control efforts on the archipelago

Morgan, A. P. — 2019-12-03

BackgroundTanzanias Zanzibar archipelago has made significant gains in malaria control over the last decade and is a target for malaria elimination. Despite consistent implementation of effective tools since 2002, elimination has not been achieved. Importation of parasites from outside of the archipelago is thought to be an important cause of malarias persistence, but this paradigm has not been studied using modern genetic tools. MethodsWe used whole-genome sequencing (WGS) to investigate the impact of importation, employing population genetic analyses of Plasmodium falciparum isolates from both the archipelago and mainland Tanzania. We assessed ancestry, levels of genetic diversity and differentiation, patterns of relatedness, and patterns of selection between these two populations by leveraging recent advances in deconvolution of genomes from polyclonal malaria infections. ResultsWe identified significant decreases in the effective population sizes in both populations in the timeframe of decreasing malaria transmission in Tanzania. Identity by descent analysis showed that parasites in the two populations shared large sections of their genomes, on the order of 5 cM, suggesting shared ancestry within the last 10 generations. Even with limited sampling,, we demonstrate a pair of isolates between the mainland and Zanzibar that are related at the expected level of half-siblings, consistent with recent importation ConclusionsThese findings suggest that importation plays an increasing role for malaria incidence on Zanzibar and demonstrate the value of genomic approaches for identifying corridors of parasite movement to the island.

Systems biology informed deep learning for inferring parameters and hidden dynamics

Yazdani, A. — 2019-12-04

Mathematical models of biological reactions at the system-level lead to a set of ordinary differential equations with many unknown parameters that need to be inferred using relatively few experimental measurements. Having a reliable and robust algorithm for parameter inference and prediction of the hidden dynamics has been one of the core subjects in systems biology, and is the focus of this study. We have developed a new systems-biology-informed deep learning algorithm that incorporates the system of ordinary differential equations into the neural networks. Enforcing these equations effectively adds constraints to the optimization procedure that manifests itself as an imposed structure on the observational data. Using few scattered and noisy measurements, we are able to infer the dynamics of unobserved species, external forcing, and the unknown model parameters. We have successfully tested the algorithm for three different benchmark problems. Author summaryThe dynamics of systems biological processes are usually modeled using ordinary differential equations (ODEs), which introduce various unknown parameters that need to be estimated efficiently from noisy measurements of concentration for a few species only. In this work, we present a new "systems-informed neural network" to infer the dynamics of experimentally unobserved species as well as the unknown parameters in the system of equations. By incorporating the system of ODEs into the neural networks, we effectively add constraints to the optimization algorithm, which makes the method robust to noisy and sparse measurements.

Clinical Viability of Magnetic Bead Implants in Muscle

Taylor, C. R. — 2022-08-03

Human movement is accomplished through muscle contraction, yet there does not exist a portable system capable of monitoring muscle length changes in real time. To address this limitation, we previously introduced magnetomicrometry, a minimally-invasive tracking technique comprising two implanted magnetic beads in muscle and a magnetic field sensor array positioned on the bodys surface adjacent the implanted beads. The implant system comprises a pair of spherical magnetic beads, each with a first coating of nickel-copper-nickel and an outer coating of Parylene C. In parallel work, we demonstrate submillimeter accuracy of magnetic bead tracking for muscle contractions in an untethered freely-roaming avian model. Here, we address the clinical viability of magnetomicrometry. Using a specialized device to insert magnetic beads into muscle in avian and lagomorph models, we collect data to assess gait metrics, bead migration, and bead biocompatibility. For these animal models, we find no gait differences post- versus pre-implantation, and bead migration towards one another within muscle does not occur for initial bead separation distances greater than 3 cm. Further, using extensive biocompatibility testing, the implants are shown to be non-irritant, non-cytotoxic, non-allergenic, and non-irritating. Our cumulative results lend support for the viability of these magnetic bead implants for implantation in human muscle. We thus anticipate their imminent use in human-machine interfaces, such as in control of prostheses, exoskeletons, and in closed-loop neuroprosthetics to aid recovery from neurological disorders.

Two distinct value-related patterns of EEG activity emerge during value-based choice, neither related to evidence accumulation

Froemer, R. — 2022-08-03

Previous work has identified characteristic neural signatures of value-based decision-making, including neural dynamics that closely resemble the ramping evidence accumulation process believed to underpin choice. Here, we test whether these signatures of the choice process can be temporally dissociated from additional, choice-independent value signals. Indeed, EEG activity during value-based choice revealed distinct spatiotemporal clusters, with a stimulus-locked cluster reflecting affective reactions to choice sets and a response-locked cluster reflecting choice difficulty. Surprisingly, neither of these clusters met the criteria for an evidence accumulation signal. Instead, we found that stimulus-locked activity can mimic an evidence accumulation process when aligned to the response. Re-analyzing four previous studies - including three perceptual decision-making studies - we show that response-locked signatures of evidence accumulation disappear when stimulus-locked and response-locked activity are modelled jointly. Collectively, our findings show that neural signatures of value can reflect choice-independent processes and look deceptively like evidence accumulation. Significance StatementTo choose, people must evaluate their options and select between them. Selection is well described by a process of accumulating evidence up to some threshold, with an electrophysiological signature in the centroparietal positivity (CPP). However, decision-making also gives rise to value signals reflecting affective reactions and other selection-unrelated processes. Measuring EEG while participants made value-based choices, we identified two spatiotemporally distinct value signals, neither reflecting evidence accumulation. Instead, we show that evidence accumulation signals found in the CPP can arise artifactually from overlapping stimulus- and response-related activity. These findings call for a significant reexamination of established links between neural and computational mechanisms of choice, while inviting deeper consideration of the array of cognitive and affective processes that occur in parallel.

Untethered Muscle Tracking Using Magnetomicrometry

Taylor, C. R. — 2022-08-04

Muscle tissue drives nearly all movement in the animal kingdom, providing power, mobility, and dexterity. Technologies for measuring muscle tissue motion, such as sonomicrometry, fluoromicrometry, and ultrasound, have significantly advanced our understanding of biomechanics. Yet, the field lacks the ability to monitor muscle tissue motion for animal behavior outside the lab. Towards addressing this issue, we previously introduced magnetomicrometry, a method that uses magnetic beads to wirelessly monitor muscle tissue length changes, and we validated magnetomicrometry via tightly-controlled in situ testing. In this study we validate the accuracy of magnetomicrometry against fluoromicrometry during untethered running in an in vivo turkey model. We demonstrate real-time muscle tissue length tracking of the freely-moving turkeys executing various motor activities, including ramp ascent and descent, vertical ascent and descent, and free roaming movement. Given the demonstrated capacity of magnetomicrometry to track muscle movement in untethered animals, we feel that this technique will enable new scientific explorations and an improved understanding of muscle function.

Heterodimerization-dependent secretion of BMPs in Drosophila

Bauer, M. — 2022-08-05

Combinatorial signaling is key to instruct context-dependent cell behaviors. During embryonic development, adult homeostasis and disease, Bone Morphogenetic Proteins (BMPs) act as dimers to instruct specific cellular responses. BMP ligands can form both homo- or heterodimers; however, obtaining direct evidence of the endogenous localization and function of each form has proven challenging. Here, we make use of precise genome editing and direct protein manipulation via protein binders to dissect the existence and functional relevance of BMP homo- and heterodimers in the Drosophila wing imaginal disc. This approach revealed in situ the existence of Dpp (BMP2/4)/Gbb (BMP5/6/7/8) heterodimers. We found that Gbb, despite being produced by all the cells of the wing imaginal disc, is only secreted by the cells in which Dpp is expressed. Dpp and Gbb form a gradient of heterodimers whereas neither Dpp nor Gbb homodimers are evident under endogenous physiological conditions. We find that the formation of heterodimers is critical for obtaining optimal signaling and long-range BMP distribution in the developing wing. These results indicate that Dpp/Gbb heterodimers are the active signal required for epithelial patterning and growth.

Doublecortin and JIP3 are neural-specific counteracting regulators of dynein-mediated retrograde trafficking

Rao, L. — 2022-08-11

Mutations in the microtubule (MT)-binding protein doublecortin (DCX) or in the MT- based molecular motor dynein result in lissencephaly. However, a functional link between DCX and dynein has not been defined. Here, we demonstrate that DCX negatively regulates dynein-mediated retrograde transport by reducing dyneins association with MTs and by disrupting the composition of the dynein motor complex. Previous work showed an increased binding of the adaptor protein C-Jun-amino-terminal kinase-interacting protein 3 (JIP3) to dynein in the absence of DCX. Using purified components, we demonstrate that JIP3 forms an active motor complex with dynein and its cofactor dynactin with two dyneins per complex. DCX competes with the binding of the second dynein, resulting in a velocity reduction of the complex. We conclude that DCX negatively regulates dynein-mediated retrograde transport through two critical interactions by regulating dynein binding to MTs and by regulating the association of JIP3 to the dynein motor complex.

Elements of divergence in germline determination in closely related species

Morita, S. — 2022-08-14

Evolutionary transitions enable the wide diversity in life histories of plants and animals. This is particularly germane in the development of the germ line in which fitness is a direct readout of evolutionary change. Here, we focused on the gem line of two distinct sea urchin species who shared a common ancestor 50 million years ago. Even though they both rely on inherited mechanisms to specify their germ line, the integration of stage-matched single cell RNA-seq (scRNA-seq) datasets from these two sea urchins revealed a variety of differences in gene expression, including a broader expression of the germ line factor Nanos2 in Lytechinus variegatus (Lv) compared to Strongylocentrotus purpuratus (Sp). In Sp, Nanos2 mRNA expression is highly restricted to the primordial germ cells (PGCs) by a lability element in its 3UTR. This element is lacking in the mRNA of Lv Nanos2, explaining how this mRNA more broadly accumulates in the Lv embryos. We discovered that the Lv Nanos2 3UTR instead leads to a germline specific translation of the protein. The results emphasize that regulatory mechanisms resulting in germline diversity rely less on transcriptional regulation and more on post-transcriptional and post-translational restrictions of key gene products, such as Nanos2. Highlights- The first integration of scRNA-seq datasets comparing two echinoderm species. - We find Nanos2 positive cells in the embryonic soma of Lytechinus variegatus, an unusual occurrence, but not in Strongylocentrous purpuratus. - We discovered that this somatic Nanos2 mRNA is lacking an important regulatory element (GNARLE) in its 3UTR - Instead, in Lv, the 3UTR of Nanos2 leads to its specific translation in the germ cells.

The neighborhood of interaction in human crowds is neither metric nor topological, but visual

Wirth, T. D. — 2022-08-19

Global patterns of collective motion in bird flocks, fish schools, and human crowds are thought to emerge from local interactions within a neighborhood of interaction, the zone in which an individual is influenced by their neighbors. Both topological and metric neighborhoods have been reported in birds, but this question has not been addressed in humans. With a topological neighborhood, an individual is influenced by a fixed number of nearest neighbors, regardless of their physical distance; whereas with a metric neighborhood, an individual is influenced by all neighbors within a fixed radius. We test these hypotheses experimentally with participants walking in real and virtual crowds, by manipulating the crowds density. Our results rule out a strictly topological neighborhood, are approximated by a metric neighborhood, but are best explained by a visual neighborhood with aspects of both. This finding has practical implications for modeling crowd behavior and understanding crowd disasters.

Molecular mechanisms of tubulogenesis revealed in the sea star hydro-vascular organ

Perillo, M. — 2022-08-26

A fundamental goal in the organogenesis field is to understand how cells organize into tubular shapes. Toward this aim, we have established the hydro-vascular organ in the sea star Patiria miniata as a model for tubulogenesis. In this animal, bilateral tubes grow out from the tip of the developing gut, and precisely extend to specific sites in the larva. This growth requires cell migration coupled with mitosis in distinct zones. Cell proliferation requires FGF signaling, whereas the three-dimensional orientation of the organ depends on Wnt signaling. Specification and maintenance of tube cell fate requires Delta/Notch signaling. Moreover, we identify target genes of the FGF pathway that contribute to tube morphology, revealing molecular mechanisms for tube outgrowth. Finally, we report that FGF activates the Six1/2 transcription factor, which serves as an evolutionarily ancient regulator of branching morphogenesis. This study uncovers novel mechanisms of tubulogenesis in vivo and we propose that cellular dynamics in the sea star hydro-vascular organ represents a key comparison for understanding the evolution of vertebrate organs. Highlights The hydro-vascular organ of the sea star presents a valuable model of tubulogenesis In this organ tube extension is driven by cell migration coupled with cell proliferation at specific growth zones The Wnt pathway controls directional outgrowth The FGF pathway promotes regionalized cell proliferation The Notch/Delta pathway is essential in cell fate repression in tubulogenesis A screen of FGF function revealed essential target gene expression, including the transcription factor Six1/2 Within a sister group to chordates, the sea star will reveal ancient mechanisms of tubulogenesis

Integrating sex-bias into studies of archaic admixture on chromosome X

Chevy, E. T. — 2022-09-01

Evidence of interbreeding between archaic hominins and humans comes from methods that infer the locations of segments of archaic haplotypes, or archaic coverage using the genomes of people living today. As more estimates of archaic coverage have emerged, it has become clear that most of this coverage is found on the autosomes-- very little is retained on chromosome X. Here, we summarize published estimates of archaic coverage on autosomes and chromosome X from extant human samples. We find on average 7.9 times more archaic coverage on autosomes than chromosome X, and identify broad continental patterns in this ratio: greatest in American samples, and least in South Asian samples. We also perform extensive simulation studies to investigate how the amount of archaic coverage, lengths of coverage, and rates of purging of archaic coverage are affected by sex-bias caused by an unequal sex ratio within the archaic introgressors. Our results generally confirm that, with increasing male sex-bias, less archaic coverage is retained on chromosome X. Ours is the first study to explicitly model such sex-bias and its potential role in creating the dearth of archaic coverage on chromosome X. Author summaryTens of thousands of years ago, humans interbred with our close hominin relatives (e.g. Neanderthals), which we know from finding segments of archaic hominin DNA in our genomes. Up to 4% of a human genome may be archaic DNA, but most of that archaic part is on the autosomes (the non-sex chromosomes). Chromosome X usually contains 3 to 10 times less archaic DNA than the autosomes. Also unlike the autosomes, it is always passed down by mothers, but only sometimes by fathers. There are several hypotheses for why chromosome X has less archaic DNA than the autosomes; one that has not been fully explored is whether the archaic hominins that interbred with our ancestors were mostly male or mostly female, known as sex-bias. In this paper, we use simulations to investigate whether sex-bias could produce less archaic DNA on chromosome X. Using simulation studies, we find that when the archaics are mostly male, modern humans end up with less archaic DNA on chromosome X than their autosomes, compared to when there is a female-bias or no sex-bias. Therefore, male sex-bias could be contributing to the difference in the amount of archaic DNA on chromosome X versus the autosomes. Of course, there are still plenty of other factors to be explored about how demography and selection have shaped our DNA. Studying patterns like this helps us learn more about early hominin natural history, and contextualizes archaic interbreeding events among other sex-biased events in human history.

Dysregulation of The Chromatin Environment Leads to Differential Alternative Splicing as A Mechanism Of Disease In a Human Model of Autism Spectrum Disorders

Leung, C. S. — 2022-09-09

Autism spectrum disorder (ASD) affects 1 in 44 children. Chromatin regulatory proteins are overrepresented among genes that contain high risk variants in ASD. Disruption of the chromatin environment leads to widespread dysregulation of gene expression, which is traditionally thought as a mechanism of disease pathogenesis associated with ASD. Alternatively, alterations in chromatin dynamics could also lead to dysregulation of alternative splicing, which is understudied as a mechanism of ASD pathogenesis. The anticonvulsant valproic acid (VPA) is a well-known environmental risk factor for ASD that acts as a class I histone deacetylase (HDAC) inhibitor. However, the precise molecular mechanisms underlying defects in human neuronal development associated with exposure to VPA are understudied. To dissect how VPA exposure and subsequent chromatin hyperacetylation influence molecular signatures involved in ASD pathogenesis, we conducted RNA sequencing (RNA-seq) in human cortical neurons that were treated with VPA. We observed that differentially expressed genes (DEGs) were enriched for mRNA splicing, mRNA processing, histone modification, and metabolism related gene sets. Furthermore, we observed widespread increase in the number and the type of alternative splicing events. Analysis of differential transcript usage (DTU) showed that exposure to VPA induces extensive alterations in transcript isoform usage across neurodevelopmentally important genes. Finally, we find that DEGs and genes that display DTU overlap with known ASD-risk genes. Together, these findings suggest that, in addition to differential gene expression, changes in alternative splicing correlated with alterations in the chromatin environment could act as an additional mechanism of disease in ASD.

Frontotemporal network engagement by human intracranial stimulation in limbic hubs

Allawala, A. B. — 2022-09-10

BackgroundPrefrontal circuits in the human brain play an important role in cognitive and affective processing. Neuromodulation therapies delivered to certain key hubs within these circuits are being used with increasing frequency to treat a host of neuropsychiatric disorders. However, the detailed neurophysiological effects of stimulation to these hubs are largely unknown. MethodsHere, we performed intracranial recordings across prefrontal networks while delivering electrical stimulation to two well-established white matter hubs involved in cognitive regulation and depression: the subcallosal cingulate (SCC) and ventral capsule/ventral striatum (VC/VS). ResultsWe demonstrate a shared frontotemporal circuit consisting of the ventromedial PFC, amygdala, and lateral orbitofrontal cortex where gamma oscillations are differentially modulated by stimulation target. Additionally, we found subject-specific responses to stimulation in the dorsal anterior cingulate cortex and demonstrate the capacity for further tuning of neural activity using current-steered stimulation. ConclusionsOur findings indicate a potential neurophysiological mechanism for the dissociable therapeutic effects seen across the SCC and VC/VS DBS targets for psychiatric neuromodulation and our results lay the groundwork for personalized, network-guided neurostimulation therapy.

Analysis of somatic mutations in senescent cells using single-cell whole-genome sequencing

Zhang, L. — 2022-09-19

SO_SCPLOWUMMARYC_SCPLOWSomatic mutations accumulate in multiple organs and tissues during aging and are a known cause of cancer. Here we tested whether mutations accumulate during replicative senescence. Cellular senescence is also a possible cause of functional decline in aging, yet also acts as an anti-cancer mechanism in vivo. Using single-cell whole-genome sequencing, we compared mutation burdens between early passage and deeply senescent human fibroblasts. The results showed that single-nucleotide variations and small insertions and deletions increased in senescent cells by about two-fold, but have the same spectrum as early passage cells, while it has been known that particular mutational signatures are found in tumor cells. In contrast, aneuploidies were observed in half the senescent cells, but largely absent in early passage cells. Thus, the patterns of mutations among senescent, normal-aged and tumor cells differ significantly.

Monkey dorsolateral prefrontal cortex represents abstract visual sequences during a no-report task

Yusif Rodriguez, N. D. R. — 2022-09-19

Monitoring sequential information is an essential component of our daily lives. Many of these sequences are abstract, in that they do not depend on the individual stimuli, but do depend on an ordered set of rules (e.g., chop then stir when cooking). Despite the ubiquity and utility of abstract sequential monitoring, little is known about its neural mechanisms. Human rostrolateral prefrontal cortex (RLPFC) exhibits specific increases in neural activity (i.e., "ramping") during abstract sequences. Monkey dorsolateral prefrontal cortex (DLPFC) has been shown to represent sequential information in motor (not abstract) sequence tasks, and contains a sub-region, area 46, with homologous functional connectivity to human RLPFC. To test the prediction that area 46 may represent abstract sequence information, and do so with parallel dynamics to those found in humans, we conducted functional magnetic resonance imaging (fMRI) in monkeys. When monkeys performed no-report abstract sequence viewing, we found that left and right area 46 responded to abstract sequential changes. Interestingly, responses to rule and number changes overlapped in right area 46 and left area 46 exhibited responses to abstract sequence rules with changes in ramping activation, similar to that observed in humans. Together, these results indicate that monkey DLPFC monitors abstract visual sequential information, potentially with a preference for different dynamics in the two hemispheres. More generally, these results show that abstract sequences are represented in functionally homologous regions across monkeys and humans. Significance StatementDaily, we complete sequences that are "abstract" because they depend on an ordered set of rules (e.g., chop then stir when cooking) rather than the identity of individual items. Little is known about how the brain tracks, or monitors, this abstract sequential information. Based on previous human work showing abstract sequence related dynamics in an analogous area, we tested if monkey dorsolateral prefrontal cortex (DLPFC), specifically area 46, represents abstract sequential information using awake monkey fMRI. We found that area 46 responded to abstract sequence changes, with a preference for more general responses on the right and dynamics similar to humans on the left. These results suggest that abstract sequences are represented in functionally homologous regions across monkeys and humans.

Dysregulation of neural activity and microglia function following exposure to the global environmental contaminant perfluorooctane sulfonate (PFOS)

Paquette, S. E. — 2022-10-03

BackgroundPer- and polyfluoroalkyl substances (PFAS) are biopersistent pollutants that have become global contaminants as a result of their diverse applications in commerce and industry. While some in vitro and epidemiological studies have explored the neurotoxic potential of perfluorooctane sulfonate (PFOS), a prevalent PFAS congener, it is unknown how developmental exposure to PFOS affects neuronal communication and other developmentally critical neural cell types, including microglia. ObjectivesWe sought to determine the extent to which PFOS exposure disrupts brain health, neuronal activity, and neuron-microglia communication during brain development. In addition, while PFOS impairs humoral immunity, its impact on innate immune cells, including resident microglia, is unclear. As such, we aimed to determine whether microglia are cellular targets of PFOS and, if so, whether disrupted microglial development and/or function could contribute to or is influenced by PFOS-induced neural dysfunction. MethodsZebrafish were chronically exposed to either control solution (0.1% DMSO), 7 {micro}M PFOS, 14 {micro}M PFOS, 28 {micro}M PFOS, or 64 {micro}M perfluorooctanoic acid (PFOA). We used in vivo imaging and gene expression analysis to assess microglial populations in the developing brain and to determine shifts in microglial state. We functionally challenged microglial using a larval brain injury model and, to assess the neuronal signaling environment, performed functional neuroimaging experiments utilizing the photoconvertible calcium indicator CaMPARI. These studies were paired with optogenetic manipulations of neurons and microglia, an untargeted metabolome wide association study (MWAS), and larval swim behavior assessments. ResultsDevelopmental PFOS exposure resulted in a shift away from the homeostatic microglia state, as determined by functional and morphological changes, as well as transcriptional upregulation of the microglia activation gene p2ry12. PFOS-induced effects on microglia state exacerbated microglia responses to brain injury in the absence of increased cell death or inflammation. PFOS exposure also heightened neural calcium activity, and optogenetic silencing of neurons or microglia independently was sufficient to normalize microglial responses to injury. An untargeted MWAS of larval brains revealed PFOS-exposed larvae had neurochemical signatures of excitatory-inhibitory imbalance. Behaviorally, PFOS-exposed larvae also exhibited anxiety-like thigmotaxis. To test whether the neuronal and microglial phenotypes were specific to PFOS, we exposed embryos to PFOA, a known immunotoxic PFAS. PFOA did not alter thigmotaxis, neuronal activity, or microglial responses, further supporting a role for neuronal activity as a critical modifier of microglial function following PFOS exposure. DiscussionTogether, this study provides the first detailed account of the effects of PFOS exposure on neural cell types in the developing brain in vivo and adds neuronal hyperactivity as an important endpoint to assess when studying the impact of toxicant exposures on microglia function.

Streetlights affect moth orientation beyond flight-to-light behaviour

Degen, J. — 2022-10-08

One of the most dramatic changes occurring on our planet in recent decades is the ever-increasing extensive use of artificial light at night, which drastically altered the environment nocturnal animals are adapted to 1,2. One nocturnal species group experiencing marked declines are moths, which are not only of great importance for species conservation, but also for their key role in food webs and in ecosystem services such as nocturnal plant pollination 3,4. Light pollution has been identified as a driver in the dramatic insect decline of the past years 5-7, yet little is known about its impact on natural insect orientation behaviour. Using harmonic radar tracking, we show that the orientation of several species of moths is significantly affected by streetlights, although only 4 % of individuals showed flight-to-light behaviour. We reveal a species-specific barrier effect of streetlights on lappet moths whenever the moon was not available as a natural celestial cue. Furthermore, streetlights increased the tortuosity of flight trajectories for both hawk moths and lappet moths. Our results provide the first spatially resolved experimental evidence for the fragmentation of landscapes by streetlights and demonstrate that light pollution affects movement patterns of moths beyond previously assumed extend, potentially affecting their reproductive success and hampering a vital ecosystem service.

Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells

Menon, N. G. — 2022-10-12

Dry Eye Disease (DED) is a complex pathology affecting millions of people with significant impact on quality of life. Corneal inflammation, including via the NF{kappa}B pathway, plays a key etiological role in DED. Recombinant human proteoglycan 4 (rhPRG4) has been shown to be a clinically effective treatment for DED that has anti-inflammatory effects in corneal epithelial cells, but the underlying mechanism is still not understood. Our goal was to understand if rhPRG4 affects TNF-stimulated inflammatory activity in corneal epithelial cells. We treated hTERT-immortalized corneal epithelial (hTCEpi) cells {+/-}TNF {+/-}rhPRG4 and performed Western blotting on cell lysate and RNA sequencing. Bioinformatics analysis revealed that rhPRG4 had a significant effect on TNF-mediated inflammation with potential effects on matricellular homeostasis. rhPRG4 reduced activation of key inflammatory pathways and decreased expression of transcripts for key inflammatory cytokines, interferons, interleu-kins, and transcription factors. TNF treatment significantly increased phosphorylation and nuclear translocation of p65, and rhPRG4 significantly reduced both these effects. RNA sequencing identified FAT10, which has not been studied in the context of DED, as a key pro-inflammatory transcript increased by TNF and decreased by rhPRG4. These results were confirmed at the protein level. In summary, rhPRG4 is able to downregulate NF{kappa}B activity in hTCEpi cells, suggesting a potential biological mechanism by which it may act as a therapeutic for DED.

GrapHiC: An integrative graph based approach for imputing missing Hi-C reads

Murtaza, G. — 2022-10-21

Hi-C experiments allow researchers to study and understand the 3D genome organization and its regulatory function. Unfortunately, sequencing costs and technical constraints severely restrict access to high-quality Hi-C data for many cell types. Existing frameworks rely on a sparse Hi-C dataset or cheaper-to-acquire ChIP-seq data to predict Hi-C contact maps with high read coverage. However, these methods fail to generalize to sparse or cross-cell-type inputs because they do not account for the contributions of epigenomic features or the impact of the structural neighborhood in predicting Hi-C reads. We propose GrapHiC, which combines Hi-C and ChIP-seq in a graph representation, allowing more accurate embedding of structural and epigenomic features. Each node represents a binned genomic region, and we assign edge weights using the observed Hi-C reads. Additionally, we embed ChIP-seq and relative positional information as node attributes, allowing our representation to capture structural neighborhoods and the contributions of proteins and their modifications for predicting Hi-C reads. Our evaluations show that GrapHiC generalizes better than the current state-of-the-art on cross-cell-type settings and sparse Hi-C inputs. Moreover, we can utilize our framework to impute Hi-C reads even when no Hi-C contact map is available, thus making high-quality Hi-C data more accessible for many cell types. Availabilityhttps://github.com/rsinghlab/GrapHiC ACM Reference FormatGhulam Murtaza, Justin Wagner, Justin M. Zook, and Ritambhara Singh. 2018. GrapHiC: An integrative graph based approach for imputing missing Hi-C reads. In Proceedings of 22nd International Workshop on Data Mining in Bioinformatics (BioKDD 23). ACM, New York, NY, USA, 16 pages. https://doi.org/XXXXXXX.XXXXXXX

Perceiving Depth from Texture and Disparity Cues: Evidence for a Non-Probabilistic Account of Cue Integration

Kemp, J. T. — 2022-10-21

The fundamental question of how the brain derives 3D information from the inherently ambiguous visual input has been approached during the last two decades with probabilistic theories of 3D perception. Probabilistic models, such as the Maximum Likelihood Estimation (MLE) model, derive from multiple independent depth cues the most probable 3D interpretations. These estimates are then combined by weighing them according to their uncertainty to obtain the most accurate and least noisy estimate. In three experiments we tested an alternative theory of cue integration termed the Intrinsic Constraint (IC) theory. This theory postulates that the visual system does not derive the most probable interpretation of the visual input, but the most stable interpretation amid variations in viewing conditions. This goal is achieved with the Vector Sum model, that represents individual cue estimates as components of a multidimensional vector whose norm determines the combined output. In contrast with the MLE model, individual cue estimates are not accurate, but linearly related to distal 3D properties through a deterministic mapping. In Experiment 1, we measured the cue-specific biases that arise when viewing single-cue stimuli of various simulated depths and show that the Vector Sum model accurately predicts an increase in perceived depth when the same cues are presented together in a combined-cue stimulus. In Experiment 2, we show how Just Noticeable Differences (JNDs) are accounted for by the IC theory and demonstrate that the Vector Sum model predicts the classic finding of smaller JNDs for combined-cue versus single-cue stimuli. Most importantly, this prediction is made through a radical re-interpretation of the JND, a hallmark measure of stimulus discriminability previously thought to estimate perceptual uncertainty. In Experiment 3, we show that biases found in cue-integration experiments cannot be attributed to flatness cues, as assumed by the MLE model. Instead, we show that flatness cues produce no measurable difference in perceived depth for monocular (3A) or binocular viewing (3B), as predicted by the Vector Sum model.

Patient-specific deep offline artificial pancreas for blood glucose regulation in type 1 diabetes

Deng, Y. — 2022-10-24

Due to insufficient insulin secretion, patients with type 1 diabetes mellitus (T1DM) are prone to blood glucose fluctuations ranging from hypoglycemia to hyperglycemia. While dangerous hypoglycemia may lead to coma immediately, chronic hyperglycemia increases patients risks for cardiorenal and vascular diseases in the long run. In principle, an artificial pancreas - a closed-loop insulin delivery system requiring patients manually input insulin dosage according to the upcoming meals - could supply exogenous insulin to control the glucose levels and hence reduce the risks from hyperglycemia. However, insulin overdosing in some type 1 diabetic patients, who are physically active, can lead to unexpected hypoglycemia beyond the control of common artificial pancreas. Therefore, it is important to take into account the glucose decrease due to physical exercise when designing the next-generation artificial pancreas. In this work, we develop a deep reinforcement learning algorithm using a T1DM dataset, containing data from wearable devices, to automate insulin dosing for patients with T1DM. In particular, we build patient-specific computational models using systems biology informed neural networks (SBINN), to mimic the glucose-insulin dynamics for a few patients from the dataset, by simultaneously considering patient-specific carbohydrate intake and physical exercise intensity.

A Taxonomy of Seizure Spread Patterns, Speed of Spread, and Associations With Structural Connectivity

Revell, A. Y. — 2022-10-26

Although seizure detection algorithms are widely used to localize seizure onset on intracranial EEG in epilepsy patients, relatively few studies focus on seizure activity beyond the seizure onset zone to direct treatment of surgical patients with epilepsy. To address this gap, we develop and compare fully automated deep learning algorithms to detect seizure activity on single channels, effectively quantifying spread when deployed across multiple channels. Across 275 seizures in 71 patients, we discover that the extent of seizure spread across the brain and the timing of seizure spread between temporal lobe regions is associated with both surgical outcomes and the brains structural connectivity between temporal lobes. Finally, we uncover a hierarchical structure of seizure spread patterns highlighting the relationship between clusters of seizures. Collectively, these findings underscore the broad utility in quantifying seizure activity past seizure onset to identify novel mechanisms of seizure evolution and its relationship to potential seizure freedom.

Examining the early distribution of the artemisinin-resistant Plasmodium falciparum kelch13 R561H mutation in Rwanda

Kirby, R. — 2022-10-26

Artemisinin resistance mutations in Plasmodium falciparum kelch13 (Pfk13) have begun to emerge in Africa. Pfk13-R561H was the first reported African mutation found in Rwanda in 2014, but limited sampling left questions about its early distribution and origin. We detected 476 parasitemias among 1873 residual blood spots from a 2014-15 Rwanda Demographic Health Survey. We sequenced 351 samples revealing 341/351 were wild type (97.03% weighted) and 4 samples (1.34% weighted) harbored R561H which were significantly spatially clustered. Our study better defines the early distribution of R561H in Rwanda and suggests that the origin may have involved higher-transmission regions.

Expanding the stdpopsim species catalog, and lessons learned forrealistic genome simulations

Lauterbur, M. E. — 2022-10-31

Simulation is a key tool in population genetics for both methods development and empirical research, but producing simulations that recapitulate the main features of genomic data sets remains a major obstacle. Today, more realistic simulations are possible thanks to large increases in the quantity and quality of available genetic data, and to the sophistication of inference and simulation software. However, implementing these simulations still requires substantial time and specialized knowledge. These challenges are especially pronounced for simulating genomes for species that are not well-studied, since it is not always clear what information is required to produce simulations with a level of realism sufficient to confidently answer a given question. The community-developed framework stdpopsim seeks to lower this barrier by facilitating the simulation of complex population genetic models using up-to-date information. The initial version of stdpopsim focused on establishing this framework using six well-characterized model species (Adrion et al., 2020). Here, we report on major improvements made in the new release of stdpopsim (version 0.2), which includes a significant expansion of the species catalog and substantial additions to simulation capabilities. Features added to improve the realism of the simulated genomes include non-crossover recombination and provision of species-specific genomic annotations. Through community-driven efforts, we expanded the number of species in the catalog more than three-fold and broadened coverage across the tree of life. During the process of expanding the catalog, we have identified common sticking points and developed best practices for setting up genome-scale simulations. We describe the input data required for generating a realistic simulation, suggest good practices for obtaining the relevant information from the literature, and discuss common pitfalls and major considerations. These improvements to stdpopsim aim to further promote the use of realistic whole-genome population genetic simulations, especially in non-model organisms, making them available, transparent, and accessible to everyone.

Chromosome-scale scaffolding of the fungus gnat genome (Diptera: Bradysia coprophila)

Urban, J. M. — 2022-11-04

BackgroundThe fungus gnat, Bradysia (Sciara) coprophila, has compelling chromosome biology. Paternal chromosomes are eliminated during spermatogenesis whereas both maternal X sister chromatids are retained. Embryos start with three copies of the X chromosome, but 1-2 copies are eliminated from somatic cells as part of sex determination, and one is eliminated in the germline to restore diploidy. These developmentally normal events present opportunities to study chromosome movements that are unusual in other systems. To support such studies, we previously generated a highly contiguous optical-map-scaffolded long-read assembly (Bcop_v1) of the male somatic genome. However, the scaffolds were not chromosome-scale, the majority of the assembly lacked chromosome assignments, and the order and orientation of the contigs along chromosomes remained unknown. FindingsMale pupae Hi-C data was used to correct, order, and orient the contigs from Bcop_v1 into chromosome-scale scaffolds, producing the updated assembly, Bcop_v2. Several orthogonal analyses allowed us to (i) identify the corresponding chromosome for each scaffold, (ii) orient them with respect to polytene maps, and (iii) determine that they were highly concordant with the chromosomes they represent. Gene annotations produced for Bcop_v1 were lifted over to Bcop_v2. Chromosomal repeat distributions highlight a potential telomeric sequence. Finally, the Hi-C data shed new light on three "fold-back regions" seen to physically interact in images of polytene X chromosomes. ConclusionsStudies of the unusual chromosome movements in Bradysia coprophila will benefit from the updated assembly (Bcop_v2) where each somatic chromosome is represented by a single scaffold.

A condensate forming tether for lariat debranching enzyme is defective in non-photosensitive trichiothiodystrophy

Townley, B. A. — 2022-11-04

The pre-mRNA life cycle requires intron processing; yet, how intron processing defects influence splicing and gene expression is unclear. Here, we find TTDN1, which is frequently mutated in non-photosensitive trichothiodystrophy (NP-TTD), functionally links intron lariat processing to the spliceosome. The conserved TTDN1 C-terminal region directly binds lariat debranching enzyme DBR1, while its N-terminal intrinsically disordered region (IDR) binds the intron binding complex (IBC). The IDR forms condensates in vitro and is needed for IBC interaction. TTDN1 loss causes significant intron lariat accumulation, as well as splicing and gene expression defects, mirroring phenotypes observed in NP-TTD patient cells. Ttdn1{Delta}/{Delta} mice recapitulate intron processing defects and neurodevelopmental phenotypes seen in NP-TTD. A DBR1-IDR fusion recruits DBR1 to the IBC and circumvents the requirement for TTDN1, indicating this tethering role as its major molecular function. Collectively, our findings unveil key functional connections between lariat processing, splicing outcomes, and NP-TTD molecular pathology.

The MuSK-BMP pathway maintains myofiber size in slow muscle through regulation of Akt-mTOR signaling

Jaime, D. — 2022-11-05

Myofiber size regulation is critical in health, disease, and aging. MuSK (muscle-specific kinase) is a BMP (bone morphogenetic protein) co-receptor that promotes and shapes BMP signaling. MuSK is expressed at all neuromuscular junctions and is also present extrasynaptically in the slow soleus muscle. To investigate the role of the MuSK-BMP pathway in vivo we generated mice lacking the BMP-binding MuSK Ig3 domain. These {Delta}Ig3-MuSK mice are viable and fertile with innervation levels comparable to wild type. In 3-month-old mice myofibers are smaller in the slow soleus, but not in the fast tibialis anterior (TA). Transcriptomic analysis revealed soleus-selective decreases in RNA metabolism and protein synthesis pathways as well as dysregulation of IGF1-Akt-mTOR pathway components. Biochemical analysis showed that Akt-mTOR signaling is reduced in soleus but not TA. We propose that the MuSK-BMP pathway acts extrasynaptically to maintain myofiber size in slow muscle by promoting protein synthetic pathways including IGF1-Akt-mTOR signaling. These results reveal a novel mechanism for regulating myofiber size in slow muscle and introduce the MuSK-BMP pathway as a target for promoting muscle growth and combatting atrophy.

Jointly aligning cells and genomic features of single-cell multi-omics data with co-optimal transport

Demetci, P. — 2022-11-10

The availability of various single-cell sequencing technologies allows one to jointly study multiple genomic features and understand how they interact to regulate cells. Although there are experimental challenges to simultaneously profile multiple features on the same single cell, recent computational methods can align the cells from unpaired multi-omic datasets. However, studying regulation also requires us to map the genomic features across different measurements. Unfortunately, most single-cell multi-omic alignment tools cannot perform these alignments or need prior knowledge. We introduce O_SCPLOWSCOOTRC_SCPLOW, a co-optimal transport-based method, which jointly aligns both cells and genomic features of unpaired single-cell multi-omic datasets. We apply O_SCPLOWSCOOTRC_SCPLOW to various single-cell multi-omic datasets with different types of measurements. Our results show that O_SCPLOWSCOOTRC_SCPLOW provides quality alignments for unsupervised cell-level and feature-level integration of datasets with sparse feature correspondences (e.g., one-to-one mappings). For datasets with dense feature correspondences (e.g., many-to-many mappings), our joint framework allows us to provide supervision on one level (e.g., cell types), thus improving alignment performance on the other (e.g., genomic features) or vice-versa. The unique joint alignment framework makes O_SCPLOWSCOOTRC_SCPLOW a helpful hypothesis-generation tool for the integrative study of unpaired single-cell multi-omic datasets. Available at: https://github.com/rsinghlab/SCOOTR.

Aliphatic residues contribute significantly to the phase separation of TDP-43 C-terminal domain

Mohanty, P. — 2022-11-10

TAR DNA binding protein 43 (TDP-43) is involved in key processes in RNA metabolism such as splicing, stability and transcription. TDP-43 dysfunction is frequently implicated in many neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and fronto-temporal dementia (FTD). The prion-like, disordered C-terminal domain (CTD) of TDP-43 is aggregation-prone and harbors the majority (~90%) of all ALS-related mutations. Recent studies have established that TDP-43 CTD can undergo liquid-liquid phase separation (LLPS) in isolation and is important for phase separation (PS) of the full-length protein under physiological conditions. While a short conserved helical region (CR, spanning residues 319-341) promotes oligomerization and is essential for LLPS, aromatic residues in the flanking disordered regions (IDR1/2) have also been found to play a critical role in PS and aggregation. However, TDP-43 CTD has a distinct sequence composition compared with other phase separating proteins, including many aliphatic residues. These residues have been suggested to modulate the apparent viscosity of the resulting phases, but their direct contribution to phase separation has been relatively ignored. Here, we utilized a multiscale simulation and experimental approach to assess the residue-level determinants of TDP-43 CTD phase separation. Single chain and condensed phase simulations performed at the atomistic and coarse-grained level respectively, identified the importance of aromatic residues (previously established) while also suggesting an essential role for aliphatic methionine residues in LLPS. In vitro experiments confirmed the role of phenylalanine, methionine, and leucine (but not alanine) residues in driving the phase separation of CTD, which have not been previously considered essential for describing the molecular grammar of PS. Finally, NMR experiments also showed that phenylalanine residues in the disordered flanking regions and methionine residues both within and outside the CR contribute important contacts to CTD interactions. Broadly, our work highlights the importance of non-alanine aliphatic residues such as methionine and leucine, and potentially valine and isoleucine, in determining the LLPS propensity, expanding the molecular grammar of protein phase separation to include critical contributions from aliphatic residues.

Expression of retrotransposons contributes to aging in Drosophila

Schneider, B. K. — 2022-11-15

Retrotransposons are a class of transposable elements capable of self-replication and insertion into new genomic locations. Across species, the mobilization of retrotransposons in somatic cells has been suggested to contribute to the cell and tissue functional decline that occurs during aging. Retrotransposon expression generally increases with age, and de novo insertions have been observed to occur during tumorigenesis. However, the extent to which new retrotransposon insertions occur during normal aging and their effect on cellular and animal function remains understudied. Here we use a single nucleus whole genome sequencing approach in Drosophila to directly test whether transposon insertions increase with age in somatic cells. Analyses of nuclei from thoraces and indirect flight muscles using a newly developed pipeline, Retrofind, revealed no significant increase in the number of transposon insertions with age. Despite this, reducing the expression of two different retrotransposons, 412 and Roo, extends lifespan, without increasing stress resistance. This suggests a key role for transposon expression and not insertion in regulating longevity. Transcriptomic analyses revealed similar changes to gene expression in 412 and Roo knockdown flies and highlighted potential changes to genes involved in proteolysis and immune function as potential contributors to the observed changes in longevity. Combined, our data show a clear link between retrotransposon expression and aging. Author SummaryWith the onset of modern medicine, the average age of the population has significantly increased, leading to more individuals living with chronic health issues. Rather than treat each age-associated disorder individually, one approach to target multiple health concerns simultaneously might to be target aging itself. Genomic instability is a hallmark of aging cells that has been proposed to be a key contributor to age-associated cellular decline. Transposons are mobile genetic elements capable of inserting into new genomic locations, thus having the potential to increase genomic instability. Consistent with this, transposon expression generally increases with age. However, the extent to which transposon insertions accumulate to disrupt the genome of cells within aging individuals has remained an open question. We specifically answer this through single cell whole genome sequencing and find that transposon insertions do not increase with age. Even though insertions did not increase, the expression of transposons is linked to aging, as reducing the expression of individual transposons extended lifespan. Transcriptome studies of these long-lived flies revealed increased expression of genes linked to proteolysis genes and to functioning of the immune system. Our study therefore establishes transposon expression, and not insertion, as a critical contributor to animal aging.

Endocytosis in Trypanosoma cruzi Depends on Proper Recruitment and Regulation of Functionally Redundant Myosin Motors

Chasen, N. M. — 2022-11-18

Utilized by the free-living kinetoplastid Bodo saltans to feed on bacterial prey, the cytostome-cytopharynx complex (SPC) is an endocytic organelle absent from all human trypanosomatid pathogens save Trypanosoma cruzi. Building upon our previous work identifying the myosin motor MyoF as the first enzymatic component of the T. cruzi SPC, we sought to expand our understanding of this distinct organelle by identifying additional protein machinery which contribute to the endocytic process. While deletion of MyoF alone did not fully ablate endocytosis, we found that deletion of both MyoF and the similarly localized MyoC produced an endocytic-null phenotype that was rescued upon complementation. To identify potential regulatory components of this motor complex, we pulled down MyoF and identified an SPC-targeted protein that contained an annotated EF-hand calcium-binding motif that was conserved across a wide range of protozoan lineages. Surprisingly, deletion of this myosin associated protein (MyAP) alone was sufficient to produce an endocytic-null phenotype, which we were able to fully rescue via complementation. The deletion of MyAP also caused the mis-localization of both cytopharynx myosins to the cytosol. While MyAP lacking the EF-hand domain was unable to complement endocytosis, it was sufficient to restore proper myosin localization. This suggested that MyAP plays two distinct roles, one in targeting myosins to the SPC and a second in regulating myosin motor activity. Transmission electron microscopy also revealed that endocytic-null mutants lacked the electron lucent lipid inclusions typically seen in the pre-lysosomal reservosomes of T. cruzi epimastigotes. Mass spectrometry based lipidomic analysis subsequently revealed a dramatic reduction in the scavenged cholesterol content in the endocytic-null mutants, which can be attributed to an inability to endocytose exogenous lipid-protein complexes for storage in the reservosomes. Overall, this work showcases the first viable endocytic-null mutants generated in T. cruzi through specific gene deletion and highlights the feasibility of leveraging this strategy towards a full dissection of the endocytic machinery and biogenesis of the SPC. ImportanceTrypanosoma cruzi chronically infects over 7 million people in the Americas and current therapeutics are insufficient to effectively cure infection. The lack of progress in developing effective vaccines or drug treatments is due, in part, to longstanding technical limitations in studying this parasite and a lack of resources committed to support research and eradication efforts. As part of its parasitic lifestyle, T. cruzi is forced to obtain basic nutrients directly from its host environment, making the development of methods to block nutrient uptake an attractive strategy to control parasite growth and transmission. While the bulk uptake of complex nutrients by T. cruzi occurs via an endocytic structure, often referred to as the cytostome-cytopharynx complex (SPC), how exactly this tubular endocytic organelle functions at a mechanistic level has remained a mystery. In this work, we investigated the contribution of several SPC targeted myosin motors and an associated protein factor to endocytic activity. By identifying and characterizing the molecular machinery responsible for nutrient uptake, we hope to both expand our basic understanding of how this deadly pathogen acquires essential nutrients from its host, while also revealing new potential therapeutic targets to impede nutrient uptake.

Effort Foraging Task reveals positive correlation between individual differences in the cost of cognitive and physical effort in humans and relationship to self-reported motivation and affect

Bustamante, L. A. — 2022-11-22

Effort-based decisions, in which people weigh potential future rewards against effort costs required to achieve those rewards, have largely been studied separately for cognitive or physical effort, yet most real-world actions incur both effort costs. What is the relationship between cognitive and physical effort costs? Here we attempt to formalize the mechanisms underlying effort-based decisions and address methodological challenges to isolate and measure these mechanisms. Patch foraging is an ecologically valid reward rate maximization problem with well-developed theoretical tools to understand choices. We developed the Effort Foraging Task, which embedded cognitive or physical effort into a patch foraging sequential decision task, to isolate and quantify the cost of both cognitive and physical effort using a computational model. Participants chose between harvesting a depleting patch, or traveling to a new patch that was costly in time and effort. Participants exit thresholds (reflecting the reward they expected to receive by harvesting when they chose to travel to a new patch) were sensitive to cognitive and physical effort demands, allowing us to quantify the perceived effort cost in monetary terms. Individual differences in cognitive and physical effort costs were positively correlated, suggesting that these are perceived and processed in common terms across different domains. We found patterns of correlation of both cognitive and physical effort costs with self-reported anxiety, cognitive function, behavioral activation, and self-efficacy. This suggests the task captures decision mechanisms associated with real-world motivation and can be used to study individual variation in effort-based decisions across domains of cost.

retro-Tango enables versatile retrograde circuit tracing in Drosophila

Sorkac, A. — 2022-11-24

Transsynaptic tracing methods are crucial tools in studying neural circuits. Although a couple of anterograde tracing methods and a targeted retrograde tool have been developed in Drosophila melanogaster, there is still need for an unbiased, user-friendly, and flexible retrograde tracing system. Here we describe retro-Tango, a method for transsynaptic, retrograde circuit tracing and manipulation in Drosophila. In this genetically encoded system, a ligand-receptor interaction at the synapse triggers an intracellular signaling cascade that results in reporter gene expression in presynaptic neurons. Importantly, panneuronal expression of the elements of the cascade renders this method versatile, enabling its use not only to test hypotheses but also to generate them. We validate retro-Tango in various circuits and benchmark it by comparing our findings with the electron microscopy reconstruction of the Drosophila hemibrain. Our experiments establish retro-Tango as a key method for circuit tracing in neuroscience research.

Expected Costs of Mental Efforts are Updated When People Exert Effort, not by Prospective Information

Nagase, A. M. — 2022-11-27

To understand why people avoid mental effort, it is crucial to reveal the mechanisms by which we learn and decide about mental effort costs. This study investigated whether mental effort cost learning aligns with temporal-difference (TD) learning or alternative mechanisms. Model-based fMRI analyses showed no correlation between cost prediction errors (CPEs) and activity in the dorsomedial frontal cortex/dorsal anterior cingulate cortex (dmFC/dACC) or striatum at the time of a fully informative effort cue about upcoming effort demands, contradicting the TD hypothesis. Instead, CPEs correlate with dmFC/dACC (positively) and caudate (negatively) activity at effort completion. Furthermore, only activity patterns at effort completion predict subsequent choices. These results show that decision policies are updated retrospectively at effort completion, updating expected costs with prediction error between experienced effort and prior expectations, demonstrating mental effort cost learning is retrospective, and imply that adaptive learning of mental effort cost does not follow canonical TD learning. Significance StatementUnderstanding how people learn about mental effort costs is essential for advancing theories of motivation and cognitive control. However, the algorithms supporting such learning remain unclear. This study addressed this gap and found that temporal-difference learning, commonly used to explain reward learning, could not account for how people learn about effort. Instead, decision policies were updated retrospectively at effort completion, based on a prediction error between experienced effort and prior expectations. These findings reveal that mental effort cost learning is fundamentally retrospective and imply that it relies on mechanisms distinct from canonical temporal-difference learning.

Age-related decline in prefrontal glutamate predicts failure to efficiently deploy working memory in the service of learning

Rmus, M. — 2022-11-29

The ability to use past experience to effectively guide decision making declines in older adulthood. Such declines have been theorized to emerge from either impairments of striatal reinforcement learning systems (RL) or impairments of recurrent networks in prefrontal and parietal cortex that support working memory (WM). Distinguishing between these hypotheses has been challenging because either RL or WM could be used to facilitate successful decision making in typical laboratory tasks. Here we investigated the neurocomputational correlates of age-related decision making deficits using an RL-WM task to disentangle these mechanisms, a computational model to quantify them, and magnetic resonance spectroscopy to link them to their molecular bases. Our results reveal that task performance is worse in older age, in a manner best explained by working memory deficits, as might be expected if cortical recurrent networks were unable to sustain persistent activity across multiple trials. Consistent with this, we show that older adults had lower levels of prefrontal glutamate, the excitatory neurotransmitter thought to support persistent activity, compared to younger adults. Individuals with the lowest prefrontal glutamate levels displayed the greatest impairments in working memory after controlling for other anatomical and metabolic factors. Together, our results suggest that lower levels of prefrontal glutamate may contribute to failures of working memory systems and impaired decision making in older adulthood.

Maternal obesity driven changes in collagen linearity of breast extracellular matrix induces invasive mammary epithelial cell phenotype

Amens, J. N. — 2022-12-02

Obesity has been linked with numerous health issues as well as an increased risk of breast cancer. Although effects of direct obesity in patient outcomes is widely studied, effects of exposure to obesity-related systemic influences in utero has been overlooked. In this study, we investigated the effect of multigenerational obesity on epithelial cell migration and invasion using decellularized breast tissues explanted from normal female mouse pups from a diet induced multigenerational obesity mouse model. We first studied the effect of multigenerational diet on the mechanical properties, adipocyte size, and collagen structure of these mouse breast tissues, and then, examined the migration and invasion behavior of normal (KTB-21) and cancerous (MDA-MB-231) human mammary epithelial cells on the decellularized matrices from each diet group. Breast tissues of mice whose dams had been fed with high-fat diet exhibited larger adipocytes and thicker and curvier collagen fibers, but only slightly elevated elastic modulus and inflammatory cytokine levels. MDA-MB-231 cancer cell motility and invasion were significantly greater on the decellularized matrices from mice whose dams were fed with high-fat diet. A similar trend was observed with normal KTB-21 cells. Our results showed that the collagen curvature was the dominating factor on this enhanced motility and stretching the matrices to equalize the collagen fiber linearity of the matrices ameliorated the observed increase in cell migration and invasion in the mice that were exposed to a high-fat diet. Previous studies indicated an increase in serum leptin concentration for those children born to an obese mother. We generated extracellular matrices using primary fibroblasts exposed to various concentrations of leptin. This produced curvier ECM and increased breast cancer cell motility for cells seeded on the decellularized ECM generated with increasing leptin concentration. Our study shows that exposure to obesity in utero is influential in determining the extracellular matrix structure, and that the resultant changes in collagen curvature is a critical factor in regulating the migration and invasion of breast cancer cells.

The Utility of Ancestral and Derived Allele Sharing for Genome-Wide Inferences of Introgression

Peede, D. — 2022-12-02

The past decade has ushered in a resurgence of studies highlighting the importance of introgression throughout the Tree of Life. Several methods exist for detecting and quantifying introgression on a genomic scale, yet the majority of these methods primarily utilize signals of derived allele sharing between donor and recipient populations. In this study, we exploit the fact that introgression will not only result in derived allele sharing but also the reintroduction of ancestral alleles to derive new estimators of the admixture proportion. Using coalescent simulations, we assess the performance of our new methods and the methods proposed in Lopez Fang et al. 2022 to assess the utility of incorporating shared ancestral variation into genome-wide inferences of introgression. Using coalescent theory, simulations, and applying our methods to human and canid data, we find that methods incorporating ancestral allele sharing are comparable to their derived allele sharing counterparts, in turn providing researchers with the opportunity to utilize more of the genomic signature of introgression.

Non-allelic homologous recombination driven translocation explains histidine-rich protein 3 deletion mechanism in Plasmodium falciparum

Hathaway, N. J. — 2022-12-08

Most malaria rapid diagnostic tests (RDTs) detect Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and PfHRP3, but deletions of pfhrp2 and phfrp3 genes make parasites undetectable by RDTs. We analyzed 19,313 public whole-genome-sequenced P. falciparum field samples to understand these deletions better. Pfhrp2 deletion only occurred by chromosomal breakage with subsequent telomere healing. Pfhrp3 deletions involved loss from pfhrp3 to the telomere and showed 3 patterns: no other associated rearrangement with evidence of telomere healing at breakpoint (Asia; Pattern 13-TARE1); associated with duplication of a chromosome 5 segment containing multidrug-resistant-1 gene (Asia; Pattern 13-5++); and most commonly, associated with duplication of a chromosome 11 segment (Americas/Africa; Pattern 13-11++). We confirmed a 13-11 hybrid chromosome with long-read sequencing, consistent with a translocation product arising from recombination between large interchromosomal ribosome-containing segmental duplications. Within most 13-11++ parasites, the duplicated chromosome 11 segments were identical. Across parasites, multiple distinct haplotype groupings were consistent with emergence due to clonal expansion of progeny from intrastrain meiotic recombination. Together, these observations suggest negative selection normally removes 13-11++ pfhrp3 deletions, and specific conditions are needed for their emergence and spread including low transmission, findings that can help refine surveillance strategies.

Structure of Phosphorylated-like ClpXP Adaptor RssB Reveals an Interface Switch for Activation

Brugger, C. — 2022-12-15

SUMMARYIn {gamma}-proteobacteria such as Escherichia coli, the general stress response is mediated by {sigma}s, the stationary phase dissociable promoter specificity subunit of RNA polymerase. {sigma}s is degraded by ClpXP during active growth in a process dependent on the RssB adaptor, which acts catalytically and is thought to be stimulated by phosphorylation of a conserved aspartate in its N-terminal receiver domain. Here we present the crystal structure of full-length RssB bound to a beryllofluoride phosphomimic. Compared to the inhibited IraD anti-adaptor-bound RssB structure, our study reveals movements and coil-to-helix transitions in the C-terminal region of the RssB receiver domain and in the inter-domain segmented helical linker, accompanied by packing of the C-terminal effector domain onto the 4-{beta}5-5 (4-5-5) "signaling" face of the RssB receiver domain. This face is often the locus of protein-protein interactions in unphosphorylated receiver domains, but its masking is unusual in their phosphorylated forms. Our structure emphasizes the remarkable plasticity that underpins regulatory strategies within the large family of response regulators.

Deep learning-based predictive identification of functional subpopulations of hematopoietic stem cells and multipotent progenitors

Wang, S. — 2022-12-20

Hematopoietic stem cells (HSCs) and multipotent progenitors (MPPs) are crucial for maintaining lifelong hematopoiesis. Developing methods to distinguish stem cells from other progenitors and evaluate stem cell functions has been a central task in stem cell research. Deep learning has been demonstrated as a powerful tool in cell image analysis and classification. In this study, we explored the possibility of using deep learning to differentiate HSCs and MPPs based on their light microscopy (DIC) images. After extensive training and validation with large image data sets, we successfully develop a three-class classifier (we named it the LSM model) that reliably differentiate long-term HSCs (LT-HSCs), short-term HSCs (ST-HSCs), and MPPs. Importantly, we demonstrated that our LSM model achieved its differentiating capability by learning the intrinsic morphological features from cell images. Furthermore, we showed that the performance of our LSM model was not affected by how these cells were identified and isolated, i.e., sorted by surface markers or intracellular GFP markers. Prospective identification of HSCs and MPPs in Evi1GFP transgenic mice by LSM model suggested that the cells with the highest GFP expression were LT-HSCs, and this prediction was substantiated later by a long-term competitive reconstitution assay. Moreover, based on DIC image data sets, we also successfully built another two-class classifier that can effectively distinguish aged HSCs from young HSCs, which both express the same surface markers but are functionally different. This finding is of particular interest since it may provide a novel quick and efficient approach, obviating the need for a time-consuming transplantation experiment, to evaluate the functional states of HSCs. Together, our study provides evidence for the first time that HSCs and MPPs can be differentiated by deep learning based on cell morphology. This novel and robust deep learning-based platform will provide a basis for the future development of a new generation stem cell identification and separation system. It may also provide new insight into molecular mechanisms underlying the self-renewal feature of stem cells.

Multiomic Analysis of Adult Diapause in Drosophila melanogaster Identifies Hallmarks of Cellular Quiescence

Hutfilz, C. — 2022-12-21

Age is a fundamental aspect of biology that underlies the efficacy of a broad range of functions. Identifying determinants for how quickly or slowly we age will contribute greatly to our understanding of age as a modifier of overall health, particularly to the advancement of therapeutic interventions designed to mitigate or delay age-associated disorders. While much work has been devoted to the study of genetic or pharmacological interventions that extend lifespan, this approach does not necessarily recapitulate the physiological profile of naturally long-lived individuals. Diapause and diapause-like states constitute natural, inducible and evolutionarily conserved examples of lifespan plasticity that are well-suited to serve as physiologically accurate models of longevity. Here, we leveraged a metabolically critical signaling organ in Drosophila, the fat body, to examine diapause-associated transcription in the context of chromatin accessibility and the regulation of lifespan. Through a combination of ATAC-seq and RNA-seq, our observations suggest chromatin is globally reorganized in diapause and may assume a poised conformation to facilitate the rapid transcription of pro-development genes upon diapause termination. We found particular significance of GAF, NELF, and RNA polymerase III in this context. Congruently, transcription during diapause appears to favor many processes supporting the maintenance of cellular quiescence and the inhibition of differentiation. Our data are consistent with a model wherein diapause induces cellular quiescence in the fat body, as was additionally supported through fluorescent microscopy and comparison with public ChIP-seq data for developmentally juvenile files. This work opens the possibility that longevity in diapause may be partially determined through a lack of mitogenic signaling from the quiescent niche, concurrent with changes to the hormonal and immunological profiles that skew metabolism towards tissue maintenance.

Identification and characterization of small molecule inhibitors of the LINE-1 retrotransposon endonuclease

D'Ordine, A. M. — 2022-12-29

The long interspersed nuclear element-1 (LINE-1 or L1) retrotransposon is the only active autonomously replicating retrotransposon in the human genome. While most of the approximately 500,000 L1 copies are no longer able to propagate, those that retain activity can harm the cell by creating mutations, generating DNA damage, and triggering the expression of inflammatory factors such as the host interferon anti-viral response. Therefore, inhibition of L1 could be used to treat a variety of diseases associated with these processes. Previous research has focused on inhibition of the L1 reverse transcriptase (RT) activity, in part due to the prevalence of well-characterized existing inhibitors to related viral enzymes. Here we present the L1 endonuclease (EN) as an additional target for reducing the detrimental effects of L1 expression. We have screened and characterized a set of structurally diverse small molecule EN inhibitors using computational, biochemical, and cellular methods. We also show that these inhibitors reduce DNA damage created by L1 and inflammation reinforced by L1 activity in senescent cells. These inhibitors could be further used to modulate endogenous L1 function to better understand the lifecycle of this ubiquitous disease-relevant element.

Expanded tRNA methyltransferase family member TRMT9B regulates synaptic growth and function

Hogan, C. A. — 2022-12-31

Nervous system function relies on the formation and function of synaptic connections between neurons. Through a genetic screen in Drosophila for new conserved synaptic genes, we identified CG42261/Fid/ TRMT9B as a negative regulator of synaptogenesis. TRMT9B has been studied for its role as a tumor suppressor in multiple carcinomas and is one of two metazoan homologs of yeast tRNA methyltransferase 9 (Trm9), which methylates tRNA wobble uridines. Members of the expanded family of tRNA methyltransferases are increasingly being associated with neurological disorders and new biochemical functions. Interestingly, whereas Trm9 homolog ALKBH8/CG17807 is ubiquitously expressed, we find that TRMT9B is enriched in the nervous system, including at synapses. However, in the absence of animal models the role of TRMT9B in the nervous system has remained unknown. Here, we generated null alleles of TRMT9B and ALKBH8, and through liquid chromatography-mass spectrometry find that ALKBH8 is responsible for canonical tRNA wobble uridine methylation under basal conditions. In the nervous system, we find that TRMT9B negatively regulates synaptogenesis through a methyltransferase-dependent mechanism in agreement with our modeling studies. Finally, we find that neurotransmitter release is impaired in TRMT9B mutants. Our findings reveal a role for TRMT9B in regulating synapse formation and function, and highlight the importance of the expanded family of tRNA methyltransferases in the nervous system.

IraM Remodels the RssB Segmented Helical Linker to Stabilize σs against Degradation by ClpXP

Brugger, C. — 2023-01-08

Upon Mg2+ starvation, a condition often associated with virulence, enterobacteria inhibit the ClpXP-dependent proteolysis of the master transcriptional regulator, {sigma}s, via IraM, a poorly understood anti-adaptor that prevents RssB-dependent loading of {sigma}s onto ClpXP. This inhibition results in {sigma}s accumulation, and expression of stress resistance genes. Here we report on the structural analysis of RssB bound to IraM, which reveals that IraM induces two folding transitions within RssB, which are amplified via a segmental helical linker. This work highlights the remarkable structural plasticity of RssB and reveals how a stress-specific RssB antagonist modulates a core stress response pathway that could be leveraged to control biofilm formation, virulence, and the development of antibiotic resistance.

Human-specific features and developmental dynamics of the brain N-glycome

Klaric, T. S. — 2023-01-11

Comparative "omics" studies have revealed unique aspects of human neurobiology, yet an evolutionary perspective of the brain N-glycome is lacking. Here, we performed multi-regional characterization of rat, macaque, chimpanzee, and human brain N-glycomes using chromatography and mass spectrometry, then integrated these data with complementary glycotranscriptomic data. We found that in primates the brain N-glycome has evolved more rapidly than the underlying transcriptomic framework, providing a mechanism for generating additional diversity. We show that brain N-glycome evolution in hominids has been characterized by an increase in complexity and (2-6)-linked N-acetylneuraminic acid along with human-specific cell-type expression of key glycogenes. Finally, by comparing the prenatal and adult human brain N-glycome, we identify region-specific neurodevelopmental pathways that lead to distinct spatial N-glycosylation profiles in the mature brain. One-Sentence SummaryEvolution of the human brain N-glycome has been marked by an increase in complexity and a shift in sialic acid linkage.

Uncertainty aversion predicts the neural expansion of semantic representations

Vives, M.-L. — 2023-01-13

Correctly identifying the meaning of a stimulus requires activating the appropriate semantic representation among many alternatives. One way to reduce this uncertainty is to differentiate semantic representations from each other, thereby expanding the semantic space. In four experiments, we test this semantic-expansion hypothesis, finding that uncertainty averse individuals exhibit increasingly differentiated and separated semantic representations. This effect is mirrored at the neural level, where uncertainty aversion predicts greater distances between activity patterns in the left inferior frontal gyrus when reading words, and enhanced sensitivity to the semantic ambiguity of these words in the ventromedial prefrontal cortex. Two direct tests of the behavioral consequences of semantic-expansion further reveal that uncertainty averse individuals exhibit reduced semantic interference and poorer generalization. Together, these findings demonstrate that the internal structure of our semantic representations is shaped in a principled manner: aversion to uncertainty acts as an organizing principle to make the world more identifiable.

The impact of modern admixture on archaic human ancestry in human populations

Witt, K. E. — 2023-01-18

Admixture, the genetic merging of parental populations resulting in mixed ancestry, has occurred frequently throughout the course of human history. Numerous admixture events have occurred between human populations across the world, as well as introgression between humans and archaic humans, Neanderthals and Denisovans. One example are genomes from populations in the Americas, as these are often mosaics of different ancestries due to recent admixture events as part of European colonization. In this study, we analyzed admixed populations from the Americas to assess whether the proportion and location of admixed segments due to recent admixture impact an individuals archaic ancestry. We identified a positive correlation between non-African ancestry and archaic alleles, as well as a slight enrichment of Denisovan alleles in Indigenous American segments relative to European segments in admixed genomes. We also identify several genes as candidates for adaptive introgression, based on archaic alleles present at high frequency in admixed American populations but low frequency in East Asian populations. These results provide insights into how recent admixture events between modern humans redistributed archaic ancestry in admixed genomes.

More than germ cells: vascular development in the early zebrafish (Danio rerio) gonad

Kossack, M. E. — 2023-01-20

Zebrafish are routinely used to model reproductive development, function, and disease, yet we still lack a clear understanding of the fundamental steps that occur during early bipotential gonad development, including when endothelial cells, pericytes, and macrophage cells arrive at the bipotential gonad to support gonad growth and differentiation. Here, we use a combination of transgenic reporters and single-cell sequencing analyses to define the arrival of different critical cell types to the larval zebrafish gonad. We determined that blood initially reaches the gonad via a vessel formed from the swim bladder artery, which we have termed the gonadal artery. We find that vascular and lymphatic development occurs concurrently in the bipotential zebrafish gonad and our data suggest that similar to what has been observed in developing zebrafish embryos, lymphatic endothelial cells in the gonad may be derived from vascular endothelial cells. We mined preexisting sequencing data sets to determine whether ovarian pericytes had unique gene expression signatures. We identified 215 genes that were uniquely expressed in ovarian pericytes that were not expressed in larval pericytes. Similar to what has been shown in the mouse ovary, our data suggest that pdgfrb+ pericytes may support the migration of endothelial tip cells during ovarian angiogenesis. Using a macrophage-driven photoconvertible protein, we found that macrophage established a nascent resident population as early as 12 dpf and can be observed removing cellular material during gonadal differentiation. This foundational information demonstrates that the early bipotential gonad contains complex cellular interactions, which likely shape the health and function of the mature, differentiated gonad. Summary SentenceDelineating the complex cellular interactions between vascular and lymphatic endothelial cells, pericytes, and macrophage in the bipotential gonad is essential for understanding the differentiation and functioning of the mature gonad. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/524593v2_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@23fff5org.highwire.dtl.DTLVardef@179612forg.highwire.dtl.DTLVardef@1db91f0org.highwire.dtl.DTLVardef@4a4557_HPS_FORMAT_FIGEXP M_FIG C_FIG

Phantom controllers: Misspecified models create the false appearance of adaptive control during value-based choice

Ritz, H. — 2023-01-20

Decision scientists have grown increasingly interested in how people adaptively control their decision making, exploring how metacognitive factors like uncertainty or conflict influence how people accumulate evidence and commit to a choice. For example, a recent study proposed that options values contribute to both the formation of a decision and the effortful invigoration of a response. Here, the control process was operationalized in a drift diffusion model as the lowering of the decision threshold on difficult trials. Reanalyzing the data from this experiment, we establish alternative explanations for these findings. We show that the reported evidence for controlled threshold adjustments can be explained away by task confounds, time-dependent collapses in decision thresholds, and stimulus-driven dynamics in alternative decision frameworks. With regard to the individual study, our findings challenge the reported evidence for this new theory of motivated control. More generally, we reveal paths and pitfalls in computational approaches to understanding when and how control guides decision-making.

Investigating the Complexity of Gene Co-expression Estimation for Single-cell Data

Zhang, J. — 2023-01-25

AO_SCPLOWBSTRACTC_SCPLOWWith the rapid advance of single-cell RNA sequencing (scRNA-seq) technology, understanding biological processes at a more refined single-cell level is becoming possible. Gene co-expression estimation is an essential step in this direction. It can annotate functionalities of unknown genes or construct the basis of gene regulatory network inference. This study thoroughly tests the existing gene co-expression estimation methods on simulation datasets with known ground truth co-expression networks. We generate these novel datasets using two simulation processes that use the parameters learned from the experimental data. We demonstrate that these simulations better capture the underlying properties of the real-world single-cell datasets than previously tested simulations for the task. Our performance results on tens of simulated and eight experimental datasets show that all methods produce estimations with a high false discovery rate potentially caused by high-sparsity levels in the data. Finally, we find that commonly used pre-processing approaches, such as normalization and imputation, do not improve the co-expression estimation. Overall, our benchmark setup contributes to the co-expression estimator development, and our study provides valuable insights for the community of single-cell data analyses.

Structural variation across 138,134 samples in the TOPMed consortium

Jun, G. — 2023-01-26

Ever larger Structural Variant (SV) catalogs highlighting the diversity within and between populations help researchers better understand the links between SVs and disease. The identification of SVs from DNA sequence data is non-trivial and requires a balance between comprehensiveness and precision. Here we present a catalog of 355,667 SVs (59.34% novel) across autosomes and the X chromosome (50bp+) from 138,134 individuals in the diverse TOPMed consortium. We describe our methodologies for SV inference resulting in high variant quality and >90% allele concordance compared to long-read de-novo assemblies of well-characterized control samples. We demonstrate utility through significant associations between SVs and important various cardio-metabolic and hemotologic traits. We have identified 690 SV hotspots and deserts and those that potentially impact the regulation of medically relevant genes. This catalog characterizes SVs across multiple populations and will serve as a valuable tool to understand the impact of SV on disease development and progression.

Long-term unsupervised recalibration of cursor BCIs

Wilson, G. H. — 2023-02-04

Intracortical brain-computer interfaces (iBCIs) require frequent recalibration to maintain robust performance due to changes in neural activity that accumulate over time. Compensating for this nonstationarity would enable consistently high performance without the need for supervised recalibration periods, where users cannot engage in free use of their device. Here we introduce a hidden Markov model (HMM) to infer what targets users are moving toward during iBCI use. We then retrain the system using these inferred targets, enabling unsupervised adaptation to changing neural activity. Our approach outperforms the state of the art in large-scale, closed-loop simulations over two months and in closed-loop with a human iBCI user over one month. Leveraging an offline dataset spanning five years of iBCI recordings, we further show how recently proposed data distribution-matching approaches to recalibration fail over long time scales; only target-inference methods appear capable of enabling long-term unsupervised recalibration. Our results demonstrate how task structure can be used to bootstrap a noisy decoder into a highly-performant one, thereby overcoming one of the major barriers to clinically translating BCIs.

An 8-cage imaging system for automated analyses of mouse behavior

Del Rosario Hernandez, T. — 2023-02-04

The analysis of mouse behavior is used in biomedical research to study brain function in health and disease. Well-established rapid assays allow for high-throughput analyses of behavior but have several drawbacks, including measurements of daytime behaviors in a nocturnal animal, effects of animal handling, and the lack of an acclimation period in the testing apparatus. We developed a novel 8-cage imaging system, with animated visual stimuli, for automated analyses of mouse behavior in 22-hour overnight recordings. Software for image analysis was developed in two open-source programs, ImageJ and DeepLabCut. The imaging system was used to measure multiple behaviors, including acclimation to the novel cage environment, day and nighttime activity, stretch-attend postures, location in various cage areas, and habituation to animated visual stimuli. These behaviors were summarized in behavioral profiles, which may be used in further studies to examine treatments for neural disorders.

GSK-3 inhibitor elraglusib enhances tumor-infiltrating immune cell activation in tumor biopsies and synergizes with anti-PD-L1 in a murine model of colorectal cancer

Huntington, K. E. — 2023-02-07

Inhibition of GSK-3 using small-molecule elraglusib has shown promising preclinical antitumor activity. Using in vitro systems, we found that elraglusib promotes immune cell-mediated tumor cell killing, enhances tumor cell pyroptosis, decreases tumor cell NF-{kappa}B-regulated survival protein expression, and increases immune cell effector molecule secretion. Using in vivo systems, we observed synergy between elraglusib and anti-PD-L1 in an immunocompetent murine model of colorectal cancer. Murine responders had more tumor-infiltrating T-cells, fewer tumor-infiltrating Tregs, lower tumorigenic circulating cytokine concentrations, and higher immunostimulatory circulating cytokine concentrations. To determine the clinical significance, we utilized human plasma samples from patients treated with elraglusib and correlated cytokine profiles with survival. Using paired tumor biopsies, we found that CD45+ tumor-infiltrating immune cells had lower expression of inhibitory immune checkpoints and higher expression of T-cell activation markers in post-elraglusib patient biopsies. These results introduce several immunomodulatory mechanisms of GSK-3 inhibition using elraglusib, providing a rationale for the clinical evaluation of elraglusib in combination with immunotherapy. Statement of significancePharmacologic inhibition of GSK-3 using elraglusib sensitizes tumor cells, activates immune cells for increased anti-tumor immunity, and synergizes with anti-PD-L1 immune checkpoint blockade. These results introduce novel biomarkers for correlations with response to therapy which could provide significant clinical utility and suggest that elraglusib, and other GSK-3 inhibitors, should be evaluated in combination with immune checkpoint blockade.

Hypoxia-inducible factor 2 is a key determinant of manganese excess and polycythemia in SLC30A10 deficiency

Prajapati, M. — 2023-02-21

Manganese is an essential yet potentially toxic metal. Initially reported in 2012, mutations in SLC30A10 are the first known inherited cause of manganese excess. SLC30A10 is an apical membrane transport protein that exports manganese from hepatocytes into bile and from enterocytes into the lumen of the gastrointestinal tract. SLC30A10 deficiency results in impaired gastrointestinal manganese excretion, leading to severe manganese excess, neurologic deficits, liver cirrhosis, polycythemia, and erythropoietin excess. Neurologic and liver disease are attributed to manganese toxicity. Polycythemia is attributed to erythropoietin excess, but the basis of erythropoietin excess in SLC30A10 deficiency has yet to be established. Here we demonstrate that erythropoietin expression is increased in liver but decreased in kidneys in Slc30a10-deficient mice. Using pharmacologic and genetic approaches, we show that liver expression of hypoxia-inducible factor 2 (Hif2), a transcription factor that mediates the cellular response to hypoxia, is essential for erythropoietin excess and polycythemia in Slc30a10-deficient mice, while hypoxia-inducible factor 1 (HIF1) plays no discernible role. RNA-seq analysis determined that Slc30a10-deficient livers exhibit aberrant expression of a large number of genes, most of which align with cell cycle and metabolic processes, while hepatic Hif2 deficiency attenuates differential expression of half of these genes in mutant mice. One such gene downregulated in Slc30a10-deficient mice in a Hif2-dependent manner is hepcidin, a hormonal inhibitor of dietary iron absorption. Our analyses indicate that hepcidin downregulation serves to increase iron absorption to meet the demands of erythropoiesis driven by erythropoietin excess. Finally, we also observed that hepatic Hif2 deficiency attenuates tissue manganese excess, although the underlying cause of this observation is not clear at this time. Overall, our results indicate that HIF2 is a key determinant of pathophysiology in SLC30A10 deficiency. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=148 HEIGHT=200 SRC="FIGDIR/small/529270v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@c25fc8org.highwire.dtl.DTLVardef@11e70feorg.highwire.dtl.DTLVardef@18c5518org.highwire.dtl.DTLVardef@26b61f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Bacillus subtilis engineered for aerospace medicine: a platform for off-planet production of pharmaceutical peptides

Vallota-Eastman, A. — 2023-02-22

1.Biologics, such as pharmaceutical peptides, have notoriously short shelf lives, insufficient for long-duration space flight missions to the Moon or Mars. To enable the sustainable presence of humans on the Moon or Mars, we must develop methods for on-site production of pharmaceutical peptides in space, a concept we call Astropharmacy. Here, we present proof-of-concept for the first step needed: a low-mass system for pharmaceutical production designed to be stable in space. To demonstrate feasibility, we engineered strains of the space-hardy spore-forming bacterium, Bacillus subtilis, to secrete two pharmaceutical peptides important for astronaut health: teriparatide (an anabolic agent for combating osteoporosis) and filgrastim (an effective countermeasure for radiation-induced neutropenia). We found that the secretion peptides from the walM and yoqH genes of B. subtilis 168 worked well for secreting teriparatide and filgrastim, respectively. In consideration of the TRISH challenge to produce a dose equivalent in 24 hours, dried spores of our engineered strains were used to produce 1 dose equivalent of teriparatide from a 2 mL culture and 1 dose equivalent of filgrastim from 52 mL of culture in 24 hours. Further optimization of strain growth conditions, expression conditions, and promoter sequences should allow higher production rates to be achieved. These strains provide the template for future optimization efforts and address the first step in the Astropharmacy, capable of on-site production, purification, and processing of biopharmaceutical compounds in platforms amenable for use in space.

Hippocampal engrams generate flexible behavioral responses and brain-wide network states

Dorst, K. E. — 2023-02-24

Memory engrams are both necessary and sufficient to mediate behavioral outputs. Defensive behaviors such as freezing and avoidance are commonly examined during hippocampal-mediated fear engram reactivation, yet how reactivation of these cellular populations across different contexts engages the brain to produce a variety of defensive behaviors is relatively unclear. To address this, we first optogenetically reactivated a tagged fear engram in the dentate gyrus (DG) subregion of the hippocampus across three distinct contexts. We found that there were differential amounts of light-induced freezing depending on the size of the context in which reactivation occurred: mice demonstrated robust light-induced freezing in the most spatially restricted of the three contexts but not in the largest. We then utilized graph theoretical analyses to identify brain-wide alterations in cFos co-activation during engram reactivation across the smallest and largest contexts. Our manipulations conferred greater positive cFos correlations and recruited regions spanning putative fear and defense systems as hubs in the respective networks. Moreover, reactivating DG-mediated engrams generated network topologies across experimental conditions, emphasizing both shared and distinct features. By identifying and manipulating the circuits supporting memory function, as well as their corresponding brain-wide activity patterns, it is thereby possible to resolve systems-level biological mechanisms mediating memorys capacity to modulate behavioral states. SIGNIFICANCE STATEMENTImplementing appropriate defensive behaviors across disparate environments is essential for survival. Memories can be used to select these responses. Recent work identified and artificially manipulated cellular ensembles within the hippocampus that mediate fear memory recall, yet how these populations engage brain-wide pathways that mediate defensive behaviors under environmental contingencies is unclear. We demonstrated here that reactivation across environments of various sizes elicits different behavioral responses and corresponding brain-wide network dynamics. These findings establish the flexibility of memory-bearing ensembles in generating brain and behavior states.

Exposure to the persistent organic pollutant 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD, dioxin) disrupts development of the zebrafish inner ear

Cintron-Rivera, L. G. — 2023-03-15

Dioxins are a class of highly toxic and persistent environmental pollutants that have been shown through epidemiological and laboratory-based studies to act as developmental teratogens. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the most potent dioxin congener, has a high affinity for the aryl hydrocarbon receptor (AHR), a ligand activated transcription factor. TCDD-induced AHR activation during development impairs nervous system, cardiac, and craniofacial development. Despite the robust phenotypes previously reported, the characterization of developmental malformations and our understanding of the molecular targets mediating TCDD-induced developmental toxicity remains limited. In zebrafish, TCDD-induced craniofacial malformations are produced, in part, by the downregulation of SRY-box transcription factor 9b (sox9b), a member of the SoxE gene family. sox9b, along with fellow SoxE gene family members sox9a and sox10, have important functions in the development of the otic placode, the otic vesicle, and, ultimately, the inner ear. Given that sox9b in a known target of TCDD and that transcriptional interactions exist among SoxE genes, we asked whether TCDD exposure impaired the development of the zebrafish auditory system, specifically the otic vesicle, which gives rise to the sensory components of the inner ear. Using immunohistochemistry, in vivo confocal imaging, and time-lapse microscopy, we assessed the impact of TCDD exposure on zebrafish otic vesicle development. We found exposure resulted in structural deficits, including incomplete pillar fusion and altered pillar topography, leading to defective semicircular canal development. The observed structural deficits were accompanied by reduced collagen type II expression in the ear. Together, our findings reveal the otic vesicle as a novel target of TCDD-induced toxicity, suggest that the function of multiple SoxE genes may be affected by TCDD exposure, and provide insight into how environmental contaminants contribute to congenital malformations. HighlightsO_LIThe zebrafish ear is necessary to detect changes in motion, sound, and gravity. C_LIO_LIEmbryos exposed to TCDD lack structural components of the developing ear. C_LIO_LITCDD exposure impairs formation of the fusion plate and alters pillar topography. C_LIO_LIThe semicircular canals of the ear are required to detect changes in movement. C_LIO_LIFollowing TCDD exposure embryos fail to establish semicircular canals. C_LI

Opposing, spatially-determined epigenetic forces impose restrictions on stochastic olfactory receptor choice

Bashkirova, E. — 2023-03-15

Olfactory receptor (OR) choice represents an example of genetically hardwired stochasticity, where every olfactory neuron expresses one out of [~]2000 OR alleles in a probabilistic, yet stereotypic fashion. Here, we propose that topographic restrictions in OR expression are established in neuronal progenitors by two opposing forces: polygenic transcription and genomic silencing, both of which are influenced by dorsoventral gradients of transcription factors NFIA, B, and X. Polygenic transcription of OR genes may define spatially constrained OR repertoires, among which one OR allele is selected for singular expression later in development. Heterochromatin assembly and genomic compartmentalization of OR alleles also vary across the axes of the olfactory epithelium and may preferentially eliminate ectopically expressed ORs with more dorsal expression destinations from this "privileged" repertoire. Our experiments identify early transcription as a potential "epigenetic" contributor to future developmental patterning and reveal how two spatially responsive probabilistic processes may act in concert to establish deterministic, precise, and reproducible territories of stochastic gene expression.

A novel miR-99b-5p-Zbp1 pathway in microglia contributes to the pathogenesis of schizophrenia

Kaurani, L. — 2023-03-21

Schizophrenia is a psychiatric disorder that is still not readily treatable. Pharmaceutical advances in the treatment of schizophrenia have mainly focused on the protein coding part of the human genome. However, the vast majority of the human transcriptome consists of non-coding RNAs. MicroRNAs are small non-coding RNAs that control the transcriptome at the systems level. In the present study we analyzed the microRNAome in blood and postmortem brains of controls and schizophrenia patients and found that miR-99b-5p was downregulated in both the prefrontal cortex and blood of patients. At the mechanistic level we show that inhibition of miR-99b-5p leads to schizophrenia-like phenotypes in mice and induced inflammatory processes in microglia linked to synaptic pruning. The miR-99b-5p-mediated inflammatory response in microglia depended on Z-DNA binding protein 1 (Zbp1) which we identified as a novel miR-99b-5p target. Antisense oligos (ASOs) against Zbp1 ameliorated the pathological phenotypes caused by miR-99b-5p inhibition. In conclusion, we report a novel miR-99b-5p-Zbp1 pathway in microglia that contributes to the pathogenesis of schizophrenia. Our data suggest that strategies to increase the levels of miR-99b-5p or inhibit Zbp1 could become a novel therapeutic strategy.

Transient gamma events delineate somatosensory modality in S1

Black, C. J. — 2023-03-31

Gamma band activity localized to the primary somatosensory cortex (S1) in humans and animals is implicated in the higher order neural processing of painful and tactile stimuli. However, it is unclear if gamma band activity differs between these distinct somatosensory modalities. Here, we coupled a novel behavioral approach with chronic extracellular electrophysiology to investigate differences in S1 gamma band activity elicited by noxious and innocuous hind paw stimulation in transgenic mice. Like prior studies, we found that trial-averaged gamma power in S1 increased following both noxious and innocuous stimuli. However, on individual trials, we noticed that evoked gamma band activity was not a continuous oscillatory signal but a series of transient spectral events. Upon further analysis we found that there was a significantly higher incidence of these gamma band events following noxious stimulation than innocuous stimulation. These findings suggest that somatosensory stimuli may be represented by specific features of gamma band activity at the single trial level, which may provide insight to mechanisms underlying acute pain.

Molecular and organizational diversity intersect to generate functional synaptic heterogeneity within and between excitatory neuronal subtypes

Medeiros, A. T. — 2023-04-02

Synaptic heterogeneity is a hallmark of nervous systems that enables complex and adaptable communication in neural circuits. To understand circuit function, it is thus critical to determine the factors that contribute to the functional diversity of synapses. We investigated the contributions of voltage-gated calcium channel (VGCC) abundance, spatial organization, and subunit composition to synapse diversity among and between synapses formed by two closely related Drosophila glutamatergic motor neurons with distinct neurotransmitter release probabilities (Pr). Surprisingly, VGCC levels are highly predictive of heterogeneous Pr among individual synapses of either low- or high-Pr inputs, but not between inputs. We find that the same number of VGCCs are more densely organized at high-Pr synapses, consistent with tighter VGCC-synaptic vesicle coupling. We generated endogenously tagged lines to investigate VGCC subunits in vivo and found that the 2{delta}-3 subunit Straightjacket along with the CAST/ELKS active zone (AZ) protein Bruchpilot, both key regulators of VGCCs, are less abundant at high-Pr inputs, yet positively correlate with Pr among synapses formed by either input. Consistently, both Straightjacket and Bruchpilot levels are dynamically increased across AZs of both inputs when neurotransmitter release is potentiated to maintain stable communication following glutamate receptor inhibition. Together, these findings suggest a model in which VGCC and AZ protein abundance intersects with input-specific spatial and molecular organization to shape the functional diversity of synapses.

Transsynaptic labeling and transcriptional control of zebrafish neural circuits

Coomer, C. — 2023-04-03

Deciphering the connectome, the ensemble of synaptic connections that underlie brain function, is a central goal of neuroscience research. Here, we report mapping of connections between presynaptic and postsynaptic partners in a living vertebrate nervous system, that of the zebrafish, through the successful adaptation of the trans-Tango genetic approach, first developed for anterograde transsynaptic tracing in Drosophila. Neural connections were visualized between synaptic partners in the larval retina and brain and followed over development. Results were corroborated by functional experiments in which optogenetic activation of retinal ganglion cells elicited responses in neurons of the optic tectum, as measured by trans-Tango-dependent expression of a genetically encoded calcium indicator. Transsynaptic signaling through trans-Tango reveals predicted as well as previously undescribed synaptic connections in the zebrafish brain, providing a valuable in vivo tool to monitor and interrogate neural circuits over time.

Differential regulation of Retinoic Acid Metabolism in Fanconi Anemia

Blaize, J. — 2023-04-06

Fanconi anemia (FA) is a rare genetic disease characterized by heterogeneous congenital abnormalities and increased risk for bone marrow failure and cancer. FA is caused by mutation of any one of 23 genes, the protein products of which function primarily in the maintenance of genome stability. An important role for the FA proteins in the repair of DNA interstrand crosslinks (ICLs) has been established in vitro. While the endogenous sources of ICLs relevant to the pathophysiology of FA have yet to be clearly determined, a role for the FA proteins in a two-tier system for the detoxification of reactive metabolic aldehydes has been established. To discover new metabolic pathways linked to FA, we performed RNA-seq analysis on non-transformed FA-D2 (FANCD2-/-) and FANCD2-complemented patient cells. Multiple genes associated with retinoic acid metabolism and signaling were differentially expressed in FA-D2 (FANCD2-/-) patient cells, including ALDH1A1 and RDH10, which encode for retinaldehyde and retinol dehydrogenases, respectively. Increased levels of the ALDH1A1 and RDH10 proteins was confirmed by immunoblotting. FA-D2 (FANCD2-/-) patient cells displayed increased aldehyde dehydrogenase activity compared to the FANCD2-complemented cells. Upon exposure to retinaldehyde, FA-D2 (FANCD2-/-) cells exhibited increased DNA double-strand breaks and checkpoint activation indicative of a defect in the repair of retinaldehyde-induced DNA damage. Our findings describe a novel link between retinoic acid metabolism and FA and identify retinaldehyde as an additional reactive metabolic aldehyde relevant to the pathophysiology of FA.

Engineering GliaTrap: a biodegradable non-swelling hydrogel with tuned release of CXCL12 to attract migrating glioblastoma cells

Suita, Y. — 2023-04-13

Glioblastoma is the most aggressive type of brain cancer with an average overall survival of 15-21 months after first diagnosis. The relapse is mainly caused by migrating glioblastoma cells that diffuse away from the tumor mass into the brain parenchyma and retain cancer stem cell (GSC) properties. Current therapeutic options are ineffective and inevitably result in relapse, indicating a high unmet medical need for innovative therapies in the treatment of invasive glioblastoma. To address this challenge, we propose a new therapeutic modality: GliaTrap, a biodegradable non-swelling, injectable hydrogel with sustained release of a chemoattractant for GSCs that lures and traps the migrating cells back to the tumor resection cavity. We developed a biodegradable and injectable hyaluronan/collagen II-based (HA/Col) hydrogel that does not swell in vivo. The hydrogel is embedded with CXCL12 loaded liposomes and is tuned for sustained release of CXCL12. The safety profile of liposome-embedded HA/Col hydrogel was determined in-vivo after stereotactic implantation in the mouse brain. The efficacy of GliaTrap to attract GSCs was determined ex vivo using a 3D tumor spheroid model and in-vivo using 3D light-sheet microscopy in orthotopic human glioblastoma xenografts. Our findings suggest that GliaTrap could represent a safe and efficacious new therapeutic approach for glioblastoma and potentially serve as a drug delivery platform to locally deliver tumor-killing agents. One Sentence SummaryGliaTrap is a biodegradable non-swelling hydrogel with tuned release of a chemoattractant to attract invading glioma cells and serve as delivery platform for local therapeutics.

Methods and considerations for estimating parameters in biophysically detailed neural models with simulation based inference

Tolley, N. — 2023-04-17

Biophysically detailed neural models are a powerful technique to study neural dynamics in health and disease with a growing number of established and openly available models. A major challenge in the use of such models is that parameter inference is an inherently difficult and unsolved problem. Identifying unique parameter distributions that can account for observed neural dynamics, and differences across experimental conditions, is essential to their meaningful use. Recently, simulation based inference (SBI) has been proposed as an approach to perform Bayesian inference to estimate parameters in detailed neural models. SBI overcomes the challenge of not having access to a likelihood function, which has severely limited inference methods in such models, by leveraging advances in deep learning to perform density estimation. While the substantial methodological advancements offered by SBI are promising, their use in large scale biophysically detailed models is challenging and methods for doing so have not been established, particularly when inferring parameters that can account for time series waveforms. We provide guidelines and considerations on how SBI can be applied to estimate time series waveforms in biophysically detailed neural models starting with a simplified example and extending to specific applications to common MEG/EEG waveforms using the the large scale neural modeling framework of the Human Neocortical Neurosolver. Specifically, we describe how to estimate and compare results from example oscillatory and event related potential simulations. We also describe how diagnostics can be used to assess the quality and uniqueness of the posterior estimates. The methods described provide a principled foundation to guide future applications of SBI in a wide variety of applications that use detailed models to study neural dynamics. Author summaryA central problem in computational neural modeling is estimating model parameters that can account for observed activity patterns. While several techniques exist to perform parameter inference in special classes of abstract neural models, there are comparatively few approaches for large scale biophysically detailed neural models. In this work, we describe challenges and solutions in applying a deep learning based statistical framework to estimate parameters in a biophysically detailed large scale neural model, and emphasize the particular difficulties in estimating parameters for time series data. Our example uses a multi-scale model designed to connect human MEG/EEG recordings to the underlying cell and circuit level generators. Our approach allows for crucial insight into how cell-level properties interact to produce measured neural activity, and provides guidelines for diagnosing the quality of the estimate and uniqueness of predictions for different MEG/EEG biomarkers.

Identifying antisense oligonucleotides to disrupt small RNA regulated antibiotic resistance via a cell-free transcription-translation platform

Tsai, M. J. — 2023-04-20

Bacterial small RNAs (sRNAs) regulate many important physiological processes in cells including antibiotic resistance and virulence genes through base pairing interactions with mRNAs. Antisense oligonucleotides (ASOs) have great potential as therapeutics against bacterial pathogens by targeting sRNAs such as MicF, which regulates outer membrane protein OmpF expression and limits permeability of antibiotics. Here, we devise a cell-free transcription-translation (TX-TL) assay to identify ASO designs that sufficiently sequester MicF. ASOs were then ordered as peptide nucleic acids conjugated to cell-penetrating peptides (CPP-PNA) to allow for effective delivery into bacteria. Subsequent minimum inhibitory concentration (MIC) assays demonstrated that simultaneously targeting the regions of MicF responsible for sequestering the start codon and the Shine-Dalgarno sequence of ompF with two different CPP-PNAs synergistically reduced the MIC for a set of antibiotics. This investigation offers a TX-TL based approach to identify novel therapeutic candidates to combat intrinsic sRNA-mediated antibiotic resistance mechanisms.

Tension sensing by FAK governs nuclear mechanotransduction, endothelial transcriptome and fate

Akhter, M. Z. — 2023-04-26

Vascular endothelium forms a restrictive barrier to defend the underlying tissue against uncontrolled influx of circulating protein and immune cells. Mechanisms that mediate the transition from restrictive to leaky endothelium, a hallmark of tissue injury exemplified by acute lung injury (ALI), remain elusive. Using endothelial cell (EC)-Fak-/-mice, we show that FAK sensing and transmission of mechanical tension to the EC nucleus governs cell fate. In FAK- deleted EC, increased EC tension induced by Rho kinase caused tyrosine phosphorylation of nuclear envelope protein, emerin at Y74/Y95, and its localization in a nuclear cap. Activated emerin stimulated DNMT3a activity and methylation of the KLF2 promoter, impairing the restrictive EC transcriptome, including S1PR1. Inhibiting emerin phosphorylation or DNMT3a activity enabled KLF2 transcription of S1PR1, rescuing the restrictive EC phenotype in EC-Fak-/- lungs. Thus, FAK sensing of tension transmission to the nucleus is crucial for maintaining a restrictive EC fate and lung homeostasis.

Up-regulation of cholesterol synthesis pathways and limited neurodegeneration in a knock-in Sod1 mutant mouse model of ALS.

Dominov, J. A. — 2023-05-05

Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative disorder affecting brain and spinal cord motor neurons. Mutations in the copper/zinc superoxide dismutase gene (SOD1) are associated with [~]20% of inherited and 1-2% of sporadic ALS cases. Much has been learned from mice expressing transgenic copies of mutant SOD1, which typically involve high-level transgene expression, thereby differing from ALS patients expressing one mutant gene copy. To generate a model that more closely represents patient gene expression, we created a knock-in point mutation (G85R, a human ALS-causing mutation) in the endogenous mouse Sod1 gene, leading to mutant SOD1G85R protein expression. Heterozygous Sod1G85R mutant mice resemble wild type, whereas homozygous mutants have reduced body weight and lifespan, a mild neurodegenerative phenotype, and express very low mutant SOD1 protein levels with no detectable SOD1 activity. Homozygous mutants exhibit partial neuromuscular junction denervation at 3-4 months of age. Spinal cord motor neuron transcriptome analyses of homozygous Sod1G85R mice revealed up-regulation of cholesterol synthesis pathway genes compared to wild type. Transcriptome and phenotypic features of these mice are similar to Sod1 knock-out mice, suggesting the Sod1G85R phenotype is largely driven by loss of SOD1 function. By contrast, cholesterol synthesis genes are down-regulated in severely affected human TgSOD1G93A transgenic mice at 4 months. Our analyses implicate dysregulation of cholesterol or related lipid pathway genes in ALS pathogenesis. The Sod1G85R knock-in mouse is a useful ALS model to examine the importance of SOD1 activity in control of cholesterol homeostasis and motor neuron survival. SIGNIFICANCE STATEMENTAmyotrophic lateral sclerosis is a devastating disease involving the progressive loss of motor neurons and motor function for which there is currently no cure. Understanding biological mechanisms leading to motor neuron death is critical for developing new treatments. Using a new knock-in mutant mouse model carrying a Sod1 mutation that causes ALS in patients, and in the mouse, causes a limited neurodegenerative phenotype similar to Sod1 loss-of-function, we show that cholesterol synthesis pathway genes are up-regulated in mutant motor neurons, whereas the same genes are down-regulated in transgenic SOD1 mice with a severe phenotype. Our data implicate dysregulation of cholesterol or other related lipid genes in ALS pathogenesis and provide new insights that could contribute to strategies for disease intervention.

Mutual inclusivity improves decision-making by smoothing out choice's competitive edge

Leng, X. — 2023-05-12

Decisions form a central bottleneck to most tasks, one that people often experience as costly. Past work proposes mitigating those costs by lowering ones threshold for deciding. Here, we test an alternative solution, one that targets the basis for most choice costs: that choosing one option sacrifices others (mutual exclusivity). Across 5 studies (N = 462), we test whether this tension can be relieved by framing choices as inclusive (allowing selection of more than one option, as in buffets). We find that inclusivity makes choices more efficient, by selectively reducing competition between potential responses as participants accumulate information for each of their options. Inclusivity also made participants feel less conflicted, especially when they couldnt decide which good option to keep or which bad option to get rid of. These inclusivity benefits were also distinguishable from the effects of manipulating decision threshold (increased urgency), which improved choices but not experiences thereof.

MuSK-BMP signaling in adult muscle stem cells maintains quiescence and regulates myofiber size

Madigan, L. A. — 2023-05-18

A central question in adult stem cell biology is elucidating the signaling pathways regulating their dynamics and function in diverse physiological and age-related contexts. Muscle stem cells in adults (Satellite Cells; SCs) are generally quiescent but can activate and contribute to muscle repair and growth. Here we tested the role of the MuSK-BMP pathway in regulating adult SC quiescence by deletion of the BMP-binding MuSK Ig3 domain ( {Delta}Ig3-MuSK). At 3 months of age SC and myonuclei numbers and myofiber size were comparable to WT. However, at 5 months of age SC density was decreased while myofiber size, myonuclear number and grip strength were increased - indicating that SCs had activated and productively fused into the myofibers over this interval. Transcriptomic analysis showed that SCs from uninjured {Delta}Ig3-MuSK mice exhibit signatures of activation. Regeneration experiments showed that {Delta}Ig3-MuSK SCs maintain full stem cell function. Expression of {Delta}Ig3-MuSK in adult SCs was sufficient to break quiescence and increase myofiber size. We conclude that the MuSK-BMP pathway regulates SC quiescence and myofiber size in a cell autonomous, age-dependent manner. Targeting MuSK-BMP signaling in muscle stem cells thus emerges a therapeutic strategy for promoting muscle growth and function in the settings of injury, disease, and aging. HighlightsO_LIMuSK, in its role as a BMP co-receptor, regulates adult muscle stem cell quiescence C_LIO_LIThe MuSK-BMP pathway acts cell autonomously C_LIO_LIIncreased muscle size and function with preservation of myonuclear density and stemness in mice with attenuated MuSK-BMP signaling C_LI

Morphogenesis in Trypanosoma cruzi epimastigotes proceeds via a highly asymmetric cell division

Campbell, P. C. — 2023-05-24

Trypanosoma cruzi is a protist parasite that is the causative agent of Chagas disease, a neglected tropical disease endemic to the Americas. T. cruzi cells are highly polarized and undergo morphological changes as they cycle within their insect and mammalian hosts. Work on related trypanosomatids has described cell division mechanisms in several life-cycle stages and identified a set of essential morphogenic proteins that serve as markers for key events during trypanosomatid division. Here, we use Cas9-based tagging of morphogenic genes, live-cell imaging, and expansion microscopy to study the cell division mechanism of the insect-resident epimastigote form of T. cruzi, which represents an understudied trypanosomatid morphotype. We find that T. cruzi epimastigote cell division is highly asymmetric, producing one daughter cell that is significantly smaller than the other. Daughter cell division rates differ by 4.9 h, which may be a consequence of this size disparity. Many of the morphogenic proteins identified in T. brucei have altered localization patterns in T. cruzi epimastigoes, which may reflect fundamental differences in the cell division mechanism of this life cycle stage, which widens and shortens the cell body to accommodate the duplicated organelles and cleavage furrow rather than elongating the cell body along the long axis of the cell, as is the case in life-cycle stages that have been studied in T. brucei. This work provides a foundation for further investigations of T. cruzi cell division and shows that subtle differences in trypansomatid cell morphology can alter how these parasites divide. Author SummaryTrypanosoma cruzi causes Chagas disease, which is among the most neglected of tropical diseases, affecting millions of people in South and Central America along with immigrant populations around the world. T. cruzi is related to other important pathogens such as Trypanosoma brucei and Leishmania spp, which have been the subject of molecular and cellular characterizations that have provided an understanding of how these organisms shape their cells and undergo division. Work in T. cruzi has lagged due to an absence of molecular tools for manipulating the parasite and the complexity of the original published genome; these issues have recently been resolved. Building on work in T. brucei, we have studied the localization of key cell cycle proteins and quantified changes in cell shape during division in an insect-resident form of T. cruzi. This work has uncovered unique adaptations to the cell division process in T. cruzi and provides insight into the range of mechanisms this family of important pathogens can employ to colonize their hosts.

A combined computational and experimental investigation of the filtration function of splenic macrophages in sickle cell disease

li, g. — 2023-06-03

Being the largest lymphatic organ in the body, the spleen also constantly controls the quality of red blood cells (RBCs) in circulation through its two major filtration components, namely interendothelial slits (IES) and red pulp macrophages. In contrast to the extensive studies in understanding the filtration function of IES, there are relatively fewer works on investigating how the splenic macrophages retain the aged and diseased RBCs, i.e., RBCs in sickle cell disease (SCD). Herein, we perform a computational study informed by companion experiments to quantify the dynamics of RBCs captured and retained by the macrophages. We first calibrate the parameters in the computational model based on microfluidic experimental measurements for sickle RBCs under normoxia and hypoxia, as those parameters are not available in the literature. Next, we quantify the impact of a set of key factors that are expected to dictate the RBC retention by the macrophages in the spleen, namely, blood flow conditions, RBC aggregation, hematocrit, RBC morphology, and oxygen levels. Our simulation results show that hypoxic conditions could enhance the adhesion between the sickle RBCs and macrophages. This, in turn, increases the retention of RBCs by as much as five-fold, which could be a possible cause of RBC congestion in the spleen of patients with SCD. Our study on the impact of RBC aggregation illustrates a clustering effect, where multiple RBCs in one aggregate can make contact and adhere to the macrophages, leading to a higher retention rate than that resulting from RBC-macrophage pair interactions. Our simulations of sickle RBCs flowing past macrophages for a range of blood flow velocities indicate that the increased blood velocity could quickly attenuate the function of the red pulp macrophages on detaining aged or diseased RBCs, thereby providing a possible rationale for the slow blood flow in the open circulation of the spleen. Furthermore, we quantify the impact of RBC morphology on their tendency to be retained by the macrophages. We find that the sickle and granular-shaped RBCs are more likely to be filtered by macrophages in the spleen. This finding is consistent with the observation of low percentages of these two forms of sickle RBCs in the blood smear of SCD patients. Taken together, our experimental and simulation results aid in our quantitative understanding of the function of splenic macrophages in retaining the diseased RBCs and provide an opportunity to combine such knowledge with the current knowledge of the interaction between IES and traversing RBCs to apprehend the complete filtration function of the spleen in SCD.

Machine Learning Classification of Alzheimer's Disease Pathology Reveals Diffuse Amyloid as a Major Predictor of Cognitive Impairment in Human Hippocampal Subregions

Stephen, T.-L. — 2023-06-05

Analyzing Alzheimers disease (AD) pathology within anatomical subregions is a significant challenge, often carried out by pathologists using a standardized, semi-quantitative approach. To augment traditional methods, a high-throughput, high-resolution pipeline was created to classify the distribution of AD pathology within hippocampal subregions. USC ADRC post-mortem tissue sections from 51 patients were stained with 4G8 for amyloid, Gallyas for neurofibrillary tangles (NFTs) and Iba1 for microglia. Machine learning (ML) techniques were utilized to identify and classify amyloid pathology (dense, diffuse and APP (amyloid precursor protein)), NFTs, neuritic plaques and microglia. These classifications were overlaid within manually segmented regions (aligned with the Allen Human Brain Atlas) to create detailed pathology maps. Cases were separated into low, intermediate, or high AD stages. Further data extraction enabled quantification of plaque size and pathology density alongside ApoE genotype, sex, and cognitive status. Our findings revealed that the increase in pathology burden across AD stages was driven mainly by diffuse amyloid. The pre and para-subiculum had the highest levels of diffuse amyloid while NFTs were highest in the A36 region in high AD cases. Moreover, different pathology types had distinct trajectories across disease stages. In a subset of AD cases, microglia were elevated in intermediate and high compared to low AD. Microglia also correlated with amyloid pathology in the Dentate Gyrus. The size of dense plaques, which may represent microglial function, was lower in ApoE4 carriers. In addition, individuals with memory impairment had higher levels of both dense and diffuse amyloid. Taken together, our findings integrating ML classification approaches with anatomical segmentation maps provide new insights on the complexity of disease pathology in AD progression. Specifically, we identified diffuse amyloid pathology as being a major driver of AD in our cohort, regions of interest and microglial responses that might advance AD diagnosis and treatment.

A Transient High-dimensional Geometry Affords Stable Conjunctive Subspaces for Efficient Action Selection

Kikumoto, A. — 2023-06-11

Flexible action selection requires cognitive control mechanisms capable of mapping the same inputs to different output actions depending on the context. From a neural state-space perspective, this requires a control representation that separates similar input neural states by context. Additionally, for action selection to be robust and time-invariant, information must be stable in time, enabling efficient readout. Here, using EEG decoding methods, we investigate how the geometry and dynamics of control representations constrain flexible action selection in the human brain. Participants performed a context-dependent action selection task. A forced response procedure probed action selection different states in neural trajectories. The result shows that before successful responses, there is a transient expansion of representational dimensionality that separated conjunctive subspaces. Further, the dynamics stabilizes in the same time window, with entry into this stable, high-dimensional state predictive of individual trial performance. These results establish the neural geometry and dynamics the human brain needs for flexible control over behavior.

TRAIL agonists rescue mice from radiation-induced lung injury

Strandberg, J. — 2023-06-13

Cancer therapy is often limited by toxicity from pneumonitis. This often-lethal side effect is known to be impacted by innate immunity, and in particular the pathways regulated by the TRAIL death receptor DR5. We investigated whether DR5 agonists could rescue mice from the lethal effects of radiation. We found that two different agonists, parenteral PEGylated trimeric-TRAIL (TLY012) and oral TRAIL-Inducing Compound #10 (TIC10/ONC201), could achieve this goal. Both compounds could completely protect mice from lethality by reducing pneumonitis, alveolar-wall thickness, and oxygen desaturation. At the molecular level, this protection appeared to be due to the inhibition of CCl22, a macrophage-derived chemokine previously associated with radiation pneumonitis and pulmonary fibrosis. The discovery that short-term treatment with TRAIL pathway agonists effectively rescues animals from high doses of radiation exposure has important translational implications. One Sentence SummaryPrevention of lethality, pneumonitis, lung fibrosis and skin dermatitis post-{psi}-irradiation by short- term treatment with innate immune TRAIL pathway agonists

WFIKKN2 is a bifunctional axon guidance cue that signals through divergent DCC family receptors

Nickerson, K. R. — 2023-06-15

Axon pathfinding is controlled by attractive and repulsive molecular cues that activate receptors on the axonal growth cone, but the full repertoire of axon guidance molecules remains unknown. The vertebrate DCC receptor family contains the two closely related members DCC and Neogenin with prominent roles in axon guidance and three additional, divergent members - Punc, Nope, and Protogenin - for which functions in neural circuit formation have remained elusive. We identified a secreted Punc/Nope/Protogenin ligand, WFIKKN2, which guides mouse peripheral sensory axons through Nope-mediated repulsion. In contrast, WFIKKN2 attracts motor axons, but not via Nope. These findings identify WFIKKN2 as a bifunctional axon guidance cue that acts through divergent DCC family members, revealing a remarkable diversity of ligand interactions for this receptor family in nervous system wiring. One-Sentence SummaryWFIKKN2 is a ligand for the DCC family receptors Punc, Nope, and Prtg that repels sensory axons and attracts motor axons.

Building methodological consensus to ensure rigor and reproducibility in zebrafish fertility research

Kossack, M. E. — 2023-06-21

Zebrafish are an increasingly popular model for studying the genetic and environmental factors that shape male and female fertility; however, the field currently lacks a standardized approach to fertility assessment. The current lack of consensus makes comparisons across studies more challenging and is an obstacle to reproducibility in the fields of reproductive biology and toxicology. Here, we review the diversity of spawning approaches used in zebrafish reproductive toxicology research to asses fertility and provide evidence that spawning parameters can result in meaningful differences in egg production and spawning success. HighlightsO_LIZebrafish fertility research lacks methodological consensus C_LIO_LIMeasures of fertility vary with age and frequency of spawning C_LIO_LIMethodological consensus will increase reproducibility in toxicology C_LI

Exposure to the aryl hydrocarbon receptor agonist dioxin disrupts formation of the muscle, nerves, and vasculature in the developing jaw

Cintron-Rivera, L. G. — 2023-06-23

Human exposures to environmental pollutants can disrupt embryonic development and impact juvenile and adult health outcomes by adversely affecting cell and organ function. Notwithstanding, environmental contamination continues to increase because of industrial development, insufficient regulations, and the mobilization of pollutants due to extreme weather events. Dioxins are a class of structurally related persistent organic pollutants that are highly toxic, carcinogenic, and teratogenic. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is the most potent dioxin compound and has been shown to induce toxic effects in developing organisms by activating the aryl hydrocarbon receptor (AHR), a ligand activated transcription factor targeted by multiple persistent organic pollutants. Contaminant-induced AHR activation results in malformations in the craniofacial cartilages and neurocranium; however, the mechanisms mediating these phenotypes are not entirely understood. In this study, we utilized the optically transparent zebrafish model to elucidate novel transcriptional and structural targets of embryonic TCDD exposure leading to craniofacial malformations. To this end, we exposed zebrafish embryos at 4 hours post fertilization (hpf) to TCDD and employed a mixed-methods approach utilizing immunohistochemistry staining, transgenic reporter lines, fixed and in vivo confocal imaging, and timelapse microscopy to determine the targets mediating TCDD-induced craniofacial phenotypes. Our data shows that embryonic TCDD exposure reduced jaw and pharyngeal arch Sox10+ chondrocytes and Tcf21+ pharyngeal mesoderm progenitors. Exposure to TCDD correspondingly led to a reduction in collagen type II deposition in Sox10+ domains. Embryonic TCDD exposure impaired development of tissues derived from or guided by Tcf21+ progenitors, namely: nerves, muscle, and vasculature. Specifically, TCDD exposure disrupted development of the hyoid and mandibular arch muscles, decreased neural innervation of the jaw, resulted in compression of cranial nerves V and VII, and led to jaw vasculature malformations. Collectively, these findings reveal novel transcriptional and structural targets of TCDD-induced toxicity, showcasing how contaminant exposures lead to congenital craniofacial malformations. HighlightsO_LIEmbryonic TCDD exposure diminishes Sox10+ craniofacial chondrocytes. C_LIO_LIFollowing TCDD exposure Col2a1 deposition is reduced in Sox10+ domains. C_LIO_LIExposure to TCDD decreases Tcf21+ progenitors and impairs muscle formation. C_LIO_LITCDD exposure leads to defects jaw innervation and cranial nerve establishment. C_LIO_LIEarly TCDD exposure results in vasculature malformations in the jaw. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=91 SRC="FIGDIR/small/546117v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@b803c4org.highwire.dtl.DTLVardef@ff4483org.highwire.dtl.DTLVardef@eafa18org.highwire.dtl.DTLVardef@c97dba_HPS_FORMAT_FIGEXP M_FIG C_FIG

Collagen type VI regulates TGFβ bioavailability in skeletal muscle

Mohassel, P. — 2023-06-24

Collagen VI-related disorders (COL6-RDs) are a group of rare muscular dystrophies caused by pathogenic variants in collagen VI genes (COL6A1, COL6A2, and COL6A3). Collagen type VI is a heterotrimeric, microfibrillar component of the muscle extracellular matrix (ECM), predominantly secreted by resident fibroadipogenic precursor cells in skeletal muscle. The absence or mislocalizatoion of collagen VI in the ECM underlies the non-cell autonomous dysfunction and dystrophic changes in skeletal muscle with an as of yet elusive direct mechanistic link between the ECM and myofiber dysfunction. Here, we conduct a comprehensive natural history and outcome study in a novel mouse model of COL6-RDs (Col6a2-/-mice) using standardized (Treat-NMD) functional, histological, and physiologic parameter. Notably, we identify a conspicuous dysregulation of the TGF{beta} pathway early in the disease process and propose that the collagen VI deficient matrix is not capable of regulating the dynamic TGF{beta} bioavailability at baseline and also in response to muscle injury. Thus, we propose a new mechanism for pathogenesis of the disease that links the ECM regulation of TGF{beta} with downstream skeletal muscle abnormalities, paving the way for developing and validating therapeutics that target this pathway.

CaBLAM! A high-contrast bioluminescent Ca2+ indicator derived from an engineered Oplophorus gracilirostris luciferase

Lambert, G. G. — 2023-06-26

Measuring ongoing cellular activity is essential to understanding the dynamic functions of biological organisms. The most popular current approach is imaging fluorescence-based genetically encoded Ca2+ indicators (GECIs). While fluorescent probes are useful in many contexts, bioluminescence-based GECIs--probes that generate light through oxidation of a small-molecule by a luciferase or photoprotein--have several distinct advantages. Because bioluminescent (BL) GECIs do not use the bright extrinsic excitation light required for fluorescence, BL GECIs do not photobleach, do not suffer from nonspecific autofluorescent background, and do not cause phototoxicity. Further, BL GECIs can be applied in contexts where directly shining photons on an imaging target is not possible. Despite these advantages, the use of BL GECIs has to date been limited by their small changes in bioluminescence intensity, high baseline signal at resting Ca2+ concentrations, and suboptimal Ca2+ affinities. Here, we describe a new BL GECI, CaBLAM (Ca2+ BioLuminescence Activity Monitor), that displays much higher dynamic range than previous BL GECIs and has a Ca2+ affinity suitable for capturing physiological changes in cytosolic Ca2+ concentration. With these improvements, CaBLAM captures single-cell and subcellular resolution activity at high frame rates in cultured neurons and in vivo, and allows multi-hour recordings in mice and behaving zebrafish. This new advance provides a robust alternative to traditional fluorescent GECIs that can enable or enhance imaging across many experimental conditions.

A FRET-based excitatory luminopsin with improved efficacy of bioluminescence-driven activation

Bjorefeldt, A. — 2023-06-26

SignificanceBioluminescent optogenetics (BL-OG) offers a unique and powerful approach to manipulate neural activity both opto- and chemogenetically using a single actuator molecule (a LuMinOpsin, LMO). AimTo further enhance the utility of BL-OG by improving the efficacy of chemogenetic (bioluminescence- driven) LMO activation. ApproachWe developed novel luciferases optimized for Forster resonance energy transfer (FRET) when fused to the fluorescent protein mNeonGreen, generating bright bioluminescent (BL) emitters spectrally tuned to Volvox Channelrhodopsin 1 (VChR1). ResultsA new LMO generated from this approach (LMO7) showed significantly stronger BL-driven opsin activation compared to previous and other new variants. We extensively benchmarked LMO7 against LMO3 (current standard), and found significantly stronger neuronal activity modulation ex vivo and in vivo, and efficient modulation of behavior. ConclusionsWe report a robust new option for achieving multiple modes of control in a single actuator, and a promising engineering strategy for continued improvement of BL-OG.

A Bioluminescent Activity Dependent (BLADe) Platform for Converting Neuronal Activity to Photoreceptor Activation

Crespo, E. L. — 2023-06-27

Genetically encoded sensors and actuators have advanced the ability to observe and manipulate cellular activity, yet few non-invasive strategies enable cells to directly couple their intracellular states to user-defined outputs. We developed a bioluminescent activity-dependent (BLADe) platform that facilitates programmable feedback through genetically encoded light generation. Using calcium (Ca{superscript 2}) flux as a model, we engineered a Ca{superscript 2}-dependent luciferase that functions as both a reporter and an activity-gated light source capable of photoactivating light-sensing actuators. In neurons, the presence of luciferin triggers Ca{superscript 2} dependent local illumination that provides activity dependent gene expression by activating a light-sensitive transcription factor and control of neural dynamics through opsin activation in single cells, populations and intact tissue. BLADe can be expanded to couple any signal that bioluminescent enzymes can be engineered to detect with the wide variety of photosensing actuators. This modular strategy of coupling an activity dependent light emitter to a light sensing actuator offers a generalizable framework for state dependent cell-autonomous control across biological systems.

Evaluating Study Design Rigor in Preclinical Cardiovascular Research: A Replication Study

Jimenez, I. C. — 2023-06-29

BackgroundMethodological rigor remains a priority in preclinical cardiovascular research to ensure experimental reproducibility and high-quality research. Limited reproducibility diminishes the translation of preclinical discoveries into medical practice. In addition, lack of reproducibility fosters uncertainty in the publics acceptance of reported research results. MethodsWe evaluated the reporting of methodological practices in preclinical cardiovascular research studies published in leading scientific journals by screening articles for the inclusion of the following study design elements (SDEs): considering sex as a biological variable, randomization, blinding, and sample size power estimation. We screened for these SDEs across articles regarding preclinical cardiovascular research studies published between 2011 and 2021. We replicated and extended a study published in 2017 by Ramirez et al. We hypothesized a higher SDE inclusion across preclinical studies over time, that preclinical studies that include human and animal substudies within the same study will exhibit greater SDE inclusion than animal-only preclinical studies, and that a difference exists in SDE usage between large and small animal models. ResultsSDE inclusion was low; with 15.2% of animal-only studies including both sexes as a biological variable, 30.4% including randomization, 32.1% including blinding, and 8.2% including sample size estimation. The incorporation of SDEs did not significantly increase over the ten-year timeframe in the screened articles. Randomization and sample size estimation differed significantly between animal and human substudies (corrected p=1.85e-05 and corrected p=3.81e-07, respectively.) ConclusionsEvidence of methodological rigor varies depending on the study type and model organisms used. From 2011-2021, SDE reporting within preclinical studies has not increased, suggesting more work is needed to foster the inclusion of rigorous study design elements in cardiovascular research.

Stochastic coding: a conserved feature of odor representations and its implications for odor discrimination

Srinivasan, S. — 2023-06-29

Sparse coding is thought to improve discrimination of sensory stimuli by reducing overlap between their representations. Two factors, however, can offset sparse codings advantages. Similar sensory stimuli have significant overlap, and responses vary across trials. To elucidate the effect of these two factors, we analyzed odor responses in the fly and mouse olfactory regions implicated in learning and discrimination -- the Mushroom Body (MB) and the Piriform Cortex (PCx). In both species, we show that neuronal responses fall along a continuum from extremely reliable across trials to extremely variable or stochastic. Computationally, we show that the range of observed variability arises from probabilistic synapses in inhibitory feedback connections within central circuits rather than sensory noise, as is traditionally assumed. We propose this coding scheme to be advantageous for coarse- and fine-odor discrimination. More reliable cells enable quick discrimination between dissimilar odors. For similar odors, however, these cells overlap, and do not provide distinguishing information. By contrast, more unreliable cells are decorrelated for similar odors, providing distinguishing information, though this requires extended training with more trials. Overall, we have uncovered a stochastic coding scheme that is conserved in vertebrates and invertebrates, and we identify a candidate mechanism, based on variability in a winner-take-all inhibitory circuit, that improves discrimination with training.

A CRISPR-based strategy for targeted sequencing in biodiversity science

Littleford-Colquhoun, B. L. — 2023-06-30

Many applications in molecular ecology require the ability to match specific DNA sequences from single- or mixed-species samples to a diagnostic reference library. Widely used methods for DNA barcoding and metabarcoding require PCR and amplicon sequencing to identify taxa based on target sequences, but the target-specific enrichment capabilities of CRISPR-Cas systems may offer advantages in some applications. We identified 54,837 CRISPR-Cas guide RNAs that may be useful for enriching chloroplast DNA across phylogenetically diverse plant species. We then tested a subset of 17 guide RNAs in vitro to enrich and sequence plant DNA strands ranging in size from diagnostic DNA barcodes of 1,428 bp to entire chloroplast genomes of 121,284 bp. We used an Oxford Nanopore sequencer to evaluate sequencing success based on both single- and mixed-species samples, which yielded mean on-target chloroplast sequence lengths of 5,755-11,367 bp, depending on the experiment. Single-species experiments yielded more on-target sequence reads and greater accuracy, but mixed-species experiments yielded superior coverage. Comparing CRISPR-based strategies to a widely used protocol for plant DNA metabarcoding with the chloroplast trnL-P6 marker, we obtained a 66-fold increase in sequence length and markedly better estimates of relative abundance for a commercially prepared mixture of plant species. Future work would benefit from developing both in vitro and in silico methods for analyses of mixed-species samples, especially when the appropriate reference genomes for contig assembly cannot be known a priori. Prior work developed CRISPR-based enrichment protocols for long-read sequencing and our experiments pioneered its use for plant DNA barcoding and chromosome assemblies that may have advantages over workflows that require PCR and short-read sequencing.

Increasing chemotherapeutic efficacy using doxorubicin and sodium bicarbonate releasing injectable chitosan-polyethylene glycol hydrogels

Ahmed, Z. — 2023-07-08

Modulation of pH is crucial to maintaining the chemical homeostasis of biological environments. The irregular metabolic pathways exhibited by cancer cells result in the production of acidic byproducts that are excreted and accumulate in the extracellular tumor microenvironment, reducing its pH. As a consequence of the lower pH in tumors, cancer cells increase the expression of metastatic phenotypes and chemotherapeutic resistance. A significant limitation in current cancer therapies is the inability to locally deliver the chemotherapy, leading to significant damage to healthy cells in systemic administration. To overcome these challenges, we present an injectable chitosan-polyethylene glycol hydrogel that is dual-loaded with doxorubicin and sodium bicarbonate providing alkaline buffering of extracellular acidity and simultaneous chemotherapeutic delivery to increase chemotherapeutic efficacy. We conducted in vitro studies of weak base chemotherapeutic and alkaline buffer release from the hydrogel. The release of doxorubicin from hydrogels increased in a low pH environment and was dependent on the encapsulated sodium bicarbonate concentration. We investigated the influence of pH on doxorubicin efficacy and viability of MCF-7 and MDA-MB-231 breast cancer cell lines. The results show a 2 to 3 fold increase in IC50 values from neutral pH to low pH, showing decreased cancer cell viability at neutral pH as compared to acidic pH. The IC50 results were shown to correlate with a decrease in intracellular uptake of doxorubicin at low pH. The proposed hydrogels were confirmed to be non-toxic to healthy MCF-10A mammary epithelial cells. Rheological studies were performed to verify that the dual loaded hydrogels were injectable. The mechanical and release properties of the hydrogels were maintained after extended storage. The chemotherapeutic activity of doxorubicin was evaluated in the presence of the proposed pH regulating hydrogels. The findings suggest a promising non-toxic, biodegradable hydrogel buffer delivery system that can achieve two simultaneous important goals of local acidosis neutralization and chemotherapeutic release.

Glutamatergic neuronal activity regulates angiogenesis and blood-retinal barrier maturation via Norrin/β-catenin signaling

Biswas, S. — 2023-07-11

Interactions among neuronal, glial and vascular components are crucial for retinal angiogenesis and blood-retinal barrier (BRB) maturation. Although synaptic dysfunction precedes vascular abnormalities in many retinal pathologies, how neuronal activity, specifically glutamatergic activity, regulates retinal angiogenesis and BRB maturation remains unclear. Using in vivo genetic studies in mice, single-cell RNA-sequencing and functional validation, we show that deep plexus angiogenesis and paracellular BRB maturation are delayed in Vglut1-/- retinas where neurons fail to release glutamate. In contrast, deep plexus angiogenesis and paracellular BRB maturation are accelerated in Gnat1-/- retinas where constitutively depolarized rods release excessive glutamate. Norrin expression and endothelial Norrin/{beta}-catenin signaling are downregulated in Vglut1-/- retinas, and upregulated in Gnat1-/- retinas. Pharmacological activation of endothelial Norrin/{beta}-catenin signaling in Vglut1-/- retinas rescued defects in deep plexus angiogenesis and paracellular BRB maturation. Our findings demonstrate that glutamatergic neuronal activity regulates retinal angiogenesis and BRB maturation by modulating endothelial Norrin/{beta}-catenin signaling.

Beyond the (geometric) mean: stochastic models undermine deterministic predictions of bet hedger evolution

Weissman, M. R. — 2023-07-12

Bet hedging is a ubiquitous strategy for risk reduction in the face of unpredictable environmental change where a lineage lowers its variance in fitness across environments at the expense of also lowering its arithmetic mean fitness. Classically, the benefit of bet hedging has been quantified using geometric mean fitness (GMF); bet hedging is expected to evolve if and only if it has a higher GMF than the wild-type. We build upon previous research on the effect of incorporating stochasticity in phenotypic distribution, environment, and reproduction to investigate the extent to which these sources of stochasticity will impact the evolution of real-world bet hedging traits. We utilize both individual-based simulations and Markov chain numerics to demonstrate that modeling stochasticity can alter the sign of selection for the bet hedger compared to deterministic predictions. We find that bet hedging can be deleterious at small population sizes and beneficial at larger population sizes. This non-monotonic dependence of the sign of selection on population size, known as sign inversion, exists across parameter space for both conservative and diversified bet hedgers. We apply our model to published data of bet hedging strategies to show that sign inversion exists for biologically relevant parameters in two study systems: Papaver dubium, an annual poppy with variable germination phenology, and Salmonella typhimurium, a pathogenic bacteria that exhibits antibiotic persistence. Taken together, our results suggest that GMF is not enough to predict when bet hedging is adaptive.

Identifying Protein Conformational States in the PDB and Comparison to AlphaFold2 Predictions

Ellaway, J. I. J. — 2023-07-13

Proteins, as molecular machines, are necessarily dynamic macromolecules that carry out essential cellular functions. Recognising their stable conformations is important for understanding the molecular mechanisms of disease. While AI-based computational methods have enabled protein structure prediction, the prediction of protein dynamics remains a challenge. Here, we present a deterministic pipeline that clusters experimentally determined protein structures to comprehensively recognise conformational states across the Protein Data Bank. Our approach clusters protein chains based on a GLObal CONformation (GLOCON) difference score, which is computed from pairwise C-alpha distances. By superposing the clustered structures, differences and similarities in conformational states can be observed. Additionally, we offer users the ability to superpose predicted models from the AlphaFold Database to the clusters of PDB structures. This clustering pipeline significantly advances researchers ability to explore the conformational landscape within the PDB. All clustered and superposed models can be viewed in Mol* on the PDBe Knowledge Base website, or accessed in as raw annotations via our GraphAPI and FTP server. The clustering package is made available as an open-source Python3 package under the Apache-2.0 license.

Validation of human telomere length trans-ancestry meta-analysis association signals identifies POP5 and KBTBD6 as novel human telomere length regulation genes

Keener, R. — 2023-07-14

Telomere length genome-wide association studies (GWAS) have become well-powered to detect novel genes in telomere length regulation. However, no prior work has validated these putative novel genes to confirm the contribution of GWAS loci to telomere length regulation. We conducted a trans-ancestry meta-analysis of 211,369 individuals. Through enrichment analyses of chromatin state and cell-type heritability we identified blood and immune cells as the most relevant cell type to examine telomere length association signals. We validated specific GWAS associations by overexpressing KBTBD6, a component of an E3 ubiquitin ligase complex, and POP5, a component of the Ribonuclease P/MRP complex, and demonstrating that both lengthened telomeres as predicted by our statistical analyses. CRISPR/Cas9 deletion of the predicted causal regions of these association peaks in K562 immortalized blood cells reduced expression of these genes, demonstrating that these loci are related to transcriptional regulation of KBTBD6 and POP5, respectively. Together our results demonstrate the utility of telomere length GWAS in the identification of novel telomere length regulation mechanisms and highlight the importance of the proteasome-ubiquitin pathway in telomere length regulation.

ChatGPT-Enhanced ROC Analysis (CERA): A Shiny Web Tool for Finding Optimal Cutoff in Biomarker Analysis

Agraz, M. — 2023-07-15

Diagnostic tests play a crucial role in establishing the presence of a specific disease in an individual. Receiver Operating Characteristic (ROC) curve analyses are essential tools that provide performance metrics for diagnostic tests. Accurate determination of the cutoff point in ROC curve analyses is the most critical aspect of the process. A variety of methods have been developed to find the optimal cutoffs. Although the R programming language provides a variety of package programs for conducting ROC curve analysis and determining the appropriate cutoffs, it typically needs coding skills and a substantial investment of time. Specifically, the necessity for data preprocessing and analysis can present a significant challenge, especially for individuals without coding experience. We have developed the CERA (ChatGPT-Enhanced ROC Analysis) tool, a user-friendly ROC curve analysis web tool using the shiny interface for faster and more effective analyses to solve this problem. CERA is not only user-friendly, but it also interacts with ChatGPT, which interprets the outputs. This allows for an interpreted report generated by R-Markdown to be presented to the user, enhancing the accessibility and understanding of the analysis results. Authors summaryMelih Agraz, after graduating from Dokuz Eylul University in Izmir, Turkiye, with a major in Mathematics from the Department of Education, he pursued his Masters degree at the same university in the field of Statistics. He furthered his education by obtaining a Ph.D. in Statistics from the Middle East Technical University, Ankara in Turkiye. As part of his academic journey, he also served as a Fulbright postdoctoral researcher at UC Berkeley in the United States. Following this, he worked as a postdoctoral researcher at Brown University and Beth Israel Hospital of Harvard Medical School. Currently, he holds the position of Assistant Professor in the Department of Statistics at Giresun University in Turkey. George Em Karniadakis is Professor of Applied Mathematics and Engineering at Brown University. He is a member of the National Academy of Engineering of USA. His interests include stochastic multiscale modeling of physical and biological systems, physics-informed machine learning, and deep neural operators. He has co-authored over 500 papers and five books, and he has the highest h-index in Applied Mathematics according to Google Scholar.

Pain reflects the informational value of nociceptive inputs.

Coll, M.-P. — 2023-07-18

Pain perception and its modulation are fundamental to human learning and adaptive behavior. This study investigated the hypothesis that pain perception is tied to pains learning function. Thirty-one participants performed a threat conditioning task where certain cues were associated with a possibility of receiving a painful electric shock. The cues that signalled potential pain or safety were regularly changed, requiring participants to continually establish new associations. Using computational models, we quantified participants pain expectations and prediction errors throughout the task and assessed their relationship with pain perception and electrophysiological responses. Our findings suggest that subjective pain perception increases with prediction error, that is when pain was unexpected. Prediction errors were also related to physiological nociceptive responses, including the amplitude of the nociceptive flexion reflex and EEG markers of cortical nociceptive processing (N2-P2 evoked potential and gamma-band power). Additionally, higher pain expectations were related to increased late event-related potential responses and alpha/beta decreases in amplitude during cue presentation. These results further strengthen the idea of a crucial link between pain and learning and suggest that understanding the influence of learning mechanisms in pain modulation could help us understand when and why pain perception is modulated in health and disease.

Enterobacter Sp. SM3 Exhibits Run-and-Tumble Motility

Johnson, S. G. — 2023-07-24

The recent discovery of the peritrichous, swarm-competent bacterium Enterobacter sp. SM3 has offered a new opportunity to elucidate the role of swarming motility in the gut microbiome. Here we present first findings of the run-and-tumble behavior of SM3 in both a control solution of motility buffer and upon bulk exposure to the chemoattractants serine and aspartate, drawing a comparison with the well-studied behavior of E. coli. It was found that SM3 runs with an average speed of approximately 30 {micro}m/s for an average duration of 0.77 s. Tumble events occur for an average duration of 0.11 s with a 69{whitebullet} average tumble angle. Both values are similar to that of E. coli. When exposed to serine, SM3 suppresses the frequency of tumble events, which in turn increases the average run duration. In addition, the average tumble angle was found to decrease in response to serine. However, when exposed to up to a millimolar concentration of aspartate, SM3 does not demonstrate a notable change in run-and-tumble parameters. These results suggest that run-and-tumble is the characteristic swimming behavior of SM3 in its planktonic state. These findings serve as a benchmark in a quest to determine the connection among swimming, swarming, and the complex dynamics of the gut microbiome. IMPORTANCEBacteria form the largest domain of living creatures on this planet. Our interactions with bacteria influence us in many ways, not the least of which being in regards to human health. A recently identified species of gut bacteria, Enterobacter sp. SM3, has been shown to reduce intestinal inflammation, suggesting that swarming could play a physiologically beneficial role. In this report, we study the motility of individual SM3 bacteria This study is an essential step towards an overarching goal to understand the influence of bacterial motility on human health.

Predicting Relative Populations of ProteinConformations without a Physics Engine UsingAlphaFold2

Monteiro da Silva, G. — 2023-07-27

This paper presents a novel approach for predicting the relative populations of protein conformations using AlphaFold 2, an AI-powered method that has revolutionized biology by enabling the accurate prediction of protein structures. While AlphaFold 2 has shown exceptional accuracy and speed, it is designed to predict proteins' single ground state conformations and is limited in its ability to predict fold switching and the effects of mutations on conformational landscapes. Here, we demonstrate how AlphaFold 2 can directly predict the relative populations of different conformations of proteins and even accurately predict changes in those populations induced by mutations by subsampling multiple sequence alignments. We tested our method against NMR experiments on two proteins with drastically different amounts of available sequence data, Abl1 kinase and the granulocyte-macrophage colony-stimulating factor, and predicted their relative state populations with accuracies in excess of 80%. Our method offers a fast and cost-effective way to predict protein conformations and their relative populations at even single point mutation resolution, making it a useful tool for pharmacology, analyzing NMR data, and studying the effects of evolution.

Genomic analysis supports Cape Lion population connectivity prior to colonial eradication and extinction

de Flamingh, A. — 2023-08-02

Extinct Cape lions (Panthera leo melanochaitus) formerly ranged throughout the grassland plains of the "Cape Flats" in what is today known as the Western Cape Province, South Africa. Cape lions were likely eradicated because of overhunting and habitat loss after European colonization. European naturalists originally described Cape lions as "Black-maned lions" and claimed that they were phenotypically distinct. However, other depictions and historical descriptions of lions from the Cape report mixed or light coloration and without black or extensively developed manes. These findings suggest that, rather than forming a distinct population, Cape lions may have had phenotypic and genotypic variation similar to other African lions. Here we investigate extinct Cape lion genome characteristics, population dynamics and genetic distinctiveness prior to their extinction. We generated genomic data from two historic Cape lions to compare to 118 existing high-coverage mitogenomes, and low-coverage nuclear genomes of 53 lions from 13 African countries. We show that, before their eradication, lions from the Cape Flats had diverse mitogenomes and nuclear genomes that clustered with lions from both southern and eastern Africa. Cape lions had high genome-wide heterozygosity and low inbreeding coefficients, indicating that populations in the Cape Flats went extinct so rapidly that genomic effects associated with long-term small population size and isolation were not detectable. Our findings do not support the characterization of Cape lions as phylogeographically distinct, as originally put forth by some European naturalists, but rather highlights how alternative knowledge-systems, e.g., Indigenous perspectives, could potentially further inform interpretations of species life histories.

Bloodmeal metabarcoding of the argasid tick (Ornithodoros turicata Duges) reveals extensive vector-host associations

Balasubramanian, S. — 2023-08-07

Molecular methods to understand host feeding patterns of arthropod vectors are critical to assess exposure risk to vector-borne disease and unveil complex ecological interactions. We build on our prior work discovering the utility of PCR-Sanger sequencing bloodmeal analysis that work remarkably well for soft ticks (Acari: Argasidae), unlike for hard ticks (Acari: Ixodidae), thanks to their unique physiology that retains vertebrate DNA from prior bloodmeals viable for years. Here, we capitalize on this feature and apply bloodmeal metabarcoding using amplicon deep sequencing to identify multiple host species in individual Ornithodoros turicata soft ticks collected from two natural areas in Texas, United States. Of 788 collected O. turicata, 394 were evaluated for bloodmeal source via metabarcoding, revealing 27 different vertebrate host species (17 mammals, 5 birds, 1 reptile, and 4 amphibians) fed upon by 274 soft ticks. Information on multiple hosts for individual O. turicata was derived from 168 of these (61%). Metabarcoding revealed more mixed vertebrate bloodmeals in O. turicata previously processed using Sanger sequencing. These data reveal wide host range of O. turicata and demonstrate the value of bloodmeal metabarcoding for understanding the ecology for known and potential tick-borne pathogens circulating among humans, domestic animals and wildlife such as relapsing fever caused by Borrelia turicatae. Our results also document, for the first time an off-host soft tick collected to have evidence of prior feeding on wild pig which is a critical observation in the context of the threat of enzootic transmission of African swine fever virus if it were introduced to the US. This research enhances our understanding of vector-host associations and offers a promising perspective for biodiversity monitoring and disease control strategies.

Structure-based development of new cyclic compoundstargeting PSD-95 PDZ3 domain.

Naik, M. T. — 2023-08-11

Aberrant BDNF signaling has been proposed to contribute to the pathophysiology of depression and other neurological disorders such as Angelman syndrome. We have previously shown that targeting the TrkB / PSD-95 nexus by peptidomimetic inhibitors is a promising approach for therapeutic intervention. Here we used structure-based knowledge to develop a new peptidomimetic compound series that fuses SynGAP-derived peptides to our prototype compound CN2097. These compounds target the PSD-95 PDZ3 domain and adjoining C helix to achieve bivalent binding that results in up to 7-fold stronger affinity compared to CN2097. These compounds were designed to improve CN2097 specificity for the PDZ3 domain and limited SAR studies have been performed to improve their resistance to proteolysis.

BRAND: A platform for closed-loop experiments with deep network models

Ali, Y. H. — 2023-08-12

Artificial neural networks (ANNs) are state-of-the-art tools for modeling and decoding neural activity, but deploying them in closed-loop experiments with tight timing constraints is challenging due to their limited support in existing real-time frameworks. Researchers need a platform that fully supports high-level languages for running ANNs (e.g., Python and Julia) while maintaining support for languages that are critical for low-latency data acquisition and processing (e.g., C and C++). To address these needs, we introduce the Backend for Realtime Asynchronous Neural Decoding (BRAND). BRAND comprises Linux processes, termed nodes, which communicate with each other in a graph via streams of data. Its asynchronous design allows for acquisition, control, and analysis to be executed in parallel on streams of data that may operate at different timescales. BRAND uses Redis to send data between nodes, which enables fast inter-process communication and supports 54 different programming languages. Thus, developers can easily deploy existing ANN models in BRAND with minimal implementation changes. In our tests, BRAND achieved <600 microsecond latency between processes when sending large quantities of data (1024 channels of 30 kHz neural data in 1-millisecond chunks). BRAND runs a brain-computer interface with a recurrent neural network (RNN) decoder with less than 8 milliseconds of latency from neural data input to decoder prediction. In a real-world demonstration of the system, participant T11 in the BrainGate2 clinical trial performed a standard cursor control task, in which 30 kHz signal processing, RNN decoding, task control, and graphics were all executed in BRAND. This system also supports real-time inference with complex latent variable models like Latent Factor Analysis via Dynamical Systems. By providing a framework that is fast, modular, and language-agnostic, BRAND lowers the barriers to integrating the latest tools in neuroscience and machine learning into closed-loop experiments.

Mammalian olfactory cortex neurons retain molecular signatures of ancestral cell types

Zeppilli, S. — 2023-08-14

The cerebral cortex diversified extensively during vertebrate evolution. Intriguingly, the three-layered mammalian olfactory cortex resembles the cortical cytoarchitecture of non-mammals yet evolved alongside the six-layered neocortex, enabling unique comparisons for investigating cortical neuron diversification. We performed single-nucleus multiome sequencing across mouse three- to six-layered cortices and compared neuron types across mice, reptiles and salamander. We identified neurons that are olfactory cortex-specific or conserved across mouse cortical areas. However, transcriptomically similar neurons exhibited area-specific epigenetic states. Additionally, the olfactory cortex showed transcriptomic divergence between lab and wild-derived mice, suggesting enhanced circuit plasticity through adult immature neurons. Finally, olfactory cortex neurons displayed marked transcriptomic similarities to reptile and salamander neurons. Together, these data indicate that the mammalian olfactory cortex retains molecular signatures representative of ancestral cortical traits.

Second Harmonic Generation Imaging Reveals Entanglement of Collagen Fibers in the Elephant Trunk Skin Dermis

Schulz, A. — 2023-08-15

Form-function relationships often have tradeoffs: if a material is tough, it is often inflexible, and vice versa. This is particularly relevant for the elephant trunk, where the skin should be protective yet elastic. To investigate how this is achieved, we used classical histochemical staining and second harmonic generation microscopy to describe the morphology and composition of elephant trunk skin. We report structure at the macro and micro scales, from the thickness of the dermis to the interaction of 10 m thick collagen fibers. We analyzed several sites along the length of the trunk, to compare and contrast the dorsal-ventral and proximal-distal skin morphologies and compositions. We find the dorsal skin of the elephant trunk can have keratin armor layers over 2mm thick, which is nearly 100 times the thickness of the equivalent layer in human skin. We also found that the structural support layer (the dermis) of elephant trunk contains a distribution of collagen-I (COL1) fibers in both perpendicular and parallel arrangement. The bimodal distribution of collagen is seen across all portions of the trunk, and is dissimilar from that of human skin where one orientation dominates within a body site. We hypothesize that this distribution of COL1 in the elephant trunk allows both flexibility and load-bearing capabilities. Additionally, when viewing individual fiber interaction of 10 m thick collagen, we find the fiber crossings per unit volume are five times more common than in human skin, suggesting that the fibers are entangled. We surmise that these intriguing structures permit both flexibility and strength in the elephant trunk. The complex nature of the elephant skin may inspire the design of materials that can combine strength and flexibility.

Primary Human Cell-Derived Extracellular Matrix from Decellularized Fibroblast Microtissues with Tissue-Dependent Composition and Microstructure

Fonseca, V. C. — 2023-08-17

Human extracellular matrix (ECM) exhibits complex protein composition and architecture depending on tissue and disease state, which remains challenging to reverse engineer. One promising approach is based on cell-secreted ECM from human fibroblasts, which can then be decellularized into an acellular biomaterial. However, fibroblasts initially seeded on rigid tissue culture plastic or biomaterial scaffolds experience aberrant mechanical cues that influence ECM deposition. Here, we show that engineered microtissues of primary human fibroblasts seeded in low-adhesion microwells can be decellularized to produce human, tissue-specific ECM. We investigate: 1) cardiac fibroblasts, as well as 2) lung fibroblasts from healthy, idiopathic fibrosis and chronic obstructive pulmonary disease donors. We demonstrate optimized culture and decellularization conditions, then characterize gene expression and protein composition. We further characterize ECM microstructure and mechanical properties. We envision that this method could be utilized for biomanufacturing of patient and tissue-specific ECM for organoid drug screening as well as implantable scaffolds. ImpactIn this study, we demonstrate a method for preparing decellularized matrix using primary human fibroblasts with tissue and disease-specific features. We aggregate single cell dispersions into engineered tissues using low adhesion microwells and show culture conditions that promote ECM deposition. We demonstrate this approach for cardiac fibroblasts as well as lung fibroblasts (both normal and diseased). We systematically investigate tissue morphology, matrix architecture, and mechanical properties, along with transcriptomic and proteomic analysis. This approach should be widely applicable for generating personalized ECM with features of patient tissues and disease state, relevant for culturing patient cells ex vivo as well as implantation for therapeutic treatments.

Control adjustment costs limit goal flexibility: Empirical evidence and a theoretical account

Grahek, I. — 2023-08-23

A cornerstone of human intelligence is the ability to flexibly adjust our cognition and behavior as our goals change. For instance, achieving some goals requires efficiency, while others require caution. Different goals require us to engage different control processes, such as adjusting how attentive and cautious we are. Here, we show that performance incurs control adjustment costs when people adjust control to meet changing goals. Across four experiments, we provide evidence of these costs, and validate a dynamical systems model explaining the source of these costs. Participants performed a single cognitively demanding task under varying performance goals (e.g., being fast or accurate). We modeled control allocation to include a dynamic process of adjusting from ones current control state to a target state for a given performance goal. By incorporating inertia into this adjustment process, our model accounts for our empirical finding that people under-shoot their target control state more (i.e., exhibit larger adjustment costs) when goals switch rather than remain fixed (Study 1). Further validating our model, we show that the magnitude of this cost is increased when: distances between target states are larger (Study 2), there is less time to adjust to the new goal (Study 3), and goal switches are more frequent (Study 4). Our findings characterize the costs of adjusting control to meet changing goals, and show that these costs emerge directly from cognitive control dynamics. In so doing, they shed new light on the sources of and constraints on flexibility of goal-directed behavior.

High-Fidelity, Hyper-Accurate, and Evolved Mutants Rewire Atomic Level Communication in CRISPR-Cas9

Skeens, E. — 2023-08-26

The Cas9-HF1, HypaCas9, and evoCas9 variants of the Cas9 endonuclease are critical tools to mitigate off-target effects in the application of CRISPR-Cas9 technology. The mechanisms by which mutations in the Rec3 domain mediate specificity in these variants are poorly understood. Here, solution NMR and molecular dynamics simulations establish the structural and dynamic effects of high-specificity mutations in Rec3, and how they propagate the allosteric signal of Cas9. We reveal conserved structural changes and peculiar dynamic differences at regions of Rec3 that interface with the RNA:DNA hybrid, transducing chemical signals from Rec3 to the catalytic HNH domain. The variants remodel the communication sourcing from the Rec3 -helix 37, previously shown to sense target DNA complementarity, either directly or allosterically. This mechanism increases communication between the DNA mismatch recognition helix and the HNH active site, shedding light on the structure and dynamics underlying Cas9 specificity and providing insight for future engineering principles.

ω-Phonetoxins inhibit voltage-gated calcium CaV2.2 ion channel splice isoforms of dorsal root ganglia

Castro-Junior, C. — 2023-08-31

Cell-specific alternative splicing of Cacna1b pre-mRNA generates functionally distinct voltage-gated CaV2.2 channels. CaV2.2 channels mediate the release of glutamate from nociceptor termini in the dorsal horn spinal cord and they are implicated in chronic pain. One alternatively spliced exon in Cacna1b, e37a, is highly expressed in dorsal root ganglia, relative to other regions of the nervous system, and it is particularly important in inflammatory hyperalgesia. Here we studied the effects of two {omega}-phonetoxins, PnTx3-4 and Ph1{beta}, derived from the spider Phoneutria nigriventer on CaV2.2 channel isoforms of dorsal root ganglia (CaV2.2 e37a and CaV2.2 e37b). Both PnTx3-4 and Ph1{beta} are known to have analgesic effects in rodent models of pain and to inhibit CaV2.2 channels. CaV2.2 e37a and CaV2.2 e37b isoforms expressed in a mammalian cell line were inhibited by PnTx3-4 and Ph1{beta} with similar potency and with similar timecourse, although CaV2.2 e37a currents were slightly, but consistently more sensitive to toxin inhibition compared to CaV2.2 e37b. The inhibitory effects of PnTx3-4 and Ph1{beta} on CaV2.2-e37a and CaV2.2-e37b channels were voltage-dependent, and both occlude the inhibitory effects of {omega}-conotoxin GVIA, consistent with a common site of action. The potency of PnTx3-4 and Ph1{beta} on both major splice isoforms in dorsal root ganglia constribute to understanding the analgesic actions of these {omega}-phonetoxins.

All-optical reporting of chloride ion driving force in the nervous system.

Selfe, J. S. — 2023-09-01

Ionic driving forces provide the net electromotive force for ion movement across receptors, channels, and transporters, and are a fundamental property of all cells. In the brain for example, fast synaptic inhibition is mediated by chloride permeable GABAA receptors, and single-cell intracellular recordings have been the only method for estimating driving forces across these receptors (DFGABAA). Here we present a new tool for quantifying inhibitory receptor driving force named ORCHID: all-Optical Reporting of CHloride Ion Driving force. We demonstrate ORCHIDs ability to provide accurate, high-throughput measurements of resting and dynamic DFGABAA from genetically targeted cell types over multiple timescales. ORCHID confirms theoretical predictions about the biophysical mechanisms that establish DFGABAA, reveals novel differences in DFGABAA between neurons and astrocytes, and affords the first in vivo measurements of intact DFGABAA. This work extends our understanding of inhibitory synaptic transmission and establishes a precedent for all-optical methods to assess ionic driving forces.

Defining RNA oligonucleotides that reverse deleterious phase transitions of RNA-binding proteins with prion-like domains

Guo, L. — 2023-09-04

RNA-binding proteins with prion-like domains, such as FUS and TDP-43, condense into functional liquids, which can transform into pathological fibrils that underpin fatal neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS)/frontotemporal dementia (FTD). Here, we define short RNAs (24-48 nucleotides) that prevent FUS fibrillization by promoting liquid phases, and distinct short RNAs that prevent and, remarkably, reverse FUS condensation and fibrillization. These activities require interactions with multiple RNA-binding domains of FUS and are encoded by RNA sequence, length, and structure. Importantly, we define a short RNA that dissolves aberrant cytoplasmic FUS condensates, restores nuclear FUS, and mitigates FUS proteotoxicity in optogenetic models and human motor neurons. Another short RNA dissolves aberrant cytoplasmic TDP-43 condensates, restores nuclear TDP-43, and mitigates TDP-43 proteotoxicity. Since short RNAs can be effectively delivered to the human brain, these oligonucleotides could have therapeutic utility for ALS/FTD and related disorders.

miR-6883 downregulates HIF1α in colorectal and breast cancer cells

Jensen-Velez, N. — 2023-09-06

Colorectal cancer (CRC) and breast cancer (BC) are deadly diseases that rank as the second and fourth leading causes of cancer-related deaths, respectively. We have previously shown that miR-6883 targets CDK4/6 and that palbociclib-mediated CDK4/6 inhibition destabilizes HIF1. We hypothesize that miR-6883 downregulates HIF1 in CRC and BC cells. miR-6883 was transfected under normoxia or hypoxia and western blot analysis revealed that miR-6883 downregulates CDK4/6 and HIF1 in CRC and BC cells, pointing to miR-6883 as a promising therapeutic to target hypoxic cancers or HIF1-deregulated tumor cells. Future studies will further investigate miR-6883 as a cancer biomarker, effects on HIF-related proteins and therapeutic uses in vivo. O_FIG O_LINKSMALLFIG WIDTH=164 HEIGHT=200 SRC="FIGDIR/small/556385v1_fig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@e0acadorg.highwire.dtl.DTLVardef@68b428org.highwire.dtl.DTLVardef@bc4dforg.highwire.dtl.DTLVardef@aa13ad_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO miR-6883 downregulates CDK4/6 and HIF1 in colorectal and breast cancer cells A) Colorectal cancer cells were treated with doses of palbociclib ranging from 0-20 {micro}M. Cell viability was measured by imaging the bioluminescent signal after addition of CellTiter-Glo reagent. B) Percent viability of colorectal cancer cells treated with palbociclib was calculated and nonlinear regression analysis was completed using GraphPad Prism software. C-H) CRC cells were transfected with miR-6883 under normoxia or hypoxia (<0.5% O2) and protein levels of CDK4/6, HIF1, Glut1, and Ran were measured by Western blot. C_FIG

Examining DNA Breathing with pyDNA-EPBD

Kabir, A. — 2023-09-12

MotivationThe two strands of the DNA double helix locally and spontaneously separate and recombine in living cells due to the inherent thermal DNA motion.This dynamics results in transient openings in the double helix and is referred to as "DNA breathing" or "DNA bubbles." The propensity to form local transient openings is important in a wide range of biological processes, such as transcription, replication, and transcription factors binding. However, the modeling and computer simulation of these phenomena, have remained a challenge due to the complex interplay of numerous factors, such as, temperature, salt content, DNA sequence, hydrogen bonding, base stacking, and others. ResultsWe present pyDNA-EPBD, a parallel software implementation of the Extended Peyrard-Bishop-Dauxois (EPBD) nonlinear DNA model that allows us to describe some features of DNA dynamics in detail. The pyDNA-EPBD generates genomic scale profiles of average base-pair openings, base flipping probability,DNA bubble probability, and calculations of the characteristically dynamic length indicating the number of base pairs statistically significantly affected by a single point mutation using the Markov Chain Monte Carlo (MCMC) algorithm.

Protein generation with evolutionary diffusion: sequence is all you need

Alamdari, S. — 2023-09-12

Deep generative models are increasingly powerful tools for the in silico design of novel proteins. Recently, a family of generative models called diffusion models has demonstrated the ability to generate biologically plausible proteins that are dissimilar to any actual proteins seen in nature, enabling unprecedented capability and control in de novo protein design. However, current state-of-the-art diffusion models generate protein structures, which limits the scope of their training data and restricts generations to a small and biased subset of protein design space. Here, we introduce a general-purpose diffusion framework, EvoDiff, that combines evolutionary-scale data with the distinct conditioning capabilities of diffusion models for controllable protein generation in sequence space. EvoDiff generates high-fidelity, diverse, and structurally-plausible proteins that cover natural sequence and functional space. We show experimentally that EvoDiff generations express, fold, and exhibit expected secondary structure elements. Critically, EvoDiff can generate proteins inaccessible to structure-based models, such as those with disordered regions, while maintaining the ability to design scaffolds for functional structural motifs. We validate the universality of our sequence-based formulation by experimentally characterizing intrinsically-disordered mitochondrial targeting signals, metal-binding proteins, and protein binders designed using EvoDiff. We envision that EvoDiff will expand capabilities in protein engineering beyond the structure-function paradigm toward programmable, sequence-first design.

Finding Drug Repurposing Candidates for Neurodegenerative Diseases using Zebrafish Behavioral Profiles

Hernandez, T. D. R. — 2023-09-13

Drug repurposing can accelerate drug development while reducing the cost and risk of toxicity typically associated with de novo drug design. Several disorders lacking pharmacological solutions and exhibiting poor results in clinical trials - such as Alzheimers disease (AD) - could benefit from a cost-effective approach to finding new therapeutics. We previously developed a neural network model, Z-LaP Tracker, capable of quantifying behaviors in zebrafish larvae relevant to cognitive function, including activity, reactivity, swimming patterns, and optomotor response in the presence of visual and acoustic stimuli. Using this model, we performed a high-throughput screening of FDA-approved drugs to identify compounds that affect zebrafish larval behavior in a manner consistent with the distinct behavior induced by calcineurin inhibitors. Cyclosporine (CsA) and other calcineurin inhibitors have garnered interest for their potential role in the prevention of AD. We generated behavioral profiles suitable for cluster analysis, through which we identified 64 candidate therapeutics for neurodegenerative disorders.

Spontaneous anterior-posterior convergences of oscillation frequencies in the alpha band

Suzuki, S. — 2023-09-13

Alpha oscillation frequencies vary along the anterior-posterior axis, but they also dynamically converge. We investigated spontaneous anterior-posterior oscillation-frequency convergences while participants rested with their eyes open or closed, by tracking their oscillatory EEG activity in an extended alpha range (5-15 Hz) with appropriate temporal ({bsim}370 ms) and spectral ({bsim}1 Hz) resolutions. Oscillation-frequency convergences were prominent in the alpha band (8-12 Hz) and our analyses revealed three primary characteristics. First, the probability of an additional site joining a frequency convergence increased as more sites frequency-converged, suggesting that synergistic interactions drive anterior-posterior frequency convergences. Second, the oscillatory power at participating sites increased as more sites frequency-converged, suggesting that the synergistic interactions are mediated by regional synchronizations, being boosted by inter-regional frequency matching, facilitating the entrainment of additional regions. Third, frequency convergences generated two opposing phase gradients, posterior-behind and posterior-ahead, potentially mediating directional information flows to and from posterior regions. These gradients became steeper as more sites frequency-converged (while maintaining relatively constant levels of phase consistency), suggesting that frequency convergences increase the directionality of anterior-posterior information flows. Interestingly, when participants closed their eyes, the opposing phase gradients spatially organized, forming posterior-ahead gradients along the midline and posterior-behind gradients within each hemisphere, suggesting that closing eyes streamlines the opposing information flows. Taken together, these results suggest that synergistic interactions drive spontaneous anterior-posterior oscillation-frequency convergences in the alpha band, which may contribute to directional flows of information to and from posterior regions.

Synergistic anti-tumor activity, reduced pERK, and immuno-stimulatory cytokine profiles with 5-FU or ONC212 plus KRAS G12D inhibitor MRTX1133 in CRC and pancreatic cancer cells independent of G12D mutation

Tajiknia, V. — 2023-09-17

KRAS mutations occur in [~]40-50% of mCRC and are associated with aggressive disease that is refractory to anti-EGFR therapies. Pancreatic cancer harbors [~]90% KRAS driver gene mutation frequency. Small molecules targeting KRAS G12C gained FDA approval for KRAS G12C-mutated NSCLC. ONC212, a fluorinated imipridone with nM anti-cancer activity has preclinical efficacy against pancreatic cancer and other malignancies. MRTX1133, identified as a noncovalent selective KRAS G12D inhibitor that suppresses G12D signaling by binding to the switch II pocket thereby inhibiting protein-protein interactions. We investigated cell viability, drug synergies, pERK suppression and cytokine, chemokine or growth factor alterations following treatment with 5-Fluorouracil (5-FU) or ONC212 plus MRTX1133 in 6 human CRC and 4 human pancreatic cancer cell lines. IC50 sensitivities ranged from 7 to 12 M for 5-FU, 0.2-0.8 M for ONC212, and >100 nM to >5,000 nM for MRTX1133 (G12D N=4: LS513 >100, HPAF-II >1,000, SNUC2B >5,000, PANC-1 >5,000). For non-G12D, the range of IC50 for MRTX1133 was >1,000 to >5,000 nM for CRC lines with G12V, G13D, or WT KRAS (N=7). Synergies between MRTX1133 plus 5-FU or ONC212 were observed regardless of KRAS G12D mutation with combination indices of <0.5 indicating strong synergy. Observed synergies were greater with MRTX1133 plus ONC212 compared to MRTX1133 plus 5-FU. pERK was suppressed with mutant but not wild-type KRAS at nM MRTX1133 doses. Immunostimulatory profiles included reduction in IL8/CXCL8, MICA, Angiopoietin 2, VEGF and TNF-alpha and increase in IL-18/IL-1F4 with MRTX treatments and combinations. Our studies reveal preclinical activity of MRTX1133 alone or synergies when combined with 5-FU or ONC212 against mCRC and pancreatic cancer cells regardless of KRAS G12D mutation. The results suggest that KRAS G12V and KRAS G13D should be further considered in clinical trials including combination therapies involving MRTX1133 and 5-FU or ONC212.

Preclinical Synergistic Combination Therapy of Lurbinectedin with Irinotecan and 5-Fluorouracil in Pancreatic Cancer

Tummala, T. — 2023-09-17

Pancreatic cancer is a devastating disease with a poor prognosis. Novel chemotherapeutics in pancreatic cancer have shown limited success, illustrating the urgent need for new treatments. Lurbinectedin (PM01183; LY-01017) received FDA approval in 2020 for metastatic small cell lung cancer on or after platinum-based chemotherapy and is currently undergoing clinical trials in a variety of tumor types. Lurbinectedin stalls and degrades RNA Polymerase II and introduces breaks in DNA, causing subsequent apoptosis. We now demonstrate lurbinectedins highly efficient killing of human-derived pancreatic tumor cell lines PANC-1, BxPC-3, and HPAF-II as a single agent. We further demonstrate that a combination of lurbinectedin and irinotecan, a topoisomerase I inhibitor with FDA approval for advanced pancreatic cancer, results in synergistic killing of pancreatic tumor cells. Western blot analysis of combination therapy indicates an upregulation of {gamma}H2AX, a DNA damage marker, and the Chk1/ATR pathway, involved in replicative stress and DNA damage response. We further demonstrate that the triple combination between lurbinectedin, irinotecan, and 5-fluorouracil (5-FU) results in highly efficient killing of tumor cells. Our results are developing insights regarding molecular mechanisms underlying therapeutic efficacy of a novel combination drug treatment for pancreatic cancer.

PD-1 Mediated Regulation of Unique Activated CD8+ T Cells by NK Cells in the Submandibular Gland

Brossay, L. — 2023-09-17

The increasing utilization of anti-PD-1 immune checkpoint blockade (ICB) has led to the emergence of immune-related adverse events (irAEs), including sicca syndrome. Interestingly, we found that the submandibular gland (SMG) of PD-1 deficient mice harbors a large population of CD8+ T cells, reminiscing ICB induced sicca. This phenotype was also observed in the SMG of both NK cell-depleted C57BL/6 animals and NK cell-deficient animals. Mechanistically, using mice conditionally deficient for PD-L1 in the NK cell lineage, we discovered that NK cells regulate CD8+ T cell homeostasis via the PD-1/PD-L1 axis in this organ. Importantly, single-cell RNA sequencing of PD-1 deficient SMG CD8+ T cells reveals a unique transcriptional profile consistent with TCR activation. These cells have limited TCR diversity and phenotypically overlap with GzmK+ CD8+ T autoimmune cells identified in primary Sjogrens syndrome patients. These insights into NK cell immunoregulation in the SMG, and the consequences of disrupted CD8+ T cell homeostasis, provide opportunities for preventing the development of irAEs. HighlightsO_LIElevated CD8+ T cells in the submandibular gland (SMG) of PD-1 deficient mice parallel sicca-like irAEs seen in ICB patients. C_LIO_LIIn addition to their previously described hyporesponsive phenotype, NK cells in the SMG regulate CD8+ T cell homeostasis through the PD-L1/PD-1 axis. C_LIO_LIPD-1 deficient SMG CD8+ T cells display unique transcriptional profiles associated with proinflammatory functions, TCR activation, interferon stimulation, and exhaustion. C_LIO_LIOligoclonal expansion and similarities in TCR sequences indicate T cell activation and a preference for recognizing specific antigens. C_LI

Chitinase 3-like-1 (CHI3L1) in the Pathogenesis of Epidermal Growth Factor Receptor Mutant Non-Small Cell Lung Cancer

Kamle, S. — 2023-09-23

Non-small cell lung cancer (NSCLC) accounts for 85% of all lung cancers. In NSCLC, 10-20% of Caucasian patients and 30-50% of Asian patients have tumors with activating mutations in the Epidermal Growth Factor Receptor (EGFR). A high percentage of these patients exhibit favorable responses to treatment with tyrosine kinase inhibitors (TKI). Unfortunately, a majority of these patients develop therapeutic resistance with progression free survival lasting 9-18 months. The mechanisms that underlie the tumorigenic effects of EGFR and the ability of NSCLC to develop resistance to TKI therapies, however, are poorly understood. Here we demonstrate that CHI3L1 is produced by EGFR activation of normal epithelial cells, transformed epithelial cells with wild type EGFR and cells with cancer-associated, activating EGFR mutations. We also demonstrate that CHI3L1 auto-induces itself and feeds back to stimulate EGFR and its ligands. Highly specific antibodies against CHI3L1 (anti-CHI3L1/FRG) and TKI, individually and in combination, abrogated the effects of EGFR activation on CHI3L1 and the ability of CHI3L1 to stimulate the EGFR axis. Anti-CHI3L1 also interacted with osimertinib to reverse TKI therapeutic resistance and induce tumor cell death and inhibit pulmonary metastasis while stimulating tumor suppressor genes including KEAP1. CHI3L1 is a downstream target of EGFR that feeds back to stimulate and activate the EGFR axis. Anti-CHI3L1 is an exciting potential therapeutic for EGFR mutant NSCLC, alone and in combination with osimertinib or other TKIs.

The MUC19 gene in Denisovans, Neanderthals, and Modern Humans: An Evolutionary History of Recurrent Introgression and Natural Selection

Villanea, F. A. — 2023-09-25

We study the gene MUC19, for which some modern humans carry a Denisovan-like haplotype. MUC19 is a mucin, a glycoprotein that forms gels with various biological functions. We find diagnostic variants for the Denisovan-like MUC19 haplotype at high frequencies in admixed Latin American individuals, and at highest frequency in 23 ancient Indigenous American individuals, all predating population admixture with Europeans and Africans. We find that the Denisovan-like MUC19 haplotype is under positive selection and carries a higher copy number of a 30 base-pair variable number tandem repeat, and that copy numbers of this repeat are exceedingly high in American populations. Finally, some Neanderthals carry the Denisovan-like MUC19 haplotype, and that it was likely introgressed into human populations through Neanderthal introgression rather than Denisovan introgression. One-Sentence SummaryModern humans and Neanderthals carry a Denisovan variant of the MUC19 gene, which is under positive selection in populations of Indigenous American ancestry.

Simulation-based Benchmarking of Ancient Haplotype Inference for Detecting Population Structure

Medina-Tretmanis, J. — 2023-09-29

Paleogenomic data has informed us about the movements, growth, and relationships of ancient populations. It has also given us context for medically relevant adaptations that appear in present-day humans due to introgression from other hominids, and it continues to help us characterize the evolutionary history of humans. However, ancient DNA (aDNA) presents several practical challenges as various factors such as deamination, high fragmentation, environmental contamination of aDNA, and low amounts of recoverable endogenous DNA, make aDNA recovery and analysis more difficult than modern DNA. Most studies with aDNA leverage only SNP data, and only a few studies have made inferences on human demographic history based on haplotype data, possibly because haplotype estimation (or phasing) has not yet been systematically evaluated in the context of aDNA. Here, we evaluate how the unique challenges of aDNA can impact phasing quality. We also develop a software tool that simulates aDNA taking into account the features of aDNA as well as the evolutionary history of the population. We measured phasing error as a function of aDNA quality and demographic history, and found that low phasing error is achievable even for very ancient individuals ([~] 400 generations in the past) as long as contamination and read depth are adequate. Our results show that population splits or bottleneck events occurring between the reference and phased populations affect phasing quality, with bottlenecks resulting in the highest average error rates. Finally, we found that using estimated haplotypes, even if not completely accurate, is superior to using the simulated genotype data when reconstructing changes in population structure after population splits between present-day and ancient populations. AvailabilityAll software used for simulation and analysis is available at github.com/Jazpy/Paleogenomic-Datasim

Cross-Tissue Specificity of Pediatric DNA Methylation Associated with Cumulative Family Adversity

Chan, M. H.-M. — 2023-10-06

BackgroundCumulative family adversity (cumulative FA), characterized by co-occurring stressors in a family context, may be biologically embedded through DNA methylation (DNAm) and contribute to later health outcomes. Materials & MethodsWe compared epigenome-wide DNAm associated with cumulative FA in buccal epithelial cells (BECs; n=218) and peripheral blood mononuclear cells (PBMCs; n=51) from 7-13-year-old children in Canada, accounting for sex, age, predicted cell-type proportion, and genetic ancestry. ResultsHigher levels of cumulative FA were associated with DNAm at seven sites, primarily in stress- and immune-related genes, only in PBMCs. Negative mother-child interaction contributed to this association. ConclusionsThe findings of this study suggested that PBMC DNAm can be used as a marker for biological embedding of cumulative FA.

Cadherin-dependent adhesion is required for muscle stem cell niche anchorage and maintenance

Hung, M. — 2023-10-06

Adhesion between stem cells and their niche provides stable anchorage and signaling cues to sustain properties such as quiescence. Skeletal muscle stem cells (MuSCs) directly adhere to an adjacent myofiber via cadherin-catenin complexes. Previous studies on N- and M-cadherin function in MuSCs revealed that while N-cadherin is required for quiescence, they are collectively dispensable for MuSC niche localization and regenerative activity. While additional cadherins are expressed at low levels, these findings raise the possibility that cadherins are unnecessary for MuSC anchorage to the niche. To address this question, we conditionally removed from MuSCs {beta}- and {gamma}-catenin and, separately, E- and T-catenin, factors essential for cadherin-dependent adhesion. Catenin-deficient MuSCs break quiescence similarly to N-/M-cadherin-deficient MuSCs, but exit the niche, and are depleted. A combination of in vivo, ex vivo, and single cell RNA sequencing approaches reveal that MuSC attrition occurs via a single fate: precocious differentiation, reentry to the niche, and fusion to myofibers. These findings indicate that cadherin-catenin-dependent adhesion is required for anchorage of MuSCs to their niche and preservation of the stem cell compartment. Furthermore, separable, cadherin-regulated functions govern niche localization, quiescence, and stem cell maintenance in MuSCs. SUMMARY STATEMENTGenetic ablation of cadherin-based adhesion in skeletal muscle stem cells triggers activation, niche exit, precocious differentiation, and subsequent depletion of the stem cell pool.

Single cell expression and chromatin access of the Toxoplasma gondii lytic cycle identifies AP2XII-8 as an essential pivotal controller of a ribosome regulon

Lou, J. — 2023-10-06

Sequential lytic cycles driven by cascading transcriptional waves underlie pathogenesis in the apicomplexan parasite Toxoplasma gondii. This parasites unique division by internal budding, short cell cycle, and jumbled up classically defined cell cycle stages have restrained in-depth transcriptional program analysis. Here, unbiased transcriptome and chromatin accessibility maps throughout the lytic cell cycle were established at the single cell level. Correlated pseudo-timeline assemblies of expression and chromatin profiles mapped transcriptional versus chromatin level transition points promoting the cell division cycle. Sequential clustering analysis identified putatively functionally related gene groups facilitating parasite division. Promoter DNA motif mapping revealed patterns of combinatorial regulation. Pseudo-time trajectory analysis revealed transcriptional bursts at different cell cycle points. The dominant burst in G1 was driven by transcription factor AP2XII-8, which engages TGCATGCG/A and TATAAGCCG motifs, and promoted the expression of a regulon encoding 40 ribosomal proteins. Overall, the study provides integrated, multi-level insights into apicomplexan transcriptional regulation.

Differential encoding of odor and place in mouse piriform and entorhinal cortex

Mena, W. — 2023-10-08

The integration of olfactory and spatial information is critical for guiding animal behavior. The lateral entorhinal cortex (LEC) is reciprocally interconnected with cortical areas for olfaction and the hippocampus and thus ideally positioned to encode odor-place associations. Here, we used mini-endoscopes to record neural activity in the mouse piriform cortex (PCx) and LEC. We show that in head-fixed mice, odor identity could be decoded from LEC ensembles, but less accurately than from PCx. In mice freely navigating a linear track, LEC ensemble activity at the odor ports was dominated by spatial information. Spatial position along the linear track could be decoded from LEC and PCx activity, however, PCx but not LEC exhibited strong behavior-driven modulation of positional information. Together, our data reveal that information about odor cues and spatial context is differentially encoded along the PCx-LEC axis. Significance statementFor most animals, the sense of smell is essential for successfully navigating the environment to find food, shelter, and mates. However, how olfactory and spatial information is integrated in the brain to support olfactory-guided behaviors remains poorly understood. In mammals, candidate brain regions thought to support odor-place associations include the olfactory (piriform, PCx) cortex, entorhinal cortex, and hippocampus. We here characterize the activity of cells in the PCx and lateral entorhinal cortex (LEC) of freely moving mice in response to odor cues presented in a linear track. Using mini-endoscope microscopy and population coding analyses, we find that information about odors, spatial location, and behavior is differentially encoded along the PCx-LEC axis.

The CLAMP GA-binding transcription factor regulates heat stress-induced transcriptional repression by associating with 3D loop anchors.

Aguilera, J. — 2023-10-10

To survive exposure to heat stress (HS), organisms activate stress response genes and repress constitutive gene expression, thereby preventing the accumulation of potentially toxic RNA and protein products. Although many studies have elucidated the mechanisms that drive HS-induced activation of stress response genes across species, little is known about the mechanisms that repress constitutively expressed genes. Using nascent RNA-sequencing, we identify the first reported transcription factor (TF) that regulates repression of constitutive genes upon heat stress across species and define direct and indirect mechanisms of action by integrating 3D genomic approaches. We demonstrate that the CLAMP (Chromatin-linked adaptor for MSL complex proteins) GA-binding transcription factor (TF) regulates [~]75% of the HS-induced repression in Drosophila, a well-established model for understanding the mechanisms of HS-induced gene regulation. Using Micro-C, we demonstrate that heat stress induces widespread changes in local 3D chromatin looping, which are significantly associated with HS-induced transcriptional changes. Overall, we identify CLAMP as the first reported TF that induces HS repression, which modulates 3D chromatin looping through both direct mechanisms and indirect mechanisms. Moreover, we present the highest-resolution heat stress 3D genomic dataset available in Drosophila, providing a key resource for generating mechanistic insights into how temperature regulates 3D genomic contacts.

Multioviz: an interactive platform for in silico perturbation and interrogation of gene regulatory networks

Xie, H. — 2023-10-11

In this paper, we aim to build a tool that will help bridge the gap between high-dimensional computation and wet-lab experimentation by allowing users to interrogate genomic signatures at multiple molecular levels and identify best next actionable steps for downstream decision making. We introduce Multioviz: a publicly accessible R package and web application platform to easily perform in silico hypothesis testing of generated gene regulatory networks. We demonstrate the utility of Multioviz by conducting an end-to-end analysis in a statistical genetics application focused on measuring the effect of in silico perturbations of complex trait architecture. By using a real data set from the Wellcome Trust Centre for Human Genetics, we both recapitulate previous findings and propose hypotheses about the genes involved in the percentage of immune CD8+ cells found in heterogeneous stocks of mice. Source code for the Multioviz R package is available at https://github.com/lcrawlab/multio-viz and an interactive version of the platform is available at multioviz.ccv.brown.edu.

Pseudo-linear Summation explains Neural Geometry of Multi-finger Movements in Human Premotor Cortex

Shah, N. P. — 2023-10-12

How does the motor cortex combine simple movements (such as single finger flexion/extension) into complex movements (such hand gestures or playing piano)? Motor cortical activity was recorded using intracortical multi-electrode arrays in two people with tetraplegia as they attempted single, pairwise and higher order finger movements. Neural activity for simultaneous movements was largely aligned with linear summation of corresponding single finger movement activities, with two violations. First, the neural activity was normalized, preventing a large magnitude with an increasing number of moving fingers. Second, the neural tuning direction of weakly represented fingers (e.g. middle) changed significantly as a result of the movement of other fingers. These deviations from linearity resulted in non-linear methods outperforming linear methods for neural decoding. Overall, simultaneous finger movements are thus represented by the combination of individual finger movements by pseudo-linear summation.

The MuSK-BMP pathway regulates synaptic Nav1.4 localization and muscle excitability

Fish, L. A. — 2023-10-29

The neuromuscular junction (NMJ) is the linchpin of nerve-evoked muscle contraction. Broadly considered, the function of the NMJ is to transduce a nerve action potential into a muscle fiber action potential (MFAP). Efficient information transfer requires both cholinergic signaling, responsible for the generation of endplate potentials (EPPs), and excitation, the activation of postsynaptic voltage-gated sodium channels (Nav1.4) to trigger MFAPs. In contrast to the cholinergic apparatus, the signaling pathways that organize Nav1.4 and muscle fiber excitability are poorly characterized. Muscle-specific kinase (MuSK), in addition to its Ig1 domain-dependent role as an agrin-LRP4 receptor, is also a BMP co-receptor that binds BMPs via its Ig3 domain and shapes BMP-induced signaling and transcriptional output. Here we probed the function of the MuSK-BMP pathway at the NMJ using mice lacking the MuSK Ig3 domain ( {Delta}Ig3-MuSK). Synapses formed normally in {Delta}Ig3-MuSK animals, but the postsynaptic apparatus was fragmented from the first weeks of life. Anatomical denervation was not observed at any age examined. Moreover, spontaneous and nerve-evoked acetylcholine release, AChR density, and endplate currents were comparable to WT. However, trains of nerve-evoked MFAPs in {Delta}Ig3-MuSK muscle were abnormal as revealed by increased jitter and blocking in single fiber electromyography. Further, nerve-evoked compound muscle action potentials (CMAPs), as well as twitch and tetanic muscle torque force production, were also diminished. Finally, Nav1.4 levels were reduced at {Delta}Ig3-MuSK synapses but not at the extrajunctional sarcolemma, indicating that the observed excitability defects are the result of impaired localization of this voltage-gated ion channel at the NMJ. We propose that MuSK plays two distinct roles at the NMJ: as an agrin-LRP4 receptor necessary for establishing and maintaining cholinergic signaling, and as a BMP co-receptor required for maintaining proper Nav1.4 density, nerve-evoked muscle excitability and force production. The MuSK-BMP pathway thus emerges as a target for modulating excitability and functional innervation, which are defective in conditions such as congenital myasthenic syndromes and aging. Significance StatementThe neuromuscular junction (NMJ) is required for nerve-evoked muscle contraction and movement, and its function is compromised during aging and disease. Although the mechanisms underlying neurotransmitter release and cholinergic response at this synapse have been studied extensively, the machinery necessary for nerve-evoked muscle excitation are incompletely characterized. We show that MuSK (Muscle-specific kinase), in its role as a BMP co-receptor, regulates NMJ structure as well as the localization of the voltage-gated sodium channels necessary for full nerve-evoked muscle fiber excitation and force production. This novel function of MuSK is structurally and mechanistically distinct from its role in organizing cholinergic machinery. The MuSK-BMP pathway thus presents a new opportunity to understand mechanisms that may preserve or enhance neuromuscular excitability in the face of aging and disease.

Biophysical modeling of frontocentral ERP generation links circuit-level mechanisms of action-stopping to a behavioral race model

Diesburg, D. A. — 2023-10-30

Human frontocentral event-related potentials (FC-ERPs) are ubiquitous neural correlates of cognition and control, but their generating multiscale mechanisms remain mostly unknown. We used the Human Neocortical Neurosolver(HNN)s biophysical model of a canonical neocortical circuit under exogenous thalamic and cortical drive to simulate the cell and circuit mechanisms underpinning the P2, N2, and P3 features of the FC-ERP observed after Stop-Signals in the Stop-Signal task (SST). We demonstrate that a sequence of simulated external thalamocortical and cortico-cortical drives can produce the FC-ERP, similar to what has been shown for primary sensory cortices. We used this model of the FC-ERP to examine likely circuit-mechanisms underlying FC-ERP features that distinguish between successful and failed action-stopping. We also tested their adherence to the predictions of the horse-race model of the SST, with specific hypotheses motivated by theoretical links between the P3 and Stop process. These simulations revealed that a difference in P3 onset between successful and failed Stops is most likely due to a later arrival of thalamocortical drive in failed Stops, rather than, for example, a difference in effective strength of the input. In contrast, the same model predicted that early thalamocortical drives underpinning the P2 and N2 differed in both strength and timing across stopping accuracy conditions. Overall, this model generates novel testable predictions of the thalamocortical dynamics underlying FC-ERP generation during action-stopping. Moreover, it provides a detailed cellular and circuit-level interpretation that supports links between these macroscale signatures and predictions of the behavioral race model. Significance statementThe frontocentral event-related potential (FC-ERP) is an easily-measurable neural correlate of cognition and control. However, the cortical dynamics that produce this signature in humans are complex, limiting the ability of researchers to make predictions about its underlying mechanisms. In this study, we used the biophysical model included in the open-source Human Neocortical Neurosolver software to simulate and evaluate the likely cellular and circuit mechanisms that underlie the FC-ERP in the Stop-Signal task. We modeled mechanisms of the FC-ERP during successful and unsuccessful stopping, generating testable predictions regarding Stop-associated computations in human frontal cortex. Moreover, the resulting model parameters provide a starting point for simulating mechanisms of the FC-ERP and other frontal scalp EEG signatures in other task conditions and contexts.

PDZD8 promotes autophagy at ER-Lysosome contact sites to regulate synaptogenesis

Thakur, R. S. — 2023-10-30

Building synaptic connections, which are often far from the soma, requires coordinating a host of cellular activities from transcription to protein turnover, placing a high demand on intracellular communication. Membrane contact sites (MCSs) formed between cellular organelles have emerged as key signaling hubs for coordinating an array of cellular activities. We have found that the endoplasmic reticulum (ER) MCS tethering protein PDZD8 is required for activity-dependent synaptogenesis. PDZD8 is sufficient to drive ectopic synaptic bouton formation through an autophagy-dependent mechanism and required for basal synapse formation when autophagy biogenesis is limited. PDZD8 functions at ER-late endosome/lysosome (LEL) MCSs to promote lysosome maturation and accelerate autophagic flux. Mutational analysis of PDZD8s SMP domain further suggests a role for lipid transfer at ER-LEL MCSs. We propose that PDZD8-dependent lipid transfer from ER to LELs promotes lysosome maturation to increase autophagic flux during periods of high demand, including activity-dependent synapse formation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/564828v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1c6b752org.highwire.dtl.DTLVardef@12d77c3org.highwire.dtl.DTLVardef@11d50corg.highwire.dtl.DTLVardef@ba404f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Low-dimensional neuronal population dynamics in anterior superior temporal gyrus reactivate phonetic representations during semantic processing

Orepic, P. — 2023-10-31

AbstractSpeech processing involves a hierarchy of cognitive layers from low-level phonetic to high-level seman-tic representations. However, interactions are not only feed-forward; Feedback mechanisms are crucial for real-time, accurate speech recognition. Yet how distant levels interface during speech processing remains unclear. Here, we analyzed intracortical recordings from 624 neurons across three human participants implanted with microelectrode arrays in the anterior superior temporal gyrus during an auditory semantic categorization task and natural speech perception. We identified distinct neural subspaces, or manifolds, for semantic and phonetic features, with a functional separation of the corresponding low-dimensional dynamics. We contrasted a bottom-up cumulative and a top-down predictive hypothesis on phonetic-semantic temporal alignment, and found phonetic alignment to word-level semantic representations, signaling top-down prediction. These effects were consistent across participants at the spiking level, and remained undetected in adjacent ECoG recordings. These findings demonstrate the reorganization of neuronal population dynamics supporting phonetic representations during semantic prediction.

Motivational context determines the impact of aversive outcomes on mental effort allocation

Prater Fahey, M. — 2023-11-01

It is well known that people will exert effort on a task if sufficiently motivated, but how they distribute these efforts across different strategies (e.g., efficiency vs. caution) remains uncertain. Past work has shown that people invest effort differently for potential positive outcomes (rewards) versus potential negative outcomes (penalties). However, this research failed to account for differences in the context in which negative outcomes motivate someone - either as punishment or reinforcement. It is therefore unclear whether effort profiles differ as a function of outcome valence, motivational context, or both. Using computational modeling and our novel Multi-Incentive Control Task, we show that the influence of aversive outcomes on ones effort profile is entirely determined by their motivational context. Participants (N:91) favored increased caution in response to larger penalties for incorrect responses, and favored increased efficiency in response to larger reinforcement for correct responses, whether positively or negatively incentivized. Statement of RelevancePeople have to constantly decide how to allocate their mental effort, and in doing so can be motivated by both the positive outcomes that effort accrues and the negative outcomes that effort avoids. For example, someone might persist on a project for work in the hopes of being promoted or to avoid being reprimanded or even fired. Understanding how people weigh these different types of incentives is critical for understanding variability in human achievement as well as sources of motivational impairments (e.g., in major depression). We show that people not only consider both potential positive and negative outcomes when allocating mental effort, but that the profile of effort they engage under negative incentives differs depending on whether that outcome is contingent on sustaining good performance (negative reinforcement) or avoiding bad performance (punishment). Clarifying the motivational factors that determine effort exertion is an important step for understanding motivational impairments in psychopathology.

Increasing hub disruption parallels dementia severity in autosomal dominant Alzheimer disease

Tu, J. C. — 2023-11-01

Hub regions in the brain, recognized for their roles in ensuring efficient information transfer, are vulnerable to pathological alterations in neurodegenerative conditions, including Alzheimer Disease (AD). Given their essential role in neural communication, disruptions to these hubs have profound implications for overall brain network integrity and functionality. Hub disruption, or targeted impairment of functional connectivity at the hubs, is recognized in AD patients. Computational models paired with evidence from animal experiments hint at a mechanistic explanation, suggesting that these hubs may be preferentially targeted in neurodegeneration, due to their high neuronal activity levels--a phenomenon termed "activity-dependent degeneration". Yet, two critical issues were unresolved. First, past research hasnt definitively shown whether hub regions face a higher likelihood of impairment (targeted attack) compared to other regions or if impairment likelihood is uniformly distributed (random attack). Second, human studies offering support for activity-dependent explanations remain scarce. We applied a refined hub disruption index to determine the presence of targeted attacks in AD. Furthermore, we explored potential evidence for activity-dependent degeneration by evaluating if hub vulnerability is better explained by global connectivity or connectivity variations across functional systems, as well as comparing its timing relative to amyloid beta deposition in the brain. Our unique cohort of participants with autosomal dominant Alzheimer Disease (ADAD) allowed us to probe into the preclinical stages of AD to determine the hub disruption timeline in relation to expected symptom emergence. Our findings reveal a hub disruption pattern in ADAD aligned with targeted attacks, detectable even in pre-clinical stages. Notably, the disruptions severity amplified alongside symptomatic progression. Moreover, since excessive local neuronal activity has been shown to increase amyloid deposition and high connectivity regions show high level of neuronal activity, our observation that hub disruption was primarily tied to regional differences in global connectivity and sequentially followed changes observed in A{beta} PET cortical markers is consistent with the activity-dependent degeneration model. Intriguingly, these disruptions were discernible 8 years before the expected age of symptom onset. Taken together, our findings not only align with the targeted attack on hubs model but also suggest that activity-dependent degeneration might be the cause of hub vulnerability. This deepened understanding could be instrumental in refining diagnostic techniques and developing targeted therapeutic strategies for AD in the future.

Using behavioral biomarkers to redefine epochs of spontaneous recovery following spinal cord injury

Eisdorfer, J. T. — 2023-11-02

The brain-spinal cord axis generates movement by assembling motor primitives into coordinated sequences. Spinal cord injury (SCI) disrupts this neuroaxis, impairing not only locomotion, but the full repertoire of behavior. Traditional scales for quantifying recovery collapse this complexity into predefined locomotor-focused criteria that obscure heterogeneity in recovery. To quantify the full behavioral repertoire following SCI, we adapted motion sequencing (MoSeq) to identify sub-second behavioral "syllables" and capture their usage and sequential organization without predefined features. We identified biomechanically distinct variants within syllable classes that are shared across injury severities and found that recovery is jointly structured by injury severity and individual mouse identity. Changes in sequences, however, unfold along a conserved temporal trajectory. By compressing behavior into a single metric, we uncovered clusters of coevolving locomotor and non-locomotor behaviors. These results frame SCI recovery with repertoire-level changes, where adaptive strategies emerge from constrained access to motor primitives and their sequences.

Neuronal LRP4 directs the development, maturation, and cytoskeletal organization of peripheral synapses

DePew, A. T. — 2023-11-03

Synapse development requires multiple signaling pathways to accomplish the myriad of steps needed to ensure a successful connection. Transmembrane receptors on the cell surface are optimally positioned to facilitate communication between the synapse and the rest of the neuron and often function as synaptic organizers to synchronize downstream signaling events. One such organizer, the LDL receptor-related protein LRP4, is a cell surface receptor most well-studied postsynaptically at mammalian neuromuscular junctions. Recent work, however, has identified emerging roles for LRP4 as a presynaptic molecule, but how LRP4 acts as a presynaptic organizer, what roles LRP4 plays in organizing presynaptic biology, and the downstream mechanisms of LRP4 are not well understood. Here we show that LRP4 functions presynaptically at Drosophila neuromuscular synapses, acting in motor neurons to instruct multiple aspects of pre- and postsynaptic development. Loss of presynaptic LRP4 results in a range of developmental defects, impairing active zone organization, synapse growth, physiological function, microtubule organization, synaptic ultrastructure, and synapse maturation. We further demonstrate that LRP4 promotes most aspects of presynaptic development via a downstream SR-protein kinase, SRPK79D. SRPK79D overexpression suppresses synaptic defects associated with loss of lrp4. These data demonstrate a function for LRP4 as a peripheral synaptic organizer acting presynaptically, highlight a downstream mechanism conserved with its CNS function, and indicate previously unappreciated roles for LRP4 in cytoskeletal organization, synapse maturation, and active zone organization, underscoring its developmental importance.

GGA1 interacts with the endosomal Na+/H+ Exchanger NHE6 governing localization to the endosome compartment

Ma, L. — 2023-11-09

Mutations in the endosomal Na+/H+ exchanger (NHE6) cause Christianson syndrome (CS), an X-linked neurological disorder. Previous studies have shown that NHE6 functions in regulation of endosome acidification and maturation in neurons. Using yeast two-hybrid screening with the NHE6 carboxyl-terminus as bait, we identify Golgi-associated, Gamma adaptin ear containing, ARF binding protein 1 (GGA1) as an interacting partner for NHE6. We corroborated the NHE6-GGA1 interaction using co-immunoprecipitation (co-IP): using over-expressed constructs in mammalian cells; and co-IP of endogenously-expressed GGA1 and NHE6 from neuroblastoma cells, as well as from mouse brain. We demonstrate that GGA1 interacts with organellar NHEs (NHE6, NHE7 and NHE9) but not with cell-surface localized NHEs (NHE1 and NHE5). By constructing hybrid NHE1/NHE6 exchangers, we demonstrate that the cytoplasmic tail of NHE6 is necessary and sufficient for interactions with GGA1. We demonstrate the co-localization of NHE6 and GGA1 in cultured, primary hippocampal neurons, using super-resolution microscopy. We test the hypothesis that the interaction of NHE6 and GGA1 functions in the localization of NHE6 to the endosome compartment. Using subcellular fractionation experiments, we show that NHE6 is mis-localized in GGA1 knockout cells wherein we find less NHE6 in endosomes but more NHE6 transport to lysosomes, and more Golgi retention of NHE6 with increased exocytosis to the surface plasma membrane. Consistent with NHE6 mis-localization, and Golgi retention, we find the intra-luminal pH in Golgi to be alkalinized. Our study demonstrates a new interaction between NHE6 and GGA1 which functions in the localization of this intra-cellular NHE to the endosome compartment.

Structure and position-specific interactions of prion-like domains in transcription factor Efg1 phase separation

Zheng, T. — 2023-11-09

Candida albicans, a prominent member of the human microbiome, can make an opportunistic switch from commensal coexistence to pathogenicity accompanied by an epigenetic shift between the white and opaque cell states. This transcriptional switch is under precise regulation by a set of transcription factors (TFs), with Enhanced Filamentous Growth Protein 1 (Efg1) playing a central role. Previous research has emphasized the importance of Egf1s prion-like domain (PrLD) and the proteins ability to undergo phase separation for the white-to-opaque transition of C. albicans. However, the underlying molecular mechanisms of Efg1 phase separation have remained underexplored. In this study, we delved into the biophysical basis of Efg1 phase separation, revealing the significant contribution of both N-terminal (N) and C-terminal (C) PrLDs. Through NMR structural analysis, we found that Efg1 N-PrLD and C-PrLD are mostly disordered though have prominent partial -helical secondary structures in both domains. NMR titration experiments suggest that the partially helical structures in N-PrLD act as hubs for self-interaction as well as Efg1 interaction with RNA. Using condensed-phase NMR spectroscopy, we uncovered diverse amino acid interactions underlying Efg1 phase separation. Particularly, we highlight the indispensable role of tyrosine residues within the transient -helical structures of PrLDs particularly in the N-PrLD compared to the C-PrLD in stabilizing phase separation. Our study provides evidence that the transient -helical structure is present in the phase separated state and highlights the particular importance of aromatic residues within these structures for phase separation. Together, these results enhance the understanding of C. albicans TF interactions that lead to virulence and provide a crucial foundation for potential antifungal therapies targeting the transcriptional switch. Statement of SignificancePhase separated condensates have been found across the domains of life and many types of cells. To understand their varied functions, seeing the residue-by-residue details of the structure and interactions of component protein constituents is essential. A set of transcription factors that phase-separate controls cell fate of the pathogenic yeast Candida albicans. Here, we examine the structural and interaction details of a main regulator of this process, Efg1, using NMR spectroscopy and biochemical assays. We find Efg1s phase-separating domains are not entirely disordered as often assumed but in fact contain helical regions that persist upon phase separation. We also reveal the balance of contacts formed in the condensed phase and the importance of specific residues and regions in phase separation.

Distinct and interdependent functions of three RING proteins regulate recombination during mammalian meiosis

Ito, M. — 2023-11-10

During meiosis, each pair of homologous chromosomes becomes connected by at least one crossover, as required for accurate segregation at the first division, and adjacent crossovers are widely separated thereby limiting total numbers. In coarsening models, crossover patterning results from nascent recombination sites competing to accrue a limiting pro-crossover RING-domain protein (COR) that diffuses between synapsed chromosomes. Here we delineate the localization dynamics of three mammalian CORs in mouse and determine their interdependencies. RNF212, HEI10 and a new member RNF212B show divergent spatiotemporal dynamics along synapsed chromosomes, including profound differences in spermatocytes and oocytes, that are not easily reconciled by elementary coarsening models. Contrasting mutant phenotypes and genetic requirements indicate that RNF212B, RNF212, and HEI10 play distinct but interdependent functions in regulating meiotic recombination and coordinating the events of meiotic prophase-I by integrating signals from DNA breaks, homolog synapsis, the cell-cycle, and incipient crossover sites. SIGNIFICANCEMeiosis produces haploid gametes by precisely halving the chromosome complement. Crossing over between homologous chromosomes (homologs) is essential for their accurate segregation and defects are associated with infertility, miscarriage, and congenital disease. Factors that ensure crossing over between each pair of homologs include mammalian RING-domain proteins RNF212, HEI10, and RNF212B, alleles of which are linked to infertility and heritable variation in crossover rate. This study focuses on understanding the functions and relationships between pro-crossover RING proteins (CORs) in mouse, providing important insights into their roles in regulating recombination, the DNA repair process that gives rise to crossovers. Notably, chromosomal localization dynamics of the three CORs are distinct and show striking sexual dimorphism with important implications for models of crossover control.

Intrinsically disordered regions and RNA binding domains contribute to protein enrichment in biomolecular condensates in Xenopus oocytes

O'Connell, L. C. — 2023-11-10

Proteins containing both intrinsically disordered regions (IDRs) and RNA binding domains (RBDs) can phase separate in vitro, forming bodies similar to cellular biomolecular condensates. However, how IDR and RBD domains contribute to in vivo recruitment of proteins to biomolecular condensates remains poorly understood. Here, we analyzed the roles of IDRs and RBDs in L-bodies, biomolecular condensates present in Xenopus oocytes. We show that a cytoplasmic isoform of hnRNPAB, which contains two RBDs and an IDR, is highly enriched in L-bodies. While both of these domains contribute to hnRNPAB self-association and phase separation in vitro and mediate enrichment into L-bodies in oocytes, neither the RBDs nor the IDR replicate the localization of full-length hnRNPAB. Our results suggest a model where the additive effects of the IDR and RBDs regulate hnRNPAB partitioning into L-bodies. This model likely has widespread applications as proteins containing RBD and IDR domains are common biomolecular condensate residents.

Multi-timescale reinforcement learning in the brain

Masset, P. — 2023-11-14

To thrive in complex environments, animals and artificial agents must learn to act adaptively to maximize fitness and rewards. Such adaptive behavior can be learned through reinforcement learning1, a class of algorithms that has been successful at training artificial agents2-6 and at characterizing the firing of dopamine neurons in the midbrain7-9. In classical reinforcement learning, agents discount future rewards exponentially according to a single time scale, controlled by the discount factor. Here, we explore the presence of multiple timescales in biological reinforcement learning. We first show that reinforcement agents learning at a multitude of timescales possess distinct computational benefits. Next, we report that dopamine neurons in mice performing two behavioral tasks encode reward prediction error with a diversity of discount time constants. Our model explains the heterogeneity of temporal discounting in both cue-evoked transient responses and slower timescale fluctuations known as dopamine ramps. Crucially, the measured discount factor of individual neurons is correlated across the two tasks suggesting that it is a cell-specific property. Together, our results provide a new paradigm to understand functional heterogeneity in dopamine neurons, a mechanistic basis for the empirical observation that humans and animals use non-exponential discounts in many situations 10-14, and open new avenues for the design of more efficient reinforcement learning algorithms.

tRNA modification enzyme-dependent redox homeostasis regulates synapse formation and memory

Madhwani, K. R. — 2023-11-14

Post-transcriptional modification of RNA regulates gene expression at multiple levels. ALKBH8 is a tRNA modifying enzyme that methylates wobble uridines in specific tRNAs to modulate translation. Through methylation of tRNA-selenocysteine, ALKBH8 promotes selenoprotein synthesis and regulates redox homeostasis. Pathogenic variants in ALKBH8 have been linked to intellectual disability disorders in the human population, but the role of ALKBH8 in the nervous system is unknown. Through in vivo studies in Drosophila, we show that ALKBH8 controls oxidative stress in the brain to restrain synaptic growth and support learning and memory. ALKBH8 null animals lack wobble uridine methylation and exhibit a global reduction in protein synthesis, including a specific decrease in selenoprotein levels. Loss of ALKBH8 or independent disruption of selenoprotein synthesis results in ectopic synapse formation. Genetic expression of antioxidant enzymes fully suppresses synaptic overgrowth in ALKBH8 null animals, confirming oxidative stress as the underlying cause of dysregulation. ALKBH8 animals also exhibit associative learning and memory impairments that are reversed by pharmacological antioxidant treatment. Together, these findings demonstrate the critical role of tRNA modification in redox homeostasis in the nervous system and reveal antioxidants as a potential therapy for ALKBH8-associated intellectual disability. Significance StatementtRNA modifying enzymes are emerging as important regulators of nervous system development and function due to their growing links to neurological disorders. Yet, their roles in the nervous system remain largely elusive. Through in vivo studies in Drosophila, we link tRNA methyltransferase-regulated selenoprotein synthesis to synapse development and associative memory. These findings demonstrate the key role of tRNA modifiers in redox homeostasis during nervous system development and highlight the potential therapeutic benefit of antioxidant-based therapies for cognitive disorders linked to dysregulation of tRNA modification.

Only cortical prediction error signals are involved in visual learning, despite availability of subcortical prediction error signals

Kim, D. — 2023-11-15

Both the midbrain systems, encompassing the ventral striatum (VS), and the cortical systems, including the dorsal anterior cingulate cortex (dACC), play roles in reinforcing and enhancing learning. However, the specific contributions of signals from these regions in learning remains unclear. To investigate this, we examined how VS and dACC are involved in visual perceptual learning (VPL) through an orientation discrimination task. In the primary experiment, subjects fasted for 5 hours before each of 14 days of training sessions and 3 days of test sessions. Subjects were rewarded with water for accurate trial responses. During the test sessions, BOLD signals were recorded from regions including VS and dACC. Although BOLD signals in both areas were associated with positive and negative RPEs, only those in dACC associated with negative RPE showed a significant correlation with performance improvement. Additionally, no significant correlation was observed between BOLD signals associated with RPEs in VS and dACC. These results suggest that although signals associated with positive and negative RPEs from both midbrain and cortical systems are readily accessible, only RPE signals in the prefrontal system, generated without linking to RPE signals in VS, are utilized for the enhancement of VPL.

Noise reduction as a unified mechanism of perceptual learning in humans, macaques, and convolutional neural networks

Cheng, Y.-A. — 2023-11-16

Visual perceptual learning (VPL), defined as long-term improvement in a visual task, is considered a crucial tool for elucidating underlying visual and brain plasticity. However, the identification of a unified theory of VPL has long been controversial. Multiple existing models have proposed diverse mechanisms, including improved signal-to-noise ratio, changes in tuning curves, and reduction of noise correlations, as major contributors to improved neural representations associated with VPL. However, each model only accounts for specific aspects of the empirical findings, and there exists no theory that can comprehensively explain all empirical results. Here, we argue that all neural changes at single units can be conceptualized as geometric transformations of population response manifolds in a high-dimensional neural space. This approach enables conflicting major models of VPL to be quantitatively tested and compared within a unified computational theory. Following this approach, we found that changes in tuning curves and noise correlations, as emphasized by previous models, make no significant contributions to improved population representations by visual training. Instead, we identified neural manifold shrinkage due to reduced trial-by-trial neural response variability, a previously unexplored factor, as the primary mechanism underlying improved population representations. Furthermore, we showed that manifold shrinkage successfully accounts for learning effects across various domains, including artificial neural responses in deep neural networks trained on typical VPL tasks, multivariate BOLD signals in humans, and multi-unit activities in monkeys. These converging results suggest that our neural geometry theory offers a quantitative and comprehensive approach to explain a wide range of empirical results and to reconcile previously conflicting models of VPL.

Androgen receptor signaling blockade enhances NK cell-mediated killing of prostate cancer cells and sensitivity to NK cell checkpoint blockade

Pinho-Schwermann, M. — 2023-11-17

BackgroundThe blockade of the androgen receptor (AR) pathway is an effective treatment for prostate cancer (PCa), but many patients progress to metastatic castration-resistant prostate cancer (mCRPC). Treatments for mCRPC include AR inhibitors (ARi), chemotherapy, PARP inhibitors, and radioligands. Checkpoint inhibitor activity is limited to a small subset of MSI-H mCRPC. AR signaling modulates CD8+ T cell function, but its impact on natural killer (NK) cell cytotoxicity is unknown. We investigated the effect of ARi on NK cell activation, cytokine secretion, NKG2A expression, and NK cell-mediated killing of PCa cells in vitro. MethodsPCa cell lines (LNCaP, 22Rv1, DU145, PC3) were co-cultured with NK-92 and treated with ARi (enzalutamide [enza], darolutamide [daro]) alone or in combination with anti-NKG2A antibody monalizumab. Immune cell-mediated tumor cell killing and cytokine secretion were quantified. NK cell expression of NKG2A and PCa cell expression of HLA-E were investigated by flow cytometry. The AR-negative cell lines PC3 and DU145 were stably transduced with an AR expression vector to evaluate the AR modulation of HLA-E. To assess the in vivo combination of NKG2A blockade and ARi therapy in vivo, Cas9 was used to genetically ablate the murine HLA-E ortholog, H2-T23, from RM-1 murine PCa cells. H2-T23 knockout and control cells were grown subcutaneously in castrated C57BL/6 mice and treated with daro or control. The activation status of peripheral blood NK cell isolated from patients with PCa before and after initiation of androgen deprivation therapy (ADT) was evaluated by flow cytometry. ResultsARi activated NK cells and significantly increased immune-mediated NK-92 cell killing of PCa cells. IFN-{gamma} and TRAIL mediated ARi-induced NK cell activation. ARi increased expression of the inhibitory receptor NKG2A on NK cells, and immune killing of PCa cells was enhanced with the combination of ARi and monalizumab. ARi also increased the expression of HLA-E, the ligand of NKG2A, on PCa cell lines. By transducing AR into AR-negative PC3 and DU145, we demonstrated that androgen signaling regulates HLA-E expression. In a mouse model of PCa, HLA-E knockout synergized with darolutamide to increase NK cell activation. NK cells derived from patients with metastatic PCa exhibited increased expression of Granzyme B and Perforin following ARi treatment. ConclusionsARi activates NK cells via IFN-{gamma} and TRAIL and promotes the killing of PCa cells. ARi also upregulates expression of HLA-E on PCa which may suppress the innate immune response against PCa. ARi-mediated NK cell killing of PCa cells was enhanced by NKG2A blockade. These results support novel immunotherapeutic strategies for PCa targeting NK activation through the combination of ARi and monalizumab. Graphical AbstractAndrogen receptor inhibitors (ARi) enhance NK cell-mediated killing of prostate cancer cells and sensitivity to NK cell checkpoint NKG2A blockade. ARi upregulate the NK cell inhibitor ligand (HLA-E) mediating suppression NK cell killing of PCa. This regulation is dependent on a functional AR signal on tumor cell lines. Adding an anti-NKG2a-HLA-E mAb with ARi further enhances the NK cell-mediated killing of PCa. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=139 SRC="FIGDIR/small/567201v2_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@ba35e6org.highwire.dtl.DTLVardef@128a2e7org.highwire.dtl.DTLVardef@def16aorg.highwire.dtl.DTLVardef@bb72e7_HPS_FORMAT_FIGEXP M_FIG C_FIG

Cortical spheroids show strain-dependent cell viability loss and neurite disruption following sustained compression injury

Gonzalez-Cruz, R. D. — 2023-11-17

Sustained compressive injury (SCI) in the brain is observed in numerous injury and pathological scenarios, including tumors, ischemic stroke, and traumatic brain injury-related tissue swelling. Sustained compressive injury is characterized by tissue loading over time, and currently, there are few in vitro models suitable to study neural cell responses to strain-dependent sustained compressive injury. Here, we present an in vitro model of sustained compressive neural injury via centrifugation. Spheroids were made from neonatal rat cortical cells seeded at 4000 cells/spheroid and cultured for 14 days in vitro. A subset of spheroids was centrifuged at 209 or 419 rad/s for 2 minutes. Modeling the physical compression of the spheroids via finite element analyses, we found that spheroids centrifuged at 209 and 419 rad/s experienced pressures of 38 kPa and 149 kPa, respectively, and compressive strains of 18% and 35%, respectively. Quantification of LIVE-DEAD assay and Hoechst 33342 nuclear staining showed that centrifuged spheroids exhibited significantly higher DNA damage than control spheroids at 2, 8, and 24 hours post-injury. Immunohistochemistry of {beta}3-tubulin networks at 2, 8, and 24 hours post-centrifugation injury showed increasing degradation of microtubules over time with increasing compressive strain. Our findings show that cellular injuries occur as a result of specific levels and timings of sustained compressive tissue strains. This experimental compressive injury model provides an in vitro platform to examine cellular injury to gain insights into brain injury that could be targeted with therapeutic strategies.

Control of T-shaped Bifurcation by Multiple Guidance Mechanisms during Dorsal Funiculus Development in the Spinal Cord

Curran, B. M. — 2023-11-18

The dorsal funiculus in the spinal cord relays somatosensory information to the brain. It is made of T-shaped bifurcation of dorsal root ganglion (DRG) sensory axons. Our previous study has shown that Slit signaling is required for proper guidance during bifurcation, but loss of Slit does not affect all DRG axons. Here, we examined the role of the extracellular molecule Netrin-1 (Ntn1). Using wholemount staining with tissue clearing, we showed that mice lacking Ntn1 have axons escaping from the dorsal funiculus at the time of bifurcation. Genetic labeling confirmed that these misprojecting axons come from DRG neurons. Single axon analysis showed that loss of Ntn1 does not affect bifurcation but rather alters turning angles. To distinguish their guidance functions, we examined mice with triple deletion of Ntn1, Slit1, and Slit2 and found a completely disorganized dorsal funiculus. Comparing mice with different genotypes using immunolabeling and single axon tracing revealed additive guidance errors, demonstrating the independent roles of Ntn1 and Slit. Moreover, the same defects were observed in embryos lacking their cognate receptors. These in vivo studies thus demonstrate the presence of multi-factorial guidance mechanisms that ensure proper formation of a common branched axonal structure during spinal cord development.

Engineering luminopsins with improved coupling efficiencies

Slaviero, A. — 2023-11-22

SignificanceLuminopsins (LMOs) are bioluminescent-optogenetic tools with a luciferase fused to an opsin that allow bimodal control of neurons by providing both optogenetic and chemogenetic access. Determining which design features contribute to the efficacy of LMOs will be beneficial for further improving LMOs for use in research. AimWe investigated the relative impact of luciferase brightness, opsin sensitivity, pairing of emission and absorption wavelength, and arrangement of moieties on the function of LMOs. ApproachWe quantified efficacy of LMOs through whole cell patch clamp recordings in HEK293 cells by determining coupling efficiency, the percentage of maximum LED induced photocurrent achieved with bioluminescent activation of an opsin. We confirmed key results by multielectrode array (MEAs) recordings in primary neurons. ResultsLuciferase brightness and opsin sensitivity had the most impact on the efficacy of LMOs, and N-terminal fusions of luciferases to opsins performed better than C-terminal and multi-terminal fusions. Precise paring of luciferase emission and opsin absorption spectra appeared to be less critical. ConclusionsWhole cell patch clamp recordings allowed us to quantify the impact of different characteristics of LMOs on their function. Our results suggest that coupling brighter bioluminescent sources to more sensitive opsins will improve LMO function. As bioluminescent activation of opsins is most likely based on Forster resonance energy transfer (FRET), the most effective strategy for improving LMOs further will be molecular evolution of luciferase-fluorescent protein-opsin fusions.

scNODE: Generative Model for Temporal Single Cell Transcriptomic Data Prediction

Zhang, J. — 2023-11-23

AO_SCPLOWBSTRACTC_SCPLOWMeasurement of single-cell gene expression at different timepoints enables the study of cell development. However, due to the resource constraints and technical challenges associated with the single-cell experiments, researchers can only profile gene expression at discrete and sparsely-sampled timepoints. This missing timepoint information impedes downstream cell developmental analyses. We propose scNODE, an end-to-end deep learning model that can predict in silico single-cell gene expression at unobserved timepoints. scNODE integrates a variational autoencoder (VAE) with neural ordinary differential equations (ODEs) to predict gene expression using a continuous and non-linear latent space. Importantly, we incorporate a dynamic regularization term to learn a latent space that is robust against distribution shifts when predicting single-cell gene expression at unobserved timepoints. Our evaluations on three real-world scRNA-seq datasets show that scNODE achieves higher predictive performance than state-of-the-art methods. We further demonstrate that scNODEs predictions help cell trajectory inference under the missing timepoint paradigm and the learned latent space is useful for in silico perturbation analysis of relevant genes along a developmental cell path. The data and code are publicly available at https://github.com/rsinghlab/scNODE.

Targeting PKC alleviates iron overload in diabetes and hemochromatosis

Banerjee, S. — 2023-11-29

Diabetes is one of the most prevalent chronic diseases worldwide. Iron overload increases the incidence of diabetes and aggravates diabetic complications that cause mortality. Reciprocally, diabetes potentially promotes body iron loading, but the mechanism remains not well understood. In this study, we demonstrated systemic iron excess and the upregulation of iron exporter ferroportin (Fpn) in the enterocytes and macrophages of multiple diabetic mouse models. Increased Fpn expression and iron efflux was also seen in the enterocytes of type 2 diabetic human patients. We further showed that protein kinase C (PKC), which is activated in hyperglycemia, was responsible for the sustained membrane expression of Fpn in physiological and in diabetic settings. For the first time, we identified that PKCs were novel binding proteins and positive regulators of Fpn. Mechanistically, hyperactive PKC promoted exocytotic membrane insertion while inhibited the endocytic trafficking of Fpn in the resting state. PKC also protected Fpn from internalization and degradation by its ligand hepcidin dependent on decreased ubiquitination and increased phosphorylation of Fpn. Importantly, the loss-of-function and pharmacological inhibition of PKC alleviated systemic iron overload in diabetes and hemochromatosis. Our study thus highlights PKC as a novel target in the control of systemic iron homeostasis.

TRAIL pathway suppression of cancer cell growth and immune cell-mediated tumor cell-killing in a senescent fibroblast-constructed tumor microenvironment

Zhang, S. — 2023-12-01

Cellular senescence and the associated secretory phenotype (SASP) promote cancer in the aging population. During aging or upon chemotherapy exposure, cellular and molecular changes occur in non-cancerous cells and alter responses to cancer therapy, primarily via modifications in the tumor microenvironment (TME) and immune response. Targeting senescent cells through removal, modulation of the SASP, or cellular reprogramming represent promising therapeutic avenues for treating cancer. We elucidate an interplay between cancer cells, immune cells, and senescent fibroblasts and describe the impact of fibroblast senescence on tumor growth and response to cancer therapy. Cytokine profiling reveals dynamic changes in SASP production during etoposide-induced senescence in IMR90 fibroblasts. We show that SASP is partially regulated by p21 (WAF1; CDKN1A), leading to the downregulation of anti-tumorigenic cytokines and upregulation of pro-tumorigenic cytokines. Senescent fibroblasts promote bystander cancer cell growth via a p21-driven SASP. These results provide strategies to target the p21-driven SASP in the TME during cancer therapy. Treatment with TRAIL or TRAIL-inducing Dordaviprone (TIC10/ONC201) reduces cell viability of tumor cells co-cultured with senescent or proliferating fibroblasts and promotes immune-mediated tumor cell-killing in co-culture with senescent IMR90 fibroblasts. ONC201 combined with senolytic drugs (e.g., Navitoclax, Lamivudine) synergizes towards tumor suppression. These results indicate that senolytic therapies may be combined with cancer therapies to target senescence-associated changes in the TME including for modulation of the senescent cytokine landscape.

Epithelial organoid supports resident memory CD8 T cell differentiation

Ulibarri, M. — 2023-12-01

Resident Memory T cells (TRM) play a vital role in regional immune defense in barrier organs. Although laboratory rodents have been extensively used to study fundamental TRM biology, poor isolation efficiency, sampling bias and low cell survival rates have limited our ability to conduct TRM-focused high-throughput assays. Here, we engineered a murine vaginal epithelial organoid (VEO)-CD8 T cell co-culture system that supports CD8 TRM differentiation in vitro. The three-dimensional VEOs established from murine adult stem cells resembled stratified squamous vaginal epithelium and induced gradual differentiation of activated CD8 T cells into epithelial TRM. These in vitro generated TRM were phenotypically and transcriptionally similar to in vivo TRM, and key tissue residency features were reinforced with a second cognate-antigen exposure during co-culture. TRM differentiation was not affected even when VEOs and CD8 T cells were separated by a semipermeable barrier, indicating soluble factors involvement. Pharmacological and genetic approaches showed that TGF-{beta} signaling played a crucial role in their differentiation. We found that the VEOs in our model remained susceptible to viral infections and the CD8 T cells were amenable to genetic manipulation; both of which will allow detailed interrogation of antiviral CD8 T cell biology in a reductionist setting. In summary, we established a robust model which captures bonafide TRM differentiation that is scalable, open to iterative sampling, and can be subjected to high throughput assays that will rapidly add to our understanding of TRM.

Single cell transcriptional changes across the blood stages of artemisinin resistant K13C580Y mutant Plasmodium falciparum upon dihydroartemisinin exposure

Oduor, C. I. — 2023-12-06

Artemisinin-based therapies have been central to malaria control, but partial resistance in Plasmodium falciparum, driven by mutations in the Kelch13 (K13) protein, threatens these gains. To investigate the molecular basis of this resistance, we applied single-cell RNA sequencing to coisogenic parasite lines, K13 wild-type (K13C580) and the artemisinin-resistant mutant (K13580Y), following a 6 hour pulse of dihydroartemisinin (DHA). This approach enabled high-resolution profiling across intraerythrocytic stages. Both lines exhibited stage-specific transcriptional responses, with pronounced changes in ring and trophozoite stages. Using Manifold Enhancement of Latent Dimensions (MELD), a computational framework for quantifying transcriptional perturbation, DHA-treatment induces stage-specific differences in protein export and metabolic pathways in K13C580 and K13580Y parasites, relating to an altered metabolic stress response state. GARP, a potential therapeutic target, was highly differentially expressed in untreated ring stages of K13580Y and K13C580. Functional assays confirmed that anti-GARP antibodies retained efficacy against K13580Y, supporting its potential as a therapeutic target. These findings provide a comprehensive view of the cellular responses related to artemisinin resistance, identify molecular features of pathogenesis, and highlight surface proteins like GARP as promising intervention targets. This work underscores the power of single-cell approaches to dissect drug responses and guide strategies to overcome resistant parasites.

Geometric-relationship specific transfer in visual perceptual learning

Tan, Q. — 2023-12-11

Visual perceptual learning (VPL) is defined as long-term improvement on a visual task as a result of visual experience. In many cases, the improvement is highly specific to the location where the target is presented, which refers to location specificity. In the current study, we investigated the effect of a geometrical relationship between the trained location and an untrained location on transfer of VPL. We found that significant transfer occurs either diagonally or along a line passing the fixation point. This indicates that whether location specificity or location transfer occurs at least partially depends on the geometrical relationship between trained location and an untrained location.

Towards a Brighter Constellation: Multi-Organ Neuroimaging of Neural and Vascular Dynamics in the Spinal Cord and Brain

Celinskis, D. — 2023-12-27

SignificancePain is comprised of a complex interaction between motor action and somatosensation that is dependent on dynamic interactions between the brain and spinal cord. This makes understanding pain particularly challenging as it involves rich interactions between many circuits (e.g., neural and vascular) and signaling cascades throughout the body. As such, experimentation on a single region may lead to an incomplete and potentially incorrect understanding of crucial underlying mechanisms. AimHere, we aimed to develop and validate new tools to enable detailed and extended observation of neural and vascular activity in the brain and spinal cord. The first key set of innovations were targeted to developing novel imaging hardware that addresses the many challenges of multi-site imaging. The second key set of innovations were targeted to enabling bioluminescent imaging, as this approach can address limitations of fluorescent microscopy including photobleaching, phototoxicity and decreased resolution due to scattering of excitation signals. ApproachWe designed 3D-printed brain and spinal cord implants to enable effective surgical implantations and optical access with wearable miniscopes or an open window (e.g., for one-or two-photon microscopy or optogenetic stimulation). We also tested the viability for bioluminescent imaging, and developed a novel modified miniscope optimized for these signals (BLmini). ResultsHere, we describe novel universal implants for acute and chronic simultaneous brain-spinal cord imaging and optical stimulation. We further describe successful imaging of bioluminescent signals in both foci, and a new miniscope, the BLmini, which has reduced weight, cost and form-factor relative to standard wearable miniscopes. ConclusionsThe combination of 3D printed implants, advanced imaging tools, and bioluminescence imaging techniques offers a new coalition of methods for understanding spinal cord-brain interactions. This work has the potential for use in future research into neuropathic pain and other sensory disorders and motor behavior.

Chitinase 3-like-1 (CHI3L1) Inhibits Innate Anti-Tumor and Tissue Remodeling ImmuneResponses by Regulating CD47-SIRPα and CD24-Siglec10-Mediated Phagocytosis

Ma, B. — 2023-12-28

Innate immune responses such as phagocytosis are critically linked to the generation of adaptive immune responses against the neoantigens in cancer and the efferocytosis that is essential for homeostasis in diseases characterized by lung injury, inflammation, and remodeling as in Chronic Obstructive Pulmonary Disease (COPD). Chitinase 3-like-1 (CHI3L1) is induced in many cancers where it inhibits adaptive immune responses by stimulating immune checkpoint molecules (ICPs) and portends a poor prognosis. CHI3L1 is also induced in COPD where it regulates epithelial cell death. Here we demonstrate that pulmonary melanoma metastasis inhibits macrophage phagocytosis by stimulating the CD47-SIRP and CD24-Siglec10 phagocytosis checkpoint pathways while inhibiting macrophage "eat me" signals from calreticulin and HMGB1. We also demonstrate that these effects on macrophage phagocytosis are mediated by CHI3L1 stimulation of the SHP-1 and SHP-2 phosphatases and the inhibition of the accumulation and phosphorylation of cytoskeleton-regulating non-muscle myosin IIa. This inhibition of innate immune responses like phagocytosis provides a mechanistic explanation for the ability of CHI3L1 to stimulate ICPs and inhibit adaptive immune responses in cancer and diseases like COPD. The ability of CHI3L1 to simultaneously inhibit innate immune responses, stimulate ICPs, inhibit T cell co-stimulation, and regulate a number of other oncogenic and inflammation pathways suggest that CHI3L1-targeted therapeutics are promising interventions in cancer, COPD and other disorders.

Overexpression of the WWE domain of RNF146 modulates poly-(ADP)-ribose dynamics at sites of DNA damage

Al-Rahahleh, R. Q. — 2023-12-29

Protein poly-ADP-ribosylation (PARylation) is a post-translational modification formed by transfer of successive units of ADP-ribose to target proteins to form poly-ADP-ribose (PAR) chains. PAR plays a critical role in the DNA damage response (DDR) by acting as a signaling platform to promote the recruitment of DNA repair factors to the sites of DNA damage that bind via their PAR-binding domains (PBDs). Several classes of PBD families have been recognized, which identify distinct parts of the PAR chain. Proteins encoding PBDs play an essential role in conveying the PAR-mediated signal through their interaction with PAR chains, which mediates many cellular functions, including the DDR. The WWE domain identifies the iso-ADP- ribose moiety of the PAR chain. We recently described the WWE domain of RNF146 as a robust genetically encoded probe, when fused to EGFP, for detection of PAR in live cells. Here, we evaluated other PBD candidates as molecular PAR probes in live cells, including several other WWE domains and an engineered macrodomain. In addition, we demonstrate unique PAR dynamics when tracked by different PAR binding domains, a finding that that can be exploited for modulation of the PAR-dependent DNA damage response.

A genetic and physiological model of renal dysfunction in Lowe syndrome

PADINJAT, R. — 2024-01-16

Lowe syndrome (LS) is an X-linked recessive genetic disorder characterized by renal dysfunction, neurodevelopmental defects, and cataract. The affected gene, OCRL encodes for a polyphosphoinositide 5-phosphatase. OCRL is localized to multiple sub-cellular locations in the endolysosomal system and defects in these organelles have been described in human cells depleted of OCRL. However, the relationship of the endolysosomal defects in OCRL depleted cells to the altered physiology of kidney cells of LS patients has not been completely determined. Here we model the kidney phenotypes of LS using a Drosophila nephrocyte model. Using this model system, we demonstrate that OCRL plays a cell-autonomous role in nephrocyte function. Deletion of the only OCRL ortholog in Drosophila (dOCRL) leads to cell-autonomous defects in larval nephrocyte structure and function. Null mutants of dOCRL (dOCRLKO) show defects in the endolysosomal system of larval nephrocytes that are associated with physiological defects in nephrocyte function. These defects could be rescued by reconstitution with a human OCRL transgene but not with a phosphatase dead version or a human LS patient derived mutation. Overall, this work provides a model system to understand the mechanisms by which the sub-cellular changes from loss of OCRL leads to defects in kidney function in human patients.

FROM DUST TO SEED: A LUNAR CHICKPEA STORY

Atkin, J. — 2024-01-22

Food sustainability is one of the most significant barriers to long-term space travel. Providing resources from Earth is not cost-efficient, and resupply missions are not viable to meet the needs of long-term life in deep space conditions. Plants in space can provide a source of nutrition and oxygen, reducing the reliance on packaged foods, reducing resupply needs, and extending the duration of missions. Using lunar regolith simulant, we employ a novel methodology to create a sustainable and productive growth medium to support the cultivation of horticultural crops on the Moon. Implementing microbial soil regeneration mechanisms derived from Earth, we leverage the interaction between Arbuscular Mycorrhizal Fungi (AMF) and Vermicompost (VC) to create a fertile LRS matrix. These amendments can sequester toxic contaminants, improve soil structure, and increase plant stress tolerance. We demonstrate the ability to produce chickpea (Cicer arietinum) in lunar regolith simulant augmented with AMF and VC under climate-controlled conditions. We cultivated chickpea to seed in a mixture containing 75% Lunar Regolith Simulant. Preliminary results suggest that higher LRS contents induce heightened stress responses. However, plants grown in 100% LRS inoculated with arbuscular mycorrhizal fungi demonstrated an average two-week survival extension compared to non-inoculated plants. This study provides, for the first time, a baseline for chickpea germination in varying mixtures of LRS and VC and will inform future studies as humanity goes back to the Moon.

First-night effect reduces the beneficial effects of sleep on visual plasticity and modifies the underlying neurochemical processes

Tamaki, M. — 2024-01-22

Individuals experience difficulty falling asleep in a new environment, termed the first night effect (FNE). However, the impact of the FNE on sleep-induced brain plasticity remains unclear. Here, using a within-subject design, we found that the FNE significantly reduces visual plasticity during sleep in young adults. Sleep-onset latency (SOL), an indicator of the FNE, was significantly longer during the first sleep session than the second session, confirming the FNE. We assessed performance gains in visual perceptual learning after sleep and increases in the excitatory-to-inhibitory neurotransmitter (E/I) ratio in early visual areas during sleep using magnetic resonance spectroscopy and polysomnography. These parameters were significantly smaller in sleep with the FNE than in sleep without the FNE; however, these parameters were not correlated with SOL. These results suggest that while the neural mechanisms of the FNE and brain plasticity are independent, sleep disturbances temporarily block the neurochemical process fundamental for brain plasticity.

Activity-Dependent Ectopic Spiking in Parvalbumin-Expressing Interneurons of the Neocortex

Theyel, B. B. — 2024-01-22

Canonically, action potentials of most mammalian neurons initiate at the axon initial segment and propagate bidirectionally: orthodromically along the distal axon, and retrogradely into the soma and dendrites. Under some circumstances action potentials may initiate ectopically, at sites distal to the axon initial segment, and propagate antidromically along the axon. These ectopic action potentials (EAPs) have been observed in experimental models of seizures and chronic pain, and more rarely in nonpathological forebrain neurons. Here we report that a large majority of parvalbumin-expressing (PV+) interneurons in upper layers of mouse neocortex, from both orbitofrontal and primary somatosensory areas, fire EAPs after sufficient activation of their somata. Somatostatin-expressing interneurons also fire EAPs, though less robustly. Ectopic firing in PV+ cells occurs in varying temporal patterns and can persist for several seconds. PV+ cells evoke strong synaptic inhibition in pyramidal neurons and interneurons and play critical roles in cortical function. Our results suggest that ectopic spiking of PV+ interneurons is common, and may contribute to both normal and pathological network functions of the neocortex. SIGNIFICANCE STATEMENTA form of neuronal firing that emerges in distal axons and terminals - the ectopic action potential (EAP) - has been detected in a few cell populations of the cerebral cortex. Previous investigations of parvalbumin-positive interneurons in neocortex had suggested only a small percentage of cells can fire EAPs. We found that a large fraction of parvalbumin-positive interneurons in the superficial layers of neocortex, including first-order and higher-order areas, can fire EAPs. These results broaden our understanding of the intrinsic firing characteristics of these critically important inhibitory interneurons.

Dissociable neural signals for reward and emotion prediction errors

Heffner, J. — 2024-01-25

Reinforcement learning models focus on reward prediction errors (PEs) as the driver of behavior. However, recent evidence indicates that deviations from emotion expectations, termed affective PEs, play a crucial role in shaping behavior. Whether there is neural separability between emotion and reward signals remains unknown. We employ electroencephalography during social learning to investigate the neural signatures of reward and affective PEs. Behavioral results reveal that while affective PEs predict choices when little is known about how a partner will behave, reward PEs become more predictive overtime as uncertainty about a partners behavior diminishes. This functional dissociation is mirrored neurally by engagement of distinct event-related potentials. The FRN indexes reward PEs while the P3b tracks affective PEs. Only the P3b predicts subsequent choices, highlighting the mechanistic influence of affective PEs during social learning. These findings present evidence for a neurobiologically viable emotion learning signal that is distinguishable--behaviorally and neurally--from reward. SignificanceFor nearly a century, scientists have asked how humans learn about their worlds. Learning models borrowed from computer science--namely, reinforcement learning--provide an elegant and simple framework that showcases how reward prediction errors are used to update ones knowledge about the environment. However, a fundamental question persists: what exactly is reward? This gap in knowledge is problematic, especially when we consider the multiplicity of social contexts where external rewards must be contextualized to gain value and meaning. We leverage electroencephalography to interrogate the role of emotion prediction errors--violations of emotional expectations--during learning. We observe distinct neural signals for reward and emotion prediction errors, suggesting that emotions may act as a bridge between external rewards and subjective value.

Computation-aided Design of Rod-Shaped Janus Base Nanopieces for Improved Tissue Penetration and Therapeutics Delivery

Lee, J. — 2024-01-27

Despite the development of various drug delivery technologies, there remains a significant need for vehicles that can improve targeting and biodistribution in "hard-to-penetrate" tissues. Some solid tumors, for example, are particularly challenging to penetrate due to their dense extracellular matrix (ECM). In this study, we have formulated a new family of rod-shaped delivery vehicles named Janus base nanopieces (Rod JBNps), which are more slender than conventional spherical nanoparticles, such as lipid nanoparticles (LNPs). These JBNp nanorods are formed by bundles of DNA-inspired Janus base nanotubes (JBNts) with intercalated delivery cargoes. To develop this novel family of delivery vehicles, we employed a computation-aided design (CAD) methodology that includes molecular dynamics and response surface methodology. This approach precisely and efficiently guides experimental designs. Using an ovarian cancer model, we demonstrated that JBNps markedly improve penetration into the dense ECM of solid tumors, leading to better treatment outcomes compared to FDA-approved spherical LNP delivery. This study not only successfully developed a rod-shaped delivery vehicle for improved tissue penetration but also established a CAD methodology to effectively guide material design.

Multiple plastid losses within photosynthetic stramenopiles revealed by comprehensive phylogenomics

Terpis, K. — 2024-02-07

Ochrophyta is a vast and morphologically diverse group of algae with complex plastids, including familiar taxa with fundamental ecological importance (diatoms or kelp), and a wealth of lesser-known and obscure organisms. The sheer diversity of ochrophytes poses a challenge for reconstructing their phylogeny, with major gaps in sampling and an unsettled placement of particular taxa yet to be tackled. We sequenced transcriptomes from 25 strategically selected representatives and used these data to build the most taxonomically comprehensive ochrophyte-centered phylogenomic supermatrix to date. We employed a combination of approaches to reconstruct and critically evaluate the relationships among ochrophytes. While generally congruent with previous analyses, the updated ochrophyte phylogenomic tree resolved the position of several taxa with previously uncertain placement, and supported a redefinition of the class Synchromophyceae. Our results indicated that the heterotrophic plastid-lacking heliozoan Actinophrys sol is not a sister lineage of ochrophytes, as proposed recently, but rather phylogenetically nested among them. In addition, we found Picophagus flagellatus to be a secondarily heterotrophic ochrophyte lacking all hallmark plastid genes, yet exhibiting mitochondrial proteins that seem to be genetic footprints of lost plastid organelle. We thus document, for the first time, plastid loss in two separate ochrophyte lineages. Altogether, our study provides a new framework for reconstructing trait evolution in ochrophytes and demonstrates that plastid loss is more common than previously thought. Issue Section: Discoveries

scGrapHiC: Deep learning-based graph deconvolution for Hi-C using single cell gene expression

Murtaza, G. — 2024-02-10

Single-cell Hi-C (scHi-C) protocol helps identify cell-type-specific chromatin interactions and sheds light on cell differentiation and disease progression. Despite providing crucial insights, scHi-C data is often underutilized due the high cost and the complexity of the experimental protocol. We present a deep learning framework, scGrapHiC, that predicts pseudo-bulk scHi-C contact maps using pseudo-bulk scRNA-seq data. Specifically, scGrapHiC performs graph deconvolution to extract genome-wide single-cell interactions from a bulk Hi-C contact map using scRNA-seq as a guiding signal. Our evaluations show that scGrapHiC, trained on 7 cell-type co-assay datasets, outperforms typical sequence encoder approaches. For example, scGrapHiC achieves a substantial improvement of 23.2% in recovering cell-type-specific Topologically Associating Domains over the baselines. It also generalizes to unseen embryo and brain tissue samples. scGrapHiC is a novel method to generate cell-type-specific scHi-C contact maps using widely available genomic signals that enables the study of cell-type-specific chromatin interactions. Availabilityhttps://github.com/rsinghlab/scGrapHiC Contactritambhara@brown.edu

Scalable nonparametric clustering with unified marker gene selection for single-cell RNA-seq data

Nwizu, C. — 2024-02-12

Clustering is commonly used in single-cell RNA-sequencing (scRNA-seq) pipelines to characterize cellular heterogeneity. However, current methods face two main limitations. First, they require user-specified heuristics which add time and complexity to bioinformatic workflows; second, they rely on post-selective differential expression analyses to identify marker genes driving cluster differences, which has been shown to be subject to inflated false discovery rates. We address these challenges by introducing nonparametric clustering of single-cell populations (NCLUSION): an infinite mixture model that leverages Bayesian sparse priors to identify marker genes while simultaneously performing clustering on single-cell expression data. NCLUSION uses a scalable variational inference algorithm to perform these analyses on datasets with up to millions of cells. Through simulations and analyses of publicly available scRNA-seq studies, we demonstrate that NCLUSION (i) matches the performance of other state-of-the-art clustering techniques with significantly reduced runtime and (ii) provides statistically robust and biologically relevant transcriptomic signatures for each of the clusters it identifies. Overall, NCLUSION represents a reliable hypothesis-generating tool for understanding patterns of expression variation present in single-cell populations.

Striatal Dopamine Can Enhance Learning, Both Fast and Slow, and Also Make it Cheaper

Westbrook, A. — 2024-02-17

Associations can be learned incrementally, via reinforcement learning (RL), or stored instantly in working memory (WM). While WM is fast, it is also capacity-limited and effortful. Striatal dopamine may promote RL plasticity, and WM, by facilitating updating and effort exertion. Yet, prior studies have failed to distinguish between dopamines effects on RL versus WM. N = 100 participants completed a paradigm isolating these systems in a double-blind study measuring dopamine synthesis with [18F]-FDOPA imaging and manipulating dopamine with methylphenidate and sulpiride. Learning is enhanced among high synthesis capacity individuals and by methylphenidate, but impaired by sulpiride. Methylphenidate also blunts effort cost learning. Computational modeling reveals that individuals with high dopamine synthesis rely more on WM, while methylphenidate boosts their RL rates. The D2 antagonist sulpiride reduces accuracy due to diminished WM involvement and faster WM decay. We conclude that dopamine enhances both slow RL, and fast WM, by promoting plasticity and reducing effort sensitivity. These results also highlight the need to control for dopamines effects on WM when testing its effects on RL.

Vaginal lactobacilli produce anti-inflammatory beta-carboline compounds.

Webber, C. A. — 2024-02-19

The vaginal microbiome is a lactobacilli-dominant, human-adapted community consistently found in people around the world. Presence of lactobacilli-dominant vaginal microbial communities, apart from Lactobacillus iners, is associated with reduced vaginal inflammation and reduced levels of pro-inflammatory cytokines. Loss of lactobacilli-dominance is associated with inflammatory conditions such as bacterial vaginosis. It remains unclear if health-associated lactobacilli actively promote the anti-inflammatory immune homeostasis of the vaginal mucosa and if so, by which mechanisms. We have identified that Lactobacillus crispatus, a key vaginal bacterial species, secretes a family of {beta}-carboline compounds with anti-inflammatory activity. These compounds suppress NF{kappa}B and interferon signaling downstream of multiple pattern recognition receptors in human macrophages and significantly dampen type I interferon receptor activation. Topical vaginal application of an anti-inflammatory {beta}-carboline compound, perlolyrine, was sufficient to significantly reduce vaginal inflammation in a mouse model of genital herpes infection. Together, we identify a key family of compounds by which vaginal lactobacilli mediate host immune homeostasis and could highlight a new therapeutic avenue for vaginal inflammation.

Monkeys engage in visual simulation to solve complex problems

Ahuja, A. — 2024-02-23

Visual simulation -- i.e., using internal reconstructions of the world to experience potential future versions of events that are not currently happening -- is among the most sophisticated capacities of the human mind. But is this ability in fact uniquely human? To answer this question, we tested monkeys on a series of experiments involving the Planko game, which we have previously used to evoke visual simulation in human participants. We found that monkeys were able to successfully play the game using a simulation strategy, predicting the trajectory of a ball through a field of planks while demonstrating a level of accuracy and behavioral signatures comparable to humans. Computational analyses further revealed that the monkeys strategy while playing Planko aligned with a recurrent neural network (RNN) that approached the task using a spontaneously learned simulation strategy. Finally, we carried out awake functional magnetic resonance imaging while monkeys played Planko. We found activity in motion-sensitive regions of the monkey brain during hypothesized simulation periods, even without any perceived visual motion cues. This neural result closely mirrors previous findings from human research, suggesting a shared mechanism of visual simulation across species. In all, these findings challenge traditional views of animal cognition, proposing that nonhuman primates possess a complex cognitive landscape, capable of invoking imaginative and predictive mental experiences to solve complex everyday problems.

Red blood cell passage through deformable interendothelial slits in the spleen: Insights into splenic filtration and hemodynamics

Li, G. — 2024-02-23

The spleen constantly clears altered red blood cells (RBCs) from the circulation, tuning the balance between RBC formation (erythropoiesis) and removal. The retention and elimination of RBCs occur predominantly in the open circulation of the spleen, where RBCs must cross submicron-wide inter-endothelial slits (IES). Several experimental and computational studies have illustrated the role of IES in filtrating the biomechanically and morphologically altered RBCs based on a rigid wall assumption. However, these studies also reported that when the size of IES is close to the lower end of clinically observed sizes (less than 0.5 m), an unphysiologically large pressure difference across the IES is required to drive the passage of normal RBCs, sparking debates on the feasibility of the rigid wall assumption. In this work, we perform a computational investigation based on dissipative particle dynamics (DPD) to explore the impact of the deformability of IES on the filtration function of the spleen. We simulate two deformable IES models, namely the passive model and the active model. In the passive model, we implement the worm-like string model to depict the IESs deformation as it interacts with blood plasma and allows RBC to traverse. In contrast, the active model involved regulating the IES deformation based on the local pressure surrounding the slit. To demonstrate the validity of the deformable model, we simulate the filtration of RBCs with varied size and stiffness by IES under three scenarios: 1) a single RBC traversing a single slit; 2) a suspension of RBCs traversing an array of slits, mimicking in vitro spleen-on-a-chip experiments; 3) RBC suspension passing through the 3D spleen filtration unit known as the splenon. Our simulation results of RBC passing through a single slit show that the deformable IES model offers more accurate predictions of the critical cell surface area to volume ratio that dictate the removal of aged RBCs from circulation compared to prior rigid-wall models. Our biophysical models of the spleen-on-a-chip indicates a hierarchy of filtration function stringency: rigid model > passive model > active model, providing a possible explanation of why the spleen-on-a-chip could overestimate the filtration function of IES. We also illustrate that the biophysical model of the splenon enables us to replicate the ex vivo experiments involving spleen filtration of malaria-infected RBCs. Taken together, our simulation findings indicate that the deformable IES model could serve as a mesoscopic representation of spleen filtration function closer to physiological reality, addressing questions beyond the scope of current experimental and computational models and enhancing our understanding of the fundamental flow dynamics and mechanical clearance processes within in the human spleen.

A worldwide ENIGMA study on epilepsy-related gray and white matter compromise across the adult lifespan

Chen, J. — 2024-03-06

ObjectivesTemporal lobe epilepsy (TLE) is commonly associated with mesiotemporal pathology and widespread alterations of grey and white matter structures. Evidence supports a progressive condition although the temporal evolution of TLE is poorly defined. This ENIGMA-Epilepsy study utilized multimodal magnetic resonance imaging (MRI) data to investigate structural alterations in TLE patients across the adult lifespan. We charted both grey and white matter changes and explored the covariance of age-related alterations in both compartments. MethodsWe studied 769 TLE patients and 885 healthy controls across an age range of 17-73 years, from multiple international sites. To assess potentially non-linear lifespan changes in TLE, we harmonized data and combined median split assessments with cross-sectional sliding window analyses of grey and white matter age-related changes. Covariance analyses examined the coupling of grey and white matter lifespan curves. ResultsIn TLE, age was associated with a robust grey matter thickness/volume decline across a broad cortico-subcortical territory, extending beyond the mesiotemporal disease epicentre. White matter changes were also widespread across multiple tracts with peak effects in temporo-limbic fibers. While changes spanned the adult time window, changes accelerated in cortical thickness, subcortical volume, and fractional anisotropy (all decreased), and mean diffusivity (increased) after age 55 years. Covariance analyses revealed strong limbic associations between white matter tracts and subcortical structures with cortical regions. ConclusionsThis study highlights the profound impact of TLE on lifespan changes in grey and white matter structures, with an acceleration of aging-related processes in later decades of life. Our findings motivate future longitudinal studies across the lifespan and emphasize the importance of prompt diagnosis as well as intervention in patients.

Reduction in activity and abundance of mitochondrial electron transport chain supercomplexes in pulmonary hypertension-induced right ventricular dysfunction

Ma, W. — 2024-03-13

Pulmonary hypertension (PH) results in RV hypertrophy, fibrosis and dysfunction resulting in RV failure which is associated with impaired RV metabolism and mitochondrial respiration. Mitochondrial supercomplexes (mSC) are assemblies of multiple electron transport chain (ETC) complexes that consist of physically associated complex I, III and IV that may enhance respiration and lower ROS generation. The goal of this study was to determine if mSCs are reduced in RV dysfunction associated with PH. We induced PH in Sprague-Dawley rats by Sugen/Hypoxia (3 weeks) followed by normoxia (4 weeks). Control and PH rats were subjected to echocardiography, blue and clear native-PAGE to assess mSC abundance and activity, and cardiomyocyte isolation to assess mitochondrial reactive oxygen species (ROS). mSC formation was also assessed in explanted human hearts with and without RV dysfunction. RV activity of CI and CIV and abundance of CI, CIII and CIV in mitochondrial mSCs was severely reduced in PH rats compared to control. There were no differences in total CI or CIV activity or abundance in smaller ETC assemblies. There were no changes in both RV and LV of expression of representative ETC complex subunits. PAT, TAPSE and RV Wall thickness significantly correlated with CIV and CI activity in mSC, but not total CI and CIV activity in the RV. Consistent with reduced mSC activity, isolated PH RV myocytes had increased mitochondrial ROS generation compared to control. Reduced mSC activity was also demonstrated in explanted human RV tissue from patients undergoing cardiac transplant with RV dysfunction. The right atrial pressure/pulmonary capillary wedge pressure ratio (RAP/PCWP, an indicator of RV dysfunction) negatively correlated with RV mSC activity level. In conclusion, reduced assembly and activity of mitochondrial mSC is correlated with RV dysfunction in PH rats and humans with RV dysfunction.

A knockoff calibration method to avoid over-clustering in single-cell RNA-sequencing

DenAdel, A. — 2024-03-13

Standard single-cell RNA-sequencing (scRNA-seq) pipelines nearly always include unsupervised clustering as a key step in identifying biologically distinct cell types. A follow-up step in these pipelines is to test for differential expression between the identified clusters. When algorithms over-cluster, downstream analyses will produce inflated P -values resulting in increased false discoveries. In this work, we present callback (Calibrated Clustering via Knockoffs): a new method for protecting against over-clustering by controlling for the impact of reusing the same data twice when performing differential expression analysis, commonly known as "double-dipping". Importantly, our approach can be applied to a wide range of clustering algorithms. Using real and simulated data, we show that callback provides state-of-the-art clustering performance and can rapidly analyze large-scale scRNA-seq studies, even on a personal laptop.

Heterochromatin spreading in cancer cells through HDAC7 mediated histone H3.3 landscape reprogramming.

Hassan, O. — 2024-03-14

Class IIa histone deacetylases (HDACs) are a family of enzymes with minimal histone deacetylase activity but can function as multi-protein interaction hubs. Here we demonstrate the expression of HDAC7, a Class IIa HDAC family member, in glioblastoma tumor tissue from 84 patients, patient-derived glioma stem cells (GSCs) from six patients, and pediatric diffuse pontine glioma (DIPG) cells from three patients. HDAC7 binds to Histone H3.3 and interacts with H3.3 and HIRA on chromatin. Targeted downregulation of HDAC7 expression with a subtype-specific siRNA inhibits the interaction of H3.3 with HIRA while increasing the association of H3.3 with DAXX and H3K9me3. This results in H3.3 being deposited on H3K9me3+/DAPI+ heterochromatin nuclear foci. Inhibition of HDAC7 triggers H3K9me3+ heterochromatin spreading, increased H3K9me3 binding in the cancer genome, and significant alterations in gene expression. Using single molecule DNA fiber approach, we show that HDAC7 inhibition results in a significant increase in replication fork speed without affecting fork symmetry. This altered replication fork speed leads to replication stress, evidenced by phosphorylation of RPA2 and impact on global DNA synthesis, resulting in reduced EdU incorporation. Finally, HDAC7 depletion leads to reduced BRCA2 expression and increased sensitivity of cancer cells to DNA damaging agents. Taken together, these studies uncover the involvement of HDAC7 in the euchromatic H3.3 chaperone network and the effect of HDAC7 depletion on chromatin dynamics, inducing epigenetic restriction and DNA damage in cancer cells.

Adaptive chunking improves effective working memory capacity in a prefrontal cortex and basal ganglia circuit

Soni, A. — 2024-03-27

How and why is working memory (WM) capacity limited? Traditional cognitive accounts focus either on limitations on the number or items that can be stored (slots models), or loss of precision with increasing load (resource models). Here we show that a neural network model of prefrontal cortex and basal ganglia can learn to reuse the same prefrontal populations to store multiple items, leading to resource-like constraints within a slot-like system, and inducing a trade-off between quantity and precision of information. Such "chunking" strategies are adapted as a function of reinforcement learning and WM task demands, mimicking human performance and normative models. Moreover, adaptive performance requires a dynamic range of dopaminergic signals to adjust striatal gating policies, providing a new interpretation of WM difficulties in patient populations such as Parkinsons disease, ADHD and schizophrenia. These simulations also suggest a computational rather than anatomical limit to WM capacity.

Balancing the risks of mating: biogeographic evidence of cleistogamy as a bet hedging strategy

Weissman, M. — 2024-04-01

Cleistogamy is a mating system in which plants produce some proportion of closed, autonomously self-pollinating flowers. Cleistogamous flowers differ from chasmogamous flowers, which are open flowers capable of outcrossing. Both dimorphic cleistogamy (cleistogamous and chasmogamous flowers produced on the same plant) and complete cleistogamy occur. Cleistogamy has been hypothesized to be a bet hedging strategy for reducing risk in the face of unpredictable pollinator availability. However, conflicting results across species and challenges connecting theory to data have prevented researchers from proving that cleistogamy is bet hedging. To test the bet hedging hypothesis, we investigated the distribution of over 400 cleistogamous species through biogeographical analyses. We find that cleistogamy is more prevalent in cooler, more variable environments. Additionally, we find that among cleistogamous species, complete cleistogamy is more likely to occur in warmer, more stable, tropical and subtropical environments. We hypothesize that the difference in distribution between complete and dimorphic cleistogamy may be driven by the opposing forces of selection to increase cleistogamy proportion and extinction risk, which we test using a heuristic Markov transition model. We conclude that the distribution of cleistogamy suggests that the strategy has evolved in variable environments, consistent with expectations for bet hedging.

TRIP12 governs DNA Polymerase β involvement in DNA damage response and repair

Inanc, B. — 2024-04-10

The multitude of DNA lesion types, and the nuclear dynamic context in which they occur, present a challenge for genome integrity maintenance as this requires the engagement of different DNA repair pathways. Specific repair controllers that facilitate DNA repair pathway crosstalk between double strand break (DSB) repair and base excision repair (BER), and regulate BER protein trafficking at lesion sites, have yet to be identified. We find that DNA polymerase {beta} (Pol{beta}), crucial for BER, is ubiquitylated in a BER complex-dependent manner by TRIP12, an E3 ligase that partners with UBR5 and restrains DSB repair signaling. Here we find that, TRIP12, but not UBR5, controls cellular levels and chromatin loading of Pol{beta}. Required for Pol{beta} foci formation, TRIP12 regulates Pol{beta} involvement after DNA damage. Notably, excessive TRIP12-mediated shuttling of Pol{beta} affects DSB formation and radiation sensitivity, underscoring its precedence for BER. We conclude that the herein discovered trafficking function at the nexus of DNA repair signaling pathways, towards Pol{beta}-directed BER, optimizes DNA repair pathway choice at complex lesion sites. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/588474v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@70c5bdorg.highwire.dtl.DTLVardef@1c60dfborg.highwire.dtl.DTLVardef@1c00ef6org.highwire.dtl.DTLVardef@1677f2_HPS_FORMAT_FIGEXP M_FIG C_FIG

SARS-CoV-2 spike S2 subunit inhibits p53 activation of p21(WAF1), TRAIL Death Receptor DR5 and MDM2 proteins in cancer cells

Zhang, S. — 2024-04-15

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and COVID-19 infection has led to worsened outcomes for patients with cancer. SARS-CoV-2 spike protein mediates host cell infection and cell-cell fusion that causes stabilization of tumor suppressor p53 protein. In-silico analysis previously suggested that SARS-CoV-2 spike interacts with p53 directly but this putative interaction has not been demonstrated in cells. We examined the interaction between SARS-CoV-2 spike, p53 and MDM2 (E3 ligase, which mediates p53 degradation) in cancer cells using an immunoprecipitation assay. We observed that SARS-CoV-2 spike protein interrupts p53-MDM2 protein interaction but did not detect SARS-CoV-2 spike bound with p53 protein in the cancer cells. We further observed that SARS-CoV-2 spike suppresses p53 transcriptional activity in cancer cells including after nutlin exposure of wild-type p53-, spike S2-expressing tumor cells and inhibits chemotherapy-induced p53 gene activation of p21(WAF1), TRAIL Death Receptor DR5 and MDM2. The suppressive effect of SARS-CoV-2 spike on p53-dependent gene activation provides a potential molecular mechanism by which SARS-CoV-2 infection may impact tumorigenesis, tumor progression and chemotherapy sensitivity. In fact, cisplatin-treated tumor cells expressing spike S2 were found to have increased cell viability as compared to control cells. Further observations on {gamma}-H2AX expression in spike S2-expressing cells treated with cisplatin may indicate altered DNA damage sensing in the DNA damage response pathway. The preliminary observations reported here warrant further studies to unravel the impact of SARS-CoV-2 and its various encoded proteins including spike on pathways of tumorigenesis and response to cancer therapeutics.

T follicular helper cell profiles differ by malaria antigen and for children compared to adults

Forconi, C. S. — 2024-04-16

BackgroundCirculating T-follicular helper (cTFH) cells have the potential to provide an additional correlate of protection against Plasmodium falciparum (Pf) as they are essential to promote B-cell production of long-lasting antibodies. Assessing the specificity of cTFH subsets to individual malaria antigens is vital to understanding the variation observed in antibody responses and identifying promising malaria vaccine candidates. MethodsUsing spectral flow cytometry and unbiased clustering analysis, we assessed antigen-specific cTFH cell recall responses in vitro to malaria vaccine candidates Pf-schizont egress antigen-1 (PfSEA-1A) and Pf-glutamic acid-rich protein (PfGARP) within a cross-section of children and adults living in a malaria-holoendemic region of western Kenya. FindingsIn children, a broad array of cTFH subsets (defined by cytokine and transcription factor expression) were reactive to both malaria antigens, PfSEA-1A and PfGARP, while adults had a narrow profile centering on cTFH17- and cTFH1/17-like subsets following stimulation with PfGARP only. InterpretationBecause TFH17 cells are involved in the maintenance of memory antibody responses within the context of parasitic infections, our results suggest that PfGARP might generate longer-lived antibody responses compared to PfSEA-1A. These findings have intriguing implications for evaluating malaria vaccine candidates as they highlight the importance of including cTFH profiles when assessing interdependent correlates of protective immunity.

Loss of developmentally derived Irf8+ macrophages promotes hyperinnervation and arrhythmia in the adult zebrafish heart

Paquette, S. E. — 2024-04-20

Recent developments in cardiac macrophage biology have broadened our understanding of the critical functions of macrophages in the heart. As a result, there is further interest in understanding the independent contributions of distinct subsets of macrophage to cardiac development and function. Here, we demonstrate that genetic loss of interferon regulatory factor 8 (Irf8)-positive embryonic-derived macrophages significantly disrupts cardiac conduction, chamber function, and innervation in adult zebrafish. At 4 months post-fertilization (mpf), homozygous irf8st96/st96 mutants have significantly shortened atrial action potential duration and significant differential expression of genes involved in cardiac contraction. Functional in vivo assessments via electro- and echocardiograms at 12 mpf reveal that irf8 mutants are arrhythmogenic and exhibit diastolic dysfunction and ventricular stiffening. To identify the molecular drivers of the functional disturbances in irf8 null zebrafish, we perform single cell RNA sequencing and immunohistochemistry, which reveal increased leukocyte infiltration, epicardial activation, mesenchymal gene expression, and fibrosis. Irf8 null hearts are also hyperinnervated and have aberrant axonal patterning, a phenotype not previously assessed in the context of cardiac macrophage loss. Gene ontology analysis supports a novel role for activated epicardial-derived cells (EPDCs) in promoting neurogenesis and neuronal remodeling in vivo. Together, these data uncover significant cardiac abnormalities following embryonic macrophage loss and expand our knowledge of critical macrophage functions in heart physiology and governing homeostatic heart health.

Complementary benefits of multivariate and hierarchical models for identifying individual differences in cognitive control

Freund, M. C. — 2024-04-28

Understanding individual differences in cognitive control is a central goal in psychology and neuroscience. Reliably measuring these differences, however, has proven extremely challenging, at least when using standard measures in cognitive neuroscience such as response times or task-based fMRI activity. While prior work has pinpointed the source of the issue -- the vast amount of cross-trial variability within these measures -- solutions remain elusive. Here, we propose one potential way forward: an analytic framework that combines hierarchical Bayesian modeling with multivariate decoding of trial-level fMRI data. Using this framework and longitudinal data from the Dual Mechanisms of Cognitive Control project, we estimated individuals neural responses associated with cognitive control within a color-word Stroop task, then assessed the reliability of these individuals responses across a time interval of several months. We show that in many prefrontal and parietal brain regions, test-retest reliability was near maximal, and that only hierarchical models were able to reveal this state of affairs. Further, when compared to traditional univariate contrasts, multivariate decoding enabled individual-level correlations to be estimated with significantly greater precision. We specifically link these improvements in precision to the optimized suppression of cross-trial variability in decoding. Together, these findings not only indicate that cognitive control-related neural responses individuate people in a highly stable manner across time, but also suggest that integrating hierarchical and multivariate models provides a powerful approach for investigating individual differences in cognitive control, one that can effectively address the issue of high-variability measures.

Phasing millions of samples achieves near perfect accuracy, enabling parent-of-origin classification of variants

Williams, C. M. — 2024-05-10

Haplotype phasing, the process of determining which genetic variants are physically located on the same chromosome, is crucial for various genetic analyses. In this study, we first benchmark SHAPEIT and Beagle, two state-of-the-art phasing methods, on two large datasets: > 8 million diverse, research-consented 23andMe, Inc. customers and the UK Biobank (UKB). We find that both perform exceptionally well. Beagles median switch error rate (SER) (after excluding single SNP switches) in white British trios from UKB is 0.026% compared to 0.00% for European ancestry 23andMe research participants; 55.6% of European ancestry 23andMe research participants have zero non-single SNP switches, compared to 42.4% of white British trios. South Asian ancestry 23andMe research participants have the highest median SER amongst the 23andMe populations, but it is still remarkably low at 0.46%. We also investigate the relationship between identity-by-descent (IBD) and SER, finding that switch errors tend to occur in regions of little or no IBD segment coverage. SHAPEIT and Beagle excel at intra-chromosomal phasing, but lack the ability to phase across chromosomes, motivating us to develop an inter-chromosomal phasing method, called HAPTIC (HAPlotype TIling and Clustering), that assigns paternal and maternal variants discretely genome-wide. Our approach uses identity-by-descent (IBD) segments to phase blocks of variants on different chromosomes. HAPTIC represents the segments a focal individual shares with their relatives as nodes in a signed graph and performs bipartite clustering on the signed graph using spectral clustering. We test HAPTIC on 1022 UKB trios, yielding a median phase error of 0.08% in regions covered by IBD segments (33.5% of sites). We also ran HAPTIC in the 23andMe database and found a median phase error rate (the rate of mismatching alleles between the inferred and true phase) of 0.92% in Europeans (93.8% of sites) and 0.09% in admixed Africans (92.7% of sites). HAPTICs precision depends heavily on data from relatives, so will increase as datasets grow larger and more diverse. HAPTIC enables analyses that require the parent-of-origin of variants, such as association studies and ancestry inference of untyped parents.

Naturalistic acute pain states decoded from neural and facial dynamics

Huang, Y. — 2024-05-12

Pain is a complex experience that remains largely unexplored in naturalistic contexts, hindering our understanding of its neurobehavioral representation in ecologically valid settings. To address this, we employed a multimodal, data-driven approach integrating intracranial electroencephalography, pain self-reports, and facial expression quantification to characterize the neural and behavioral correlates of naturalistic acute pain in twelve epilepsy patients undergoing continuous monitoring with neural and audiovisual recordings. High self-reported pain states were associated with elevated blood pressure, increased pain medication use, and distinct facial muscle activations. Using machine learning, we successfully decoded individual participants high versus low self-reported pain states from distributed neural activity patterns (mean AUC = 0.70), involving mesolimbic regions, striatum, and temporoparietal cortex. High self-reported pain states exhibited increased low-frequency activity in temporoparietal areas and decreased high-frequency activity in mesolimbic regions (hippocampus, cingulate, and orbitofrontal cortex) compared to low pain states. This neural pain representation remained stable for hours and was modulated by pain onset and relief. Objective facial expression changes also classified self-reported pain states, with results concordant with electrophysiological predictions. Importantly, we identified transient periods of momentary pain as a distinct naturalistic acute pain measure, which could be reliably differentiated from affect-neutral periods using intracranial and facial features, albeit with neural and facial patterns distinct from self-reported pain. These findings reveal reliable neurobehavioral markers of naturalistic acute pain across contexts and timescales, underscoring the potential for developing personalized pain interventions in real-world settings.

Cone bipolar cell synapses generate transient versus sustained signals in parallel ON pathways of the mouse retina

Kuo, S. P. — 2024-05-14

Parallel processing is a fundamental organizing principle in the nervous system and understanding how parallel neural circuits generate distinct outputs from common inputs is a key goal of neuroscience. In the mammalian retina, divergence of cone signals into multiple feedforward bipolar cell pathways forms the initial basis for parallel retinal circuits dedicated to specific visual functions. Here, we used patch-clamp electrophysiology, electron microscopy and two photon imaging of a fluorescent glutamate sensor to examine how kinetically-distinct responses arise in transient versus sustained ON alpha RGCs (ON-T and ON-S RGCs) of the mouse retina. We directly compared the visual response properties of these RGCs with their presynaptic bipolar cell partners, which we identified using 3D electron microscopy reconstruction. Different ON bipolar cell subtypes (type 5i, type 6 and type 7) had indistinguishable light-driven responses whereas extracellular glutamate signals around RGC dendrites and postsynaptic excitatory currents measured in ON-T and ON-S RGCs in response to the identical stimuli used to probe bipolar cells were kinetically distinct. Anatomical examination of the bipolar cell axon terminals presynaptic to ON-T and ON-S RGCs suggests that bipolar subtype-specific differences in the size of synaptic ribbon-associated vesicle pools may contribute to transient versus sustained kinetics. Our findings indicate that feedforward bipolar cell synapses are a primary point of divergence in kinetically distinct visual pathways.

Patterned Electrical Brain Stimulation by a Wireless Network of Implantable Microdevices

Lee, J. — 2024-05-26

Transmitting meaningful information into brain circuits by electronic means is a challenge facing brain-computer interfaces. A key goal is to find an approach to inject spatially structured local current stimuli across swaths of sensory areas of the cortex. Here, we introduce a fully wireless approach to multipoint patterned electrical microstimulation by a spatially distributed epicortically implanted network of silicon microchips to target specific areas of the cortex. Each sub-millimeter-sized microchip harvests energy from an external radio-frequency source and converts this into biphasic current injected focally into tissue by a pair of integrated microwires. The amplitude, period, and repetition rate of injected current from each chip are controlled across the implant network by implementing a pre-scheduled, collision-free bitmap wireless communication protocol featuring sub-millisecond latency. As an in vivo demonstration, a network of 30 wireless stimulators was chronically implanted into motor and sensory areas of the cortex in a freely moving rat for three months. We explored the effects of patterned intracortical electrical stimulation on trained animal behavior at average RF powers well below safety limits.

Perirhinal and postrhinal cortices are necessary for retrieving latently-acquired stimulus associations

Peng, X. — 2024-05-26

The perirhinal and postrhinal cortices reside in the parahippocampal region of the medial temporal lobe. They receive and process sensory information from cortical and subcortical sources and support hippocampal functions via direct connectivity and indirectly via the entorhinal cortex. Previous studies using the sensory preconditioning paradigm have shown that the perirhinal and postrhinal cortices are necessary for associating cues during preconditioning and retrieving the associations during conditioning. However, the question of whether these regions are also required for stimulus association retrieval during re-exposure to the preconditioning cue had not been addressed. Using a chemogenetic approach, we temporarily suppressed the perirhinal or postrhinal cortex in adult male rats during the preconditioning cue test phase. Both suppression groups showed impaired sensory preconditioning compared to sham surgery controls, as indicated by significantly reduced preferential responding. Implications of our findings are discussed in relation to the importance of the PER and POR in context processing and episodic-like memory in animal models. Significance statementAlthough Pavlovian conditioning between two stimuli is widely used in neuroscience, associative learning in the real-world is often too complex to be modeled by first-order conditioning alone. Proper responding may require integrating multiple associations via common elements, i.e. higher-order conditioning, the neural basis of which is not well understood. Whereas existing research indicates the perirhinal and postrhinal cortices in the medial temporal lobe contribute to certain forms of sensory preconditioning (SPC, a type of higher-order conditioning), we suggest their contributions lie in the encoding and retrieving latently acquired associations, processes which are central to all forms of SPC. We argue this interpretation also captures perirhinal and postrhinal contributions to contextual and episodic memory, hence offering a unifying explanation regarding the contributions of these cortices across multiple paradigms.

Right ventricular stiffening and anisotropy alterations in pulmonary hypertension: Mechanisms and relations to function

Neelakantan, S. — 2024-05-29

AimsPulmonary hypertension (PH) results in an increase in RV afterload, leading to RV dysfunction and failure. The mechanisms underlying maladaptive RV remodeling are poorly understood. In this study, we investigated the multiscale and mechanistic nature of RV free wall (RVFW) biomechanical remodeling and its correlations with RV function adaptations. Methods and ResultsMild and severe models of PH, consisting of hypoxia (Hx) model in Sprague-Dawley (SD) rats (n=6 each, Control and PH) and Sugen-hypoxia (SuHx) model in Fischer (CDF) rats (n=6 each, Control and PH), were used. Organ-level function and tissue-level stiffness and microstructure were quantified through in-vivo and ex-vivo measures, respectively. Multiscale analysis was used to determine the association between fiber-level remodeling, tissue-level stiffening, and organ-level dysfunction. Animal models with different PH severity provided a wide range of RVFW stiffening and anisotropy alterations in PH. Decreased RV-pulmonary artery (PA) coupling correlated strongly with stiffening but showed a weaker association with the loss of RVFW anisotropy. Machine learning classification identified the range of adaptive and maladaptive RVFW stiffening. Multiscale modeling revealed that increased collagen fiber tautness was a key remodeling mechanism that differentiated severe from mild stiffening. Myofiber orientation analysis indicated a shift away from the predominantly circumferential fibers observed in healthy RVFW specimens, leading to a significant loss of tissue anisotropy. ConclusionMultiscale biomechanical analysis indicated that although hypertrophy and fibrosis occur in both mild and severe PH, certain fiber-level remodeling events, including increased tautness in the newly deposited collagen fibers and significant reorientations of myofibers, contributed to excessive biomechanical maladaptation of the RVFW leading to severe RV-PA uncoupling. Collagen fiber remodeling and the loss of tissue anisotropy can provide an improved understanding of the transition from adaptive to maladaptive remodeling. Translational perspectiveRight ventricular (RV) failure is a leading cause of mortality in patients with pulmonary hypertension (PH). RV diastolic and systolic impairments are evident in PH patients. Stiffening of the RV wall tissue and changes in the wall anisotropy are expected to be major contributors to both impairments. Global assessments of the RV function remain inadequate in identifying patients with maladaptive RV wall remodeling primarily due to their confounded and weak representation of RV fiber and tissue remodeling events. This study provides novel insights into the underlying mechanisms of RV biomechanical remodeling and identifies the adaptive-to-maladaptive transition across the RV biomechanics-function spectrum. Our analysis dissecting the contribution of different RV wall remodeling events to RV dysfunction determines the most adverse fiber-level remodeling to RV dysfunction as new therapeutic targets to curtail RV maladaptation and, in turn, RV failure in PH.

An active electronic bidirectional interface for high resolution interrogation of the spinal cord

Parker, S. R. — 2024-06-02

ObjectiveEpidural electrical stimulation (EES) has shown promise as both a clinical therapy and research tool for studying nervous system function. However, available clinical EES paddles are limited to using a small number of contacts due to the burden of wires necessary to connect each contact to the therapeutic delivery device, limiting the treatment area or density of epidural electrode arrays. We aimed to eliminate this burden using advanced on-paddle electronics. ApproachWe developed a smart EES paddle with a 60-electrode programmable array, addressable using an active electronic multiplexer embedded within the electrode paddle body. The electronics are sealed in novel, ultra-low profile hermetic packaging. We conducted extensive reliability testing on the novel array, including a battery of ISO 10993-1 biocompatibility tests and determination of the hermetic package leak rate. We then evaluated the EES device in vivo, placed on the epidural surface of the ovine lumbosacral spinal cord for 15 months. Main resultsThe active paddle array performed nominally when implanted in sheep for over 15 months and no device-related malfunctions were observed. The onboard multiplexer enabled bespoke electrode arrangements across, and within, experimental sessions. We identified stereotyped responses to stimulation in lower extremity musculature, and examined local field potential responses to EES using high-density recording bipoles. Finally, spatial electrode encoding enabled machine learning models to accurately perform EES parameter inference for unseen stimulation electrodes, reducing the need for extensive training data in future deep models. SignificanceWe report the development and chronic large animal in vivo evaluation of a high-density EES paddle array containing active electronics. Our results provide a foundation for more advanced computation and processing to be integrated directly into devices implanted at the neural interface, opening new avenues for the study of nervous system function and new therapies to treat neural injury and dysfunction.

Multi-Modality Deep Infarct: Non-invasive identification of infarcted myocardium using composite in-silico-human data learning

Mehdi, R. R. — 2024-06-03

Myocardial infarction (MI) continues to be a leading cause of death worldwide. The precise quantification of infarcted tissue is crucial to diagnosis, therapeutic management, and post-MI care. Late gadolinium enhancement-cardiac magnetic resonance (LGE-CMR) is regarded as the gold standard for precise infarct tissue localization in MI patients. A fundamental limitation of LGE-CMR is the invasive intravenous introduction of gadolinium-based contrast agents that present potential high-risk toxicity, particularly for individuals with underlying chronic kidney diseases. Herein, we develop a completely non-invasive methodology that identifies the location and extent of an infarct region in the left ventricle via a machine learning (ML) model using only cardiac strains as inputs. In this transformative approach, we demonstrate the remarkable performance of a multi-fidelity ML model that combines rodent-based in-silico-generated training data (low-fidelity) with very limited patient-specific human data (high-fidelity) in predicting LGE ground truth. Our results offer a new paradigm for developing feasible prognostic tools by augmenting synthetic simulation-based data with very small amounts of in-vivo human data. More broadly, the proposed approach can significantly assist with addressing biomedical challenges in healthcare where human data are limited.

Insula uses overlapping codes for emotion in self and others

Xiao, J. — 2024-06-08

In daily life, we must recognize others emotions so we can respond appropriately. This ability may rely, at least in part, on neural responses similar to those associated with our own emotions. We hypothesized that the insula, a cortical region near the junction of the temporal, parietal, and frontal lobes, may play a key role in this process. We recorded local field potential (LFP) activity in human neurosurgical patients performing two tasks, one focused on identifying their own emotional response and one on identifying facial emotional responses in others. We found matching patterns of gamma- and high-gamma band activity for the two tasks in the insula. Three other regions (MTL, ACC, and OFC) clearly encoded both self- and other-emotions, but used orthogonal activity patterns to do so. These results support the hypothesis that the insula plays a particularly important role in mediating between experienced vs. observed emotions.

Peptidomimetic inhibitors targeting TrkB/PSD-95 signaling improves cognition and seizure outcomes in an Angelman Syndrome mouse model

Huie, E. Z. — 2024-06-08

Angelman syndrome (AS) is a rare genetic neurodevelopmental disorder with profoundly debilitating symptoms with no FDA-approved cure or therapeutic. Brain-derived neurotrophic factor (BDNF), and its receptor TrkB, have a well-established role as regulators of synaptic plasticity, dendritic outgrowth, dendritic spine formation and maintenance. Previously, we reported that the association of PSD-95 with TrkB is critical for intact BDNF signaling in the AS mouse model, as illustrated by attenuated PLC{gamma} and PI3K signaling and intact MAPK pathway signaling. These data suggest that drugs tailored to enhance the TrkB-PSD-95 interaction may provide a novel approach for the treatment of AS and a variety of NDDs. To evaluate this critical interaction, we synthesized a class of high-affinity PSD-95 ligands that bind specifically to the PDZ3 domain of PSD-95, denoted as Syn3 peptidomimetic ligands. We evaluated Syn3 and its analog D-Syn3 (engineered using dextrorotary (D)-amino acids) in vivo using the Ube3a exon 2 deletion mouse model of AS. Following systemic administration of Syn3 and D-Syn3, we demonstrated improvement in the seizure domain of AS. Learning and memory using the novel object recognition assay also illustrated improved cognition following Syn3 and D-Syn3, along with restored long-term potentiation. Finally, D-Syn3 treated mice showed a partial rescue in motor learning. Neither Syn3 nor D-Syn3 improved gross exploratory locomotion deficits, nor gait impairments that have been well documented in the AS rodent models. These findings highlight the need for further investigation of this compound class as a potential therapeutic for AS and other genetic NDDs.

Combining supervised and unsupervised analyses to quantify behavioral phenotypes and validate therapeutic efficacy in a triple transgenic mouse model of Alzheimer's disease

Del Rosario Hernandez, T. — 2024-06-08

Behavioral testing is an essential tool for evaluating cognitive function and dysfunction in preclinical research models. This is of special importance in the study of neurological disorders such as Alzheimers disease. However, the reproducibility of classic behavioral assays is frequently compromised by interstudy variation, leading to ambiguous conclusions about the behavioral markers characterizing the disease. Here, we identify age- and genotype-driven differences between 3xTg-AD and non-transgenic control mice using a low-cost, highly customizable behavioral assay that requires little human intervention. Through behavioral phenotyping combining both supervised and unsupervised behavioral classification methods, we are able to validate the preventative effects of the immunosuppressant cyclosporine A in a rodent model of Alzheimers disease, as well as the partially ameliorating effects of candidate drugs nebivolol and cabozantinib. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=158 SRC="FIGDIR/small/597924v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1c61729org.highwire.dtl.DTLVardef@1abd0corg.highwire.dtl.DTLVardef@1719439org.highwire.dtl.DTLVardef@1ba4491_HPS_FORMAT_FIGEXP M_FIG C_FIG

Single-cell transcriptome unravels spermatogonial stem cells and dynamic heterogeneity of spermatogenesis in seasonal breeding teleost

Yang, Y. — 2024-06-08

Seasonal spermatogenesis in fish is driven by spermatogonial stem cells (SSCs), which undergo a complex cellular process to differentiate into mature sperm. In this study, we characterized spermatogenesis in the large yellow croaker (Larimichthys crocea), a marine fish of significant commercial value, based on a high-resolution single-cell RNA-seq atlas of testicular cells from three distinct developmental stages- juvenile, adult differentiating and regressed testes. We detailed continuous developmental trajectory of spermatogenic cells, from spermatogonia to spermatids, elucidating the molecular events involved in spermatogenesis. We uncovered dynamic heterogeneity in cellular compositions throughout the annual reproductive cycle, accompanied by strong molecular signatures within specific testicular cells. Notably, we identified a distinct population of SSCs and observed a critical metabolic transition from glycolysis to oxidative phosphorylation, enhancing our understanding of the biochemical and molecular characteristics of SSCs. Additionally, we elucidated the interactions between somatic cells and spermatogonia, illuminating the mechanisms that regulate SSCs development. Overall, this work enhances our understanding of spermatogenesis in seasonal breeding teleost and provides essential insights for the further conservation and culture of SSCs. Summary statementOur study reveals new insights into the development of spermatogonial stem cells (SSCs), potentially impacting further conservation and culture of SSCs in teleost.

Mutation and cell state compatibility is required and targetable in Ph+ acute lymphoblastic leukemia minimal residual disease

Winter, P. S. — 2024-06-10

Efforts to cure BCR::ABL1 B cell acute lymphoblastic leukemia (Ph+ ALL) solely through inhibition of ABL1 kinase activity have thus far been insufficient despite the availability of tyrosine kinase inhibitors (TKIs) with broad activity against resistance mutants. The mechanisms that drive persistence within minimal residual disease (MRD) remain poorly understood and therefore untargeted. Utilizing 13 patient-derived xenograft (PDX) models and clinical trial specimens of Ph+ ALL, we examined how genetic and transcriptional features co-evolve to drive progression during prolonged TKI response. Our work reveals a landscape of cooperative mutational and transcriptional escape mechanisms that differ from those causing resistance to first generation TKIs. By analyzing MRD during remission, we show that the same resistance mutation can either increase or decrease cellular fitness depending on transcriptional state. We further demonstrate that directly targeting transcriptional state-associated vulnerabilities at MRD can overcome BCR::ABL1 independence, suggesting a new paradigm for rationally eradicating MRD prior to relapse. Finally, we illustrate how cell mass measurements of leukemia cells can be used to rapidly monitor dominant transcriptional features of Ph+ ALL to help rationally guide therapeutic selection from low-input samples. HIGHLIGHTSO_LIRelapse after remission on TKI can harbor mutations in ABL1, RAS, or neither C_LIO_LIMutations and development-like cell state dictate fitness in residual disease C_LIO_LICo-targeting cell state and ABL1 markedly reduces MRD C_LIO_LIBiophysical measurements provide an integrative, rapid measurement of cell state C_LI

Surviving Medical School During a Pandemic: Experiences of New York Medical Students During the Height of SARS-CoV-2

Knight, L. M. — 2024-06-21

BackgroundThe COVID-19 pandemic dramatically altered the landscape of medical education. While patients overwhelmed hospital systems, lockdowns and social distancing recommendations took priority, and medical education was pushed online. Early in 2020, New York State (NYS) was hit especially hard by COVID-19. ObjectiveThis study sought to understand the effect of the COVID-19 pandemic on medical students well-being and education. MethodsNYS medical students responded to a six-question survey during April and May 2020. Questions assessed self-reported changes in stress levels, academic performance, and board preparation efforts. Open-ended data was analyzed using a modified grounded theory approach. Results488 responses across 11 medical schools were included (response rate of 5.8%). Major themes included: standardized test-related stressors (23%), study-related changes (19%), education and training concerns (17%), financial stressors (12%), and additional family obligations (12%). Second year students reported more stress/anxiety than students in other years (95.9%, p-value< 0.00001). Reported stress/anxiety, effects on exam preparation, and anticipated academic effect varied by geographics. ConclusionsWhile all NYS medical students reported being greatly affected, those closest to the NY City pandemic epi-center and closest to taking the Step 1 exam were the most distressed. Lack of flexibility of the medical education system during this public health emergency contributed to worsened student well-being. It is time to make plans for supporting the long-term mental health needs of these physicians-in-training and to examine ways the academic medical community can better adapt to the needs of students affected by a large public health emergency in the future.

Gut-to-brain regulation of Drosophila aging through neuropeptide F, insulin and juvenile hormone

Chen, J. — 2024-06-30

Dietary restriction slows aging in many animals, while in some cases the sensory signals from diet alone are sufficient to retard or accelerate lifespan. The digestive tract is a candidate location to sense nutrients, where neuropeptides secreted by enteroendocrine cells (EEC) produce systemic signals in response to food. Here we measure how Drosophila neuropeptide F (NPF) is secreted into adult circulation by enteroendocrine cells and find that specific enteroendocrine cells differentially respond to dietary sugar and yeast. Lifespan is increased when gut NPF is genetically depleted, and this manipulation is sufficient to blunt the longevity benefit conferred by dietary restriction. Depletion of NPF receptors at insulin producing neurons of the brain also increases lifespan, consistent with observations where loss of gut NPF decreases neuronal insulin secretion. The longevity conferred by repressing gut NPF and brain NPF receptors is reversed by treating adults with a juvenile hormone (JH) analog. JH is produced by the adult corpora allata, and inhibition of the insulin receptor at this tissue decreases JH titer and extends lifespan, while this longevity is restored to wild type by treating adults with a JH analog. Overall, enteroendocrine cells of the gut modulate Drosophila aging through interorgan communication mediated by a gut- brain-corpora allata axis, and insulin produced in the brain impacts lifespan through its control of JH titer. These data suggest that we should consider how human incretins and their analogs, which are used to treat obesity and diabetes, may impact aging. Significance StatementNeuropeptide F (NPF) produced in the Drosophila gut is an insulin-regulatory hormone (incretin) that is secreted into adult circulation in response to feeding and diet. Suppression of gut NPF extends Drosophila longevity, as does knockdown of Neuropeptide F receptors at the insulin-producing medial neurosecretory cells in the brain that control the titer of juvenile hormone. Gut hormones and brain insulin regulate lifespan because they control juvenile hormone titer, which itself is the master endocrine regulator of Drosophila aging. Gut NPF modulates Drosophila aging through the integration of nutrient sensing, insulin signaling and juvenile hormone. Given the role of incretin-mimetic drugs to treat diabetes and obesity, it may be time to consider how incretin analogs could impact human aging.

Aspartic proteases are abundant and active in acidified wound fluid

Barbosa da Silva, E. — 2024-06-30

Wound healing necessitates a balance between synthesis and breakdown of extracellular matrix components, which is tightly regulated by proteases and their inhibitors. Studies have shown that treatment of poorly healing wounds with acid results in improved healing. In this study, we systematically evaluated changes in proteolytic activity of murine wound fluid upon acidification. A library of 228 synthetic peptides served as reporters of protease activity at pH 7.4, pH 5.0 and pH 3.5. The peptide digestion patterns differed at each pH, revealing that proteases active at pH 7.4 are inactivated at pH 3.5. Notably, aspartic acid proteases emerged as the dominant active enzymes at pH 3.5 and their activity was inhibited by pepstatin. Using a fluorogenic substrate, we quantified aspartic protease activity across varying pH levels and demonstrated optimal activity between pH 3.0 and 3.8. This activity was detectable as early as one day post-injury and persisted over the following ten days. Importantly, human wound fluid exhibited the same activity profile, validating the mouse model as a relevant system for studying acid-mediated wound healing processes.

The evolutionary modifications of a GoLoco motif in the AGS protein facilitate micromere formation in the sea urchin embryo

Emura, N. — 2024-07-02

The evolutionary introduction of asymmetric cell division (ACD) into the developmental program facilitates the formation of a new cell type, contributing to developmental diversity and, eventually, to species diversification. The micromere of the sea urchin embryo may serve as one of those examples: An ACD at the 16-cell stage forms micromeres unique to echinoids among echinoderms. We previously reported that a polarity factor, Activator of G-protein Signaling (AGS), plays a crucial role in micromere formation. However, AGS and its associated ACD factors are present in all echinoderms and across most metazoans. This raises the question of what evolutionary modifications of AGS protein or its surrounding molecular environment contributed to the evolutionary acquisition of micromeres only in echinoids. In this study, we learned that the GoLoco motifs at the AGS C-terminus play critical roles in regulating micromere formation in sea urchin embryos. Further, other echinoderms AGS or chimeric AGS that contain the C-terminus of AGS orthologs from various organisms showed varied localization and function in micromere formation. In contrast, the sea star or the pencil urchin orthologs of other ACD factors were consistently localized at the vegetal cortex in the sea urchin embryo, suggesting that AGS may be a unique variable factor that facilitates ACD diversity among echinoderms. Consistently, sea urchin AGS appears to facilitate micromere-like cell formation and accelerate the enrichment timing of the germline factor Vasa during early embryogenesis of the pencil urchin, an ancestral type of sea urchin. Based on these observations, we propose that the molecular evolution of a single polarity factor facilitates ACD diversity while preserving the core ACD machinery among echinoderms and beyond during evolution. HighlightsO_LIEvolutionary modifications of GoLoco motifs are critical for AGS function in micromere formation in the sea urchin embryo. C_LIO_LIThe chimeric AGS, which contains the C-terminus of AGS orthologs from various organisms, suggests that human LGN, pencil urchin AGS, and Drosophila Pins compensate for the activity of sea urchin AGS. C_LIO_LISea urchin AGS (SpAGS) regulates the localization of the conserved asymmetric cell division (ACD) machinery members at the vegetal cortex. C_LIO_LISpAGS is a variable factor facilitating ACD diversity during species diversification. C_LI

Bulk and Single-cell Transcriptomic Brain Data Identify Overlapping Processes and Cell-types with Human AUD and Mammalian Models of Alcohol Use

Huggett, S. B. — 2024-07-03

This study explores the neurobiological underpinnings of alcohol use disorder (AUD) by integrating bulk and single-cell transcriptomic data from humans, primates, and mice across three brain regions associated with addiction (i.e., prefrontal cortex (PFC), nucleus accumbens (NAc), and central amygdala (CeA)). We compared AUD RNA expression and cell-type abundance from 92 human brain to data from 53 primates and 90 mice engaged in diverse alcohol use paradigms. The findings revealed significant and reproducible correlations between human AUD and mammalian models of alcohol use that vary by tissue, species, and behavioral paradigm. The strongest correlations occurred between primate and mouse models of binge drinking (i.e., high drinking in the dark). Certain primate models demonstrated that the brain RNA correlations with human alcohol use disorder (AUD) were approximately 40% as strong as the correlations observed within human samples themselves. By integrating single-cell transcriptomic data, this study observed decreased oligodendrocyte proportions in the PFC and NAc of human AUD with similar trends in animal models. Gene co-expression network analyses revealed conserved systems associated with human AUD and animal models of heavy/binge alcohol consumption. Gene co-expression networks were enriched for pathways related to inflammation, myelination, and synaptic plasticity and the genes within them accounted for [~]20% of the heritability in human alcohol consumption. Identified hub genes were associated with relevant traits (e.g., impulsivity, motivation) in humans and mice. This study sheds light on conserved biological entities underlying AUD and chronic alcohol use, providing insights into the cellular, genetic, and neuromolecular basis across species.

Methamphetamine-induced adaptation of learning rate dynamics depend on baseline performance.

Kirschner, H. — 2024-07-08

The ability to calibrate learning according to new information is a fundamental component of an organisms ability to adapt to changing conditions. Yet, the exact neural mechanisms guiding dynamic learning rate adjustments remain unclear. Catecholamines appear to play a critical role in adjusting the degree to which we use new information over time, but individuals vary widely in the manner in which they adjust to changes. Here, we studied the effects of a low dose of methamphetamine (MA), and individual differences in these effects, on probabilistic reversal learning dynamics in a within-subject, double-blind, randomized design. Participants first completed a reversal learning task during a drug-free baseline session to provide a measure of baseline performance. Then they completed the task during two sessions, one with MA (20 mg oral) and one with placebo (PL). First, we showed that, relative to PL, MA modulates the ability to dynamically adjust learning from prediction errors. Second, this effect was more pronounced in participants who performed moderately low at baseline. These results present novel evidence for the involvement of catecholaminergic transmission on learning flexibility and highlights that baseline performance modulates the effect of the drug.

Aberration correction in long GRIN lens-based microendoscopes for extended field-of-view two-photon imaging in deep brain regions

Sattin, A. — 2024-07-24

Two-photon (2P) fluorescence imaging through gradient index (GRIN) lens-based endoscopes is fundamental to investigate the functional properties of neural populations in deep brain circuits. However, GRIN lenses have intrinsic optical aberrations, which severely degrade their imaging performance. GRIN aberrations decrease the signal-to-noise ratio (SNR) and spatial resolution of fluorescence signals, especially in lateral portions of the field-of-view (FOV), leading to restricted FOV and smaller number of recorded neurons. This is especially relevant for GRIN lenses of several millimeters in length, which are needed to reach the deeper regions of the rodent brain. We have previously demonstrated a novel method to enlarge the FOV and improve the spatial resolution of 2P microendoscopes based on GRIN lenses of length < 4.1 mm (Antonini et al. eLife 2020). However, previously developed microendoscopes were too short to reach the most ventral regions of the mouse brain. In this study, we combined optical simulations with fabrication of aspherical polymer microlenses through three-dimensional (3D) microprinting to correct for optical aberrations in long (length > 6 mm) GRIN lens-based microendoscopes (diameter, 500 {micro}m). Long corrected microendoscopes had improved spatial resolution, enabling imaging in significantly enlarged FOVs. Moreover, using synthetic calcium data we showed that aberration correction enabled detection of cells with higher SNR of fluorescent signals and decreased cross-contamination between neurons. Finally, we applied long corrected microendoscopes to perform large-scale and high precision recordings of calcium signals in populations of neurons in the olfactory cortex, a brain region laying approximately 5 mm from the brain surface, of awake head-tethered mice. Long corrected microendoscopes are powerful new tools enabling population imaging with unprecedented large FOV and high spatial resolution in the most ventral regions of the mouse brain.

Going around the bend to understand the role of leg coalescence in metachronal swimming

Tack, N. B. — 2024-07-25

Many of the most abundant aquatic invertebrates display metachronal swimming by sequentially beating closely spaced flexible appendages. Common biophysical mechanisms like appendage spatial asymmetry and phase drive the success and performance of this locomotor mode, which is generally explained by the need to maximize thrust production. However, the potential role of these mechanisms in drag reduction, another important contributor to the overall swimming performance, has yet to be evaluated. We present a comprehensive overview of the morphological, functional, and physical mechanisms promoting drag reduction during metachronal swimming by exploring appendage differential bending and leg grouping (coalescence). We performed -CT and in-vivo velocimetry measurements of shrimp (Palaemonetes vulgaris) to design a five-legged robotic metachronal analog. This test platform enabled simultaneous flow and force measurements to quantify the thrust and drag forces produced by flexible and stiff pleopods (legs) beating independently or coalescing. We tested the hypothesis that coalescence and bending effectively reduce drag during the recovery stroke (RS). The curved cross-section of the pleopods enables passive asymmetrical bending during the RS to reduce their coefficient of drag by up to 75.8% relative to stiff pleopods. Bending promotes physical interactions facilitating the coalescence of three pleopods at any time during the RS to reduce drag such that the mean net thrust produced during coalescence is increased by 30.2%. These improvements are explained by the production of a weaker wake compared with stiff and non-coalescing pleopods. Our results describe fundamental biological and physical components of metachronal propulsion that may aid the development of novel bio-inspired underwater vehicles. Summary statementShrimp swimming legs bend nearly horizontally and cluster together during metachronal propulsion to reduce drag and improve the overall swimming performance.

Exploring the impacts of human breast milk functional lipidome on infant health and growth outcomes in early life using lipid bioinformatics

Ganeshalingam, M. — 2024-07-26

Human breast milk lipidome is complex, and how changes in the functional lipid metabolism converge systematically to alter infants health outcomes is poorly understood. We used human breast milk and infant-mother dyads as a test system to demonstrate how the application of improved lipid bioinformatics can be effective in discerning systematic changes in functional lipid metabolism providing novel discoveries of how lactational programming in early life can influence infant health and growth outcomes. The study consisted of 40 mother-infant dyads where breast milk, maternal diet, infant anthropometrics [fat mass index (FMI), length z score, BMI z score, fat-free mass index (FFMI)], and infant atopic disease outcome (ear infection, cold, wheezing, diarrhea, and eczema) were collected at one and four months postpartum. Integrated Lipid Bioinformatics analyses were conducted using XLSTAT, Metaboanalyst 5.0. R software, Lipid Search, Xcalibur, and Cytoscape software. The results showed breast milk lipidome ordinated into distinct clusters based on maternal BMI status, and differences in developmental and atopic disease outcomes following redundancy analysis. Specifically, lipids from obese mothers clustered with FMI and eczema, while lipids from non-obese mothers clustered with FFM and wheezing. Receiver operating analysis was effective in identifying potential lipid biomarkers that were significantly associated with infant FMI, FFMI, and eczema during early life. Sphingolipid and glycerophospholipid pathways were significantly associated with the altered breast milk lipidome impacting infant development and atopic disease outcome during the first year of life. The findings following the advanced lipid bioinformatics suggest that the breastmilk functional lipid metabolism appears to play a key role in lipid-mediated lactational programming influencing development and atopic disease outcome, and present opportunities for potential dietary intervention in early life.

Cognitive sequences in obsessive-compulsive disorder are supported by frontal cortex ramping activity and mediated by symptom severity

Doyle, H. — 2024-07-29

Completing sequences is a routine part of daily life. Many are abstract, defined by a rule governing the order rather than the identity of individual steps (e.g., getting dressed). In obsessive-compulsive disorder (OCD), excessive ritualistic behaviors suggest a disruption in abstract sequence completion. Executing abstract sequences requires at least two levels in a hierarchy of cognitive control: abstract sequential control (tracking steps) and task switching (shifting between tasks). While task switching has been studied in OCD, little is known in a sequential context. Understanding both hierarchical control types is key to uncovering how abstract sequences with nested task switches are processed in OCD. Previous studies showed that the rostrolateral prefrontal cortex (RLPFC) supports abstract sequence monitoring in healthy individuals with an increase in activity across each sequence, a dynamic known as "ramping". Ramping outside the RLPFC is potentially indicative of other sequence-related processes such as progress towards a goal and increasing working memory load. Therefore, we hypothesized that abstract sequential control deficits would correspond to altered ramping dynamics in RLPFC and other cortical regions. Second, we predicted task switching deficits in OCD, coupled with altered activity in cortical regions canonically implicated in task level control. We found partial support for both hypotheses. Abstract sequential control did not show behavioral differences in OCD but did show increased overall ramping in the anterior cingulate cortex (ACC) and superior frontal sulcus (SFS) and ramping differences in additional, novel cortical regions according to abstract sequence complexity. In contrast, behavioral differences were observed for task switching in OCD without neural differences between the groups. Together, these results suggest a group of areas support sequential control differentially in OCD than in healthy controls, despite behavioral similarity, and that this observation is likely not the result of neural deficits in task switching. These findings thus provide insight into OCD during complex behaviors more similar to daily life where sequence and task level control are intertwined and may inform future potential treatment.

Monkey lateral prefrontal cortex subregions differentiate between perceptual exposure to visual stimuli

Leaman, K. — 2024-07-29

Each day, humans must parse visual stimuli with varying amounts of perceptual experience, ranging from incredibly familiar to entirely new. Even when choosing a novel to buy at a bookstore, one sees covers they have repeatedly experienced intermixed with recently released titles. Visual exposure to stimuli has distinct neural correlates in the lateral prefrontal cortex (LPFC) of nonhuman primates. However, it is currently unknown if this function may be localized to specific subregions within LPFC. Specifically, we aimed to determine whether the posterior fundus of area 46 (p46f), an area that responds to deviations from learned sequences, also responds to less frequently presented stimuli outside of the sequential context. We compare responses in p46f to the adjacent subregion, posterior ventral area 46 (p46v), which we propose may be more likely to show exposure-dependent responses due to its proximity to novelty responsive regions. To test whether p46f or p46v represent perceptual exposure, we performed awake functional magnetic resonance imaging (fMRI) on three male monkeys as they observed visual stimuli that varied in their number of daily presentations. Here we show that p46v, but not p46f, shows preferential activation to stimuli with low perceptual exposure, further localizing exposure-dependent effects in monkey LPFC. These results align with previous research that has found novelty responses in ventral LPFC and are consistent with the proposal that p46f performs a sequence-specific function. Further, they expand on our knowledge of the specific role of LPFC subregions and localize perceptual exposure processing within this broader brain region.

Calcium phosphate nanoclusters modify periodontium remodeling and minimize orthodontic relapse

Cuylear, D. L. — 2024-07-30

Orthodontic relapse is one of the most prevalent concerns of orthodontic therapy. Relapse results in patients teeth reverting towards their pretreatment positions, which increases the susceptibility to functional problems, dental disease, and substantially increases the financial burden for retreatment. This phenomenon is thought to be induced by rapid remodeling of the periodontal ligament (PDL) in the early stages and poor bone quality in the later stages. Current therapies, including fixed or removable retainers and fiberotomies, have limitations with patient compliance and invasiveness. Approaches using biocompatible biomaterials, such as calcium phosphate polymer-induced liquid precursors (PILP), is an ideal translational approach for minimizing orthodontic relapse. Here, post-orthodontic relapse is reduced after a single injection of high concentration PILP (HC-PILP) nanoclusters by altering PDL remodeling in the early stage of relapse and improving trabecular bone quality in the later phase. HC-PILP nanoclusters are achieved by using high molecular weight poly aspartic acid (PASP, 14 kDa) and poly acrylic acid (PAA, 450 kDa), which resulted in a stable solution of high calcium and phosphate concentrations without premature precipitation. In vitro results show that HC-PILP nanoclusters prevented collagen type-I mineralization, which is essential for the tooth-periodontal ligament (PDL)-bone interphase. In vivo experiments show that the PILP nanoclusters minimize relapse and improve the trabecular bone quality in the late stages of relapse. Interestingly, PILP nanoclusters also altered the remodeling of the PDL collagen during the early stages of relapse. Further in vitro experiments showed that PILP nanoclusters alter the fibrillogenesis of collagen type-I by impacting the protein secondary structure. These findings propose a novel approach for treating orthodontic relapse and provide additional insight into the PILP nanoclusters structure and properties on collagenous structure repair.

Unsupervised learning as a computational principle works in visual learning of natural scenes, but not of artificial stimuli

Watanabe, T. — 2024-07-31

Unsupervised learning--learning through repeated exposure without instruction or reward--is central to both machine learning and human cognition, including language acquisition and statistical learning. However, its role in visual perceptual learning (VPL) remains debated, as previous studies have not shown VPL for task-irrelevant but visible features, particularly in artificial stimuli. Here, we show that task-irrelevant exposure to natural scene images induces robust VPL, while artificial images that lack complex structure characteristics of natural scene images, known as higher-order statistics, do not. Behavioral and fMRI results suggest that although unsupervised learning underlies VPL, it can be suppressed by top-down attention. Higher-order statistics may evade this suppression, possibly because their slower processing reaches visual areas beyond V1 outside the optimal temporal window for attentional suppression. These findings suggest that unsupervised learning underlies VPL, but its occurrence depends on both higher-order stimulus structure and the brains attentional gating mechanisms.

Synergistic combination therapy with ONC201 or ONC206, Enzalutamide and Darolutamide in preclinical studies of castration-resistant prostate cancer

Wu, L. J. — 2024-08-01

Androgen receptor (AR) signaling plays a primary role in prostate cancer progression. Non-steroidal anti- androgens (NSAA) including enzalutamide, and apalutamide have been used to treat patients with advanced disease. However, patients with metastatic castration-resistant prostate cancer (mCPRC) develop resistance, resulting in limited overall survival benefit. Darolutamide is a novel next-generation androgen receptor- signaling inhibitor that is FDA approved for non-metastatic castration resistant prostate cancer (nmCRPC). Imipridone ONC201/TIC10 is first-in-class small molecule that activates the integrated stress response (ISR) and upregulates TNF-related apoptosis-inducing ligand (TRAIL). Our study investigates ISR and AR signaling in anti-tumor efficacy with ONC201 and enzalutamide or darolutamide against mCRPC cells. mCRPC cell lines 22RV1, LNCaP, DU145 and PC3 were treated with ONC201, darolutamide, and enzalutamide as single agents or in combinations. Combinations of ONC201 and darolutamide or enzalutamide demonstrated synergistic effects in mCRPC cells. Combinations of ONC201 and darolutamide or enzalutamide reduced PSA levels in LNCaP cells and induced of ATF4 in both LNCaP and 22RV1 cell lines. Darolutamide synergized with ONC201 regardless of AR status or castration sensitivity in vitro. Flow cytometric analysis showed increased intra-tumoral NK cells in mice treated with ONC201 and combination of ONC201 and darolutamide. Trends of increased TRAIL activation within NK cells were also observed in treatment groups. ONC201 and darolutamide demonstrated anti-tumor effects in vivo in the 22RV1 CRPC model. Our results prompt further translational and clinical studies with imipridones ONC201 or ONC201 in combination with enzalutamide or darolutamide for treatment of castrate resistant advanced or metastatic prostate cancer.

MRGM: An enhanced catalog of mouse gut microbial genomes substantially broadening taxonomic and functional landscapes

Kim, N. — 2024-08-12

Mouse gut microbiome research is pivotal for understanding the human gut microbiome, providing insights into disease modeling, host-microbe interactions, and the dietary influence on the gut microbiome. To enhance the translational value of mouse gut microbiome studies, we need detailed and high-quality catalogs of mouse gut microbial genomes. We introduce the Mouse Reference Gut Microbiome (MRGM), a comprehensive catalog with 42,245 non-redundant mouse gut bacterial genomes across 1,524 species. MRGM marks a 40% increase in the known taxonomic diversity of mouse gut microbes, capturing previously underrepresented lineages through refined genome quality assessment techniques. MRGM not only broadens the taxonomic landscape but also enriches the functional landscape of the mouse gut microbiome. Using deep learning, we have elevated the Gene Ontology annotation rate for mouse gut microbial proteins from 3.2% with orthology to 60%, marking an over 18-fold increase. MRGM supports both DNA- and marker-based taxonomic profiling by providing custom databases, surpassing previous catalogs in performance. Finally, taxonomic and functional comparisons between human and mouse gut microbiota reveal diet-driven divergences in their taxonomic composition and functional enrichment. Overall, our study highlights the value of high-quality microbial genome catalogs in advancing our understanding of the co-evolution between gut microbes and their host.

An instantaneous voice synthesis neuroprosthesis

Wairagkar, M. — 2024-08-19

Brain computer interfaces (BCIs) have the potential to restore communication to people who have lost the ability to speak due to neurological disease or injury. BCIs have been used to translate the neural correlates of attempted speech into text1-3. However, text communication fails to capture the nuances of human speech such as prosody, intonation and immediately hearing ones own voice. Here, we demonstrate a "brain-to-voice" neuroprosthesis that instantaneously synthesizes voice with closed-loop audio feedback by decoding neural activity from 256 microelectrodes implanted into the ventral precentral gyrus of a man with amyotrophic lateral sclerosis and severe dysarthria. We overcame the challenge of lacking ground-truth speech for training the neural decoder and were able to accurately synthesize his voice. Along with phonemic content, we were also able to decode paralinguistic features from intracortical activity, enabling the participant to modulate his BCI-synthesized voice in real-time to change intonation, emphasize words, and sing short melodies. These results demonstrate the feasibility of enabling people with paralysis to speak intelligibly and expressively through a BCI.

Immune checkpoint inhibitor treatment does not impair ovarian or endocrine function in a mouse model of triple negative breast cancer

De La Cruz, P. — 2024-08-19

BackgroundRepresenting 15-20% of all breast cancer cases, triple negative breast cancer (TNBC) is diagnosed more frequently in reproductive-age women and exhibits higher rates of disease metastasis and recurrence when compared with other subtypes. Few targeted treatments exist for TNBC, and many patients experience infertility and endocrine disruption as a result of frontline chemotherapy treatment. While they are a promising option for less toxic therapeutic approaches, little is known about the effects of immune checkpoint inhibitors on reproductive and endocrine function. ResultsOur findings in a syngeneic TNBC mouse model revealed that therapeutically relevant immunotherapies targeting PD-1, LAG-3, and TIM-3 had no effect on the quality and abundance of ovarian follicles, estrus cyclicity, or hormonal homeostasis. Similarly, in a tumor-free mouse model, we found that ovarian architecture, follicle abundance, estrus cyclicity, and ovulatory efficiency remain unchanged by PD-1 blockade. ConclusionsTaken together, our results suggest that immunotherapy may be a promising component of fertility-sparing therapeutic regimens for patients that wish to retain ovarian and endocrine function after cancer treatment.

An IL-17-DUOX2 axis controls gastrointestinal colonization by Candida albicans

Kakade, P. — 2024-08-19

Candida albicans is a ubiquitous fungus in the human gut microbiome as well as a prevalent cause of opportunistic mucosal and systemic disease. There is currently little understanding, however, as to how crosstalk between C. albicans and the host regulates colonization of this key niche. Here, we performed expression profiling on ileal and colonic tissues in germ-free mice colonized with C. albicans to define the global response to this fungus. We reveal that Duox2 and Duoxa2, encoding dual NADPH oxidase activity, are upregulated in both the ileum and colon, and that induction requires the C. albicans yeast-hyphal transition and the hyphal-specific toxin candidalysin. Hosts lacking the IL-17 receptor failed to upregulate Duox2/Duoxa2 in response to C. albicans, while addition of IL-17A to colonoids induced these genes together with the concomitant production of hydrogen peroxide. To directly define the role of Duox2/Duoxa2 in fungal colonization, antibiotic-treated mice lacking intestinal DUOX2 activity were evaluated for C. albicans colonization and host responses. Surprisingly, loss of DUOX2 function reduced fungal colonization at extended time points (>17 days colonization) and increased the proportion of hyphal cells in the gut. IL-17A levels were also elevated in C. albicans-colonized mice lacking functional DUOX2 highlighting cross-regulation between this cytokine and DUOX2. Together, these experiments reveal novel links between fungal cells, candidalysin toxin and the host IL-17-DUOX2 axis, and that a complex interplay between these factors regulates C. albicans filamentation and colonization in the gut.

Dissecting contributions of pulmonary arterial remodeling to right ventricular afterload in pulmonary hypertension

Neelakantan, S. — 2024-08-19

Pulmonary hypertension (PH) is defined as an elevation in the right ventricle (RV) afterload, characterized by increased hemodynamic pressure in the main pulmonary artery (PA). Elevations in RV afterload increase RV wall stress, resulting in RV remodeling and potentially RV failure. From a biomechanical standpoint, the primary drivers for RV afterload elevations include increases in pulmonary vascular resistance (PVR) in the distal vasculature and decreases in vessel compliance in the proximal PA. However, the individual contributions of the various vascular remodeling events toward the progression of PA pressure elevations and altered vascular hemodynamics remain elusive. In this study, we used a subject-specific one-dimensional (1D) fluid-structure interaction (FSI) model to investigate the alteration of pulmonary hemodynamics in PH and to quantify the contributions of vascular stiffening and increased resistance towards increased main pulmonary artery (MPA) pressure. We used a combination of subject-specific hemodynamic measurements, ex-vivo mechanical testing of arterial tissue specimens, and ex-vivo X-ray micro-tomography imaging to develop the 1D-FSI model and dissect the contribution of PA remodeling events towards alterations in the MPA pressure waveform. Both the amplitude and pulsatility of the MPA pressure waveform were analyzed. Our results indicated that increased distal resistance has the greatest effect on the increase in maximum MPA pressure, while increased stiffness caused significant elevations in the characteristic impedance. The method presented in this study will serve as an essential step toward understanding the complex interplay between PA remodeling events that leads to the most severe adverse effect on RV dysfunction.

Population coding under the scale-invariance of high-dimensional noise

Moosavi, S. A. — 2024-08-26

High-dimensional neural activities exhibiting scale-invariant, power-law noise spectra are ubiquitously observed across various brain regions and species. However, their impact on information coding remains unclear. We provide scaling conditions for noise covariance that clarify the boundedness of information and establish a quantitative relation between information capacity and population size, based on the properties of scale-invariant noise covariance observed in stimulus-evoked activities of mouse V1 neurons. Our analysis reveals that sublinearly scaling small noise components align sufficiently with the signal direction, enabling neurons to convey stimulus information unboundedly as population size increases. These findings demonstrate that the quasi-universal scaling of neural noise covariance lays a foundation for understanding the scaling and boundedness of population codes, highlighting the critical need to consider the full spectrum of high-dimensional noise.

Adaptation to yeast restriction through the evolution of resource acquisition, macronutrient reserve, and ovarian function

Dasgupta, P. — 2024-08-26

Optimization of reproduction under dietary adversity is an important aspect of diet-dependent adaptation. Yet, little is known about the mechanism of such adaptive evolution. Here, we investigated a set of experimentally evolved populations of Drosophila melanogaster where early-life fecundity evolved as an adaptation to chronic protein restriction. We tested the role of resource acquisition and macronutrient storage, and changes in ovarian function that could have allowed such reproductive adaptation. We show that higher early-life fecundity was associated with the increased larval feeding rate, aiding in accumulation of higher protein content at the time of eclosion. Further evidence also suggests increase in reproductively allocated lipid content. Evolved females were found to have larger ovaries that also had a higher number of mature, post-vitellogenic oocytes that seem to readily allow the attainment of higher fecundity. Our results further show the evolution of plasticity in ovariole count (depending on mating status) and total egg-chamber count in females. These results are important in understanding the putative role of genetic variance and covariances that aid or limit the evolution of reproductive optimization, especially under nutritional adversity.

Modulation of blood-tumor barrier transcriptional programs improves intra-tumoral drug delivery and potentiates chemotherapy in GBM

Macias, J. L. J. — 2024-08-28

Glioblastoma (GBM) is the most common malignant primary brain tumor. GBM has an extremely poor prognosis and new treatments are badly needed. Efficient drug delivery to GBM is a major obstacle as the blood-brain barrier (BBB) prevents passage of the majority of cancer drugs into the brain. It is also recognized that the blood-brain tumor barrier (BTB) in the growing tumor represents a challenge. The BTB is heterogeneous and poorly characterized, but similar to the BBB it can prevent therapeutics from reaching effective intra-tumoral doses, dramatically hindering their potential. Here, we identified a 12-gene signature associated with the BTB, with functions related to vasculature development, morphogenesis and cell migration. We identified CDH5 as a core molecule in this set and confirmed its over-expression in GBM vasculature using spatial transcriptomics of GBM patient specimens. We found that the indirubin-derivative, 6-bromoindirubin acetoxime (BIA), could downregulate CDH5 and other BTB signature genes, causing endothelial barrier disruption in endothelial monolayers and BBB 3D spheroids in vitro. Treatment of tumor-bearing mice with BIA enabled increased intra-tumoral accumulation of the BBB non-penetrant chemotherapeutic drug cisplatin and potentiated cisplatin-mediated DNA damage by targeting DNA repair pathways. Finally, using an injectable BIA nanoparticle formulation, PPRX-1701, we significantly improved the efficacy of cisplatin in patient-derived GBM xenografts and prolonged their survival. Overall, our work reveals potential targets at the BTB for improved chemotherapy delivery and the bifunctional properties of BIA as a BTB modulator and potentiator of chemotherapy, supporting its further development.

Neuroendocrine differentiation (ND) in sensitivity of neuroendocrine tumor (NET) cells to ONC201/TIC10 cancer therapeutic

Ding, E. — 2024-08-30

Prostate cancer (PCa) neuroendocrine tumor (NET)-like cells with low or absent androgen receptor (AR) signaling cause hormone therapy resistance and poor prognosis. Small cell lung carcinoma (SCLC), a high-grade NET, presents with metastasis early and has poor survival. ONC201/TIC10 is a first-in-class cancer therapeutic with clinical activity in diffuse gliomas and neuroendocrine tumors. We hypothesized that markers of neuroendocrine differentiation, activation of the integrated stress response (ISR) and the TRAIL pathway, as well as the expression of ClpP, contribute to neuroendocrine tumor cell death and sensitivity to ONC201. We show that PCa and SCLC cell lines (N=6) are sensitive to ONC201, regardless of the extent of neuroendocrine differentiation. Endogenous levels of some NET markers (CgA, FoxO1, ENO2, PGP9.5, SOX2) are present in a spectrum in PCa and SCLC cell lines. Overexpression of neural transcription factor BRN2 in DU145 PCa cells does not increase expression of NET differentiation markers FoxO1, ENO2, PGP9.5, and CgA at 48 hours. However, the transient BRN2 overexpression showed slight decreases in some NET markers on the spectrum while maintaining sensitivity of PCa cells to ONC201 before any phenotypic change related to NET differentiation. Our results show that ONC201 has preclinical activity against PCa including those without NET markers or in PCa cells with transient overexpression of neural transcription factor BRN2. Our results have relevance to activity of ONC201 in PCa where most castrate-resistant androgen-independent cancers are not therapy resistant due to NET differentiation. Importantly, NET differentiation does not promote resistance to ONC201 supporting further clinical investigations across the spectrum of PCa.

Neuroendocrine Prostate Cancer Drivers SOX2 and BRN2 Confer Differential Responses to Imipridones ONC201, ONC206, and ONC212 in Prostate Cancer Cell Lines

Purcell, C. — 2024-08-30

Prostate cancer (PCa) is the leading cause death from cancer in men worldwide. Approximately 30% of castrate-resistant PCas become refractory to therapy due to neuroendocrine differentiation (NED) that is present in <1% of androgen-sensitive tumors. First-in-class imipridone ONC201/TIC10 has shown clinical activity against midline gliomas, neuroendocrine tumors and PCa. We explored the question of whether NED promotes sensitivity to imipridones ONC201 and ONC206 by inducible overexpression of SOX2 and BRN2, well-known neuroendocrine drivers, in human PCa cell lines DU145 or LNCaP. Slight protection from ONC201 or ONC206 with SOX2 and BRN2 overexpression was observed in the inducible LNCaP cells but not in the DU145 cells. At 2 months, there was an apparent increase in CLpP expression in LNCaP SOX2-overexpressing cells but this did not confer enhanced sensitivity to ONC201. DU145 SOX2-overexpressing cells had a significantly reduced ONC201 sensitivity than DU145 control cells. The results support the idea that treatment of castrate-resistant prostate cancer by imipridones may not be significantly impacted by neuroendocrine differentiation as a therapy-resistance mechanism. The results support further testing of imipridones across subtypes of androgen-sensitive and castrate-resistant prostate cancer.

Inferring in vivo murine cerebrospinal fluid flow using artificial intelligence velocimetry with moving boundaries and uncertainty quantification

Toscano, J. D. — 2024-08-30

Cerebrospinal fluid (CSF) flow is crucial for clearing metabolic waste from the brain, a process whose dysregulation is linked to neurodegenerative diseases like Alzheimers. Traditional approaches like particle tracking velocimetry (PTV) are limited by their reliance on single-plane two-dimensional measurements, which fail to capture the complex dynamics of CSF flow fully. To overcome these limitations, we employ Artificial Intelligence Velocimetry (AIV) to reconstruct three-dimensional velocities, infer pressure and wall shear stress, and quantify flow rates. Given the experimental nature of the data and inherent variability in biological systems, robust uncertainty quantification (UQ) is essential. Towards this end, we have modified the baseline AIV architecture to address aleatoric uncertainty caused by noisy experimental data, enhancing our measurement refinement capabilities. We also implement UQ for the model and epistemic uncertainties arising from the governing equations and network representation. Toward this end, we test multiple governing laws, representation models, and initializations. Our approach not only advances the accuracy of CSF flow quantification but also can be adapted to other applications that use physics-informed machine learning to reconstruct fields from experimental data, providing a versatile tool for inverse problems.

TRAIL receptor agonist TLY012 in combination with PD-1 inhibition promotes tumor regression in an immune-competent mouse model of pancreatic ductal adenocarcinoma

Louie, A. D. — 2024-08-30

Pancreatic ductal adenocarcinoma (PDAC) has an immunosuppressed, apoptosis-resistant phenotype. TLY012 is a pegylated recombinant Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL), an orphan drug for chronic pancreatitis and systemic sclerosis. Innate immune TRAIL signaling suppresses cancer. We hypothesized that combination of immune checkpoint-blocking anti-PD-1 antibody and TLY012 would have synergistic anti-tumor efficacy in immune-competent PDAC-bearing mice. PDAC tumor-bearing C57Bl/6 mice treated 10 mg/kg anti-mouse PD-1 antibody twice weekly and 10 mg/kg TLY012 three times weekly had reduced tumor growth and tumor volume at 70 days compared to either drug alone (all p<0.005). B-cell activating factor (BAFF), which promotes PDAC tumors, decreased to 44% of control mice with dual treatment at 7 days and remained decreased at 3 months. Long-term dual treatment showed the highest levels of proinflammatory cytokines interferon gamma (average 5.6 times control level, p=0.046), CCL5 (average 14.1 times control level, p=0.048), and interleukin-3 (IL-3, average 71.1 times control level, p=0.0053). Flow cytometry showed trends toward decreased circulating regulatory T cells, increased NK cells, and a higher proportion of CD8+ T cells within tumors in dual treatment group. In summary, combination of anti-PD-1 and TLY012 prevented growth of PDAC in an immunocompetent mouse model while increasing tumor-infiltrating CD8+ T cells, decreasing circulating T-regulatory cells and altering cytokine expression of CCL5, interferon gamma and IL-3 to promote proinflammatory, antitumor effects. Combining TLY012 and anti-mouse PD-1 creates changes in immune cell and cytokine levels to induce a more proinflammatory immune environment that contributes to decreased PDAC tumor growth.

Developmental and physiological impacts of pathogenic human huntingtin protein in the nervous system

Hana, T. A. — 2024-09-02

Huntingtons Disease (HD) is a neurodegenerative disorder, part of the nine identified inherited polyglutamine (polyQ) diseases. Most commonly, HD pathophysiology manifests in middle-aged adults with symptoms including progressive loss of motor control, cognitive decline, and psychiatric disturbances. Associated with the pathophysiology of HD is the formation of insoluble fragments of the huntingtin protein (htt) that tend to aggregate in the nucleus and cytoplasm of neurons. To track both the intracellular progression of the aggregation phenotype as well as the physiological deficits associated with mutant htt, two constructs of human HTT were expressed with varying polyQ lengths, non-pathogenic-htt (Q15, NP-htt) and pathogenic-htt (Q138, P-htt), with an N-terminal RFP tag for in vivo visualization. P-htt aggregates accumulate in the ventral nerve cord cell bodies as early as 24 hours post hatching and significant aggregates form in the segmental nerve branches at 48 hours post hatching. Organelle trafficking up-and downstream of aggregates formed in motor neurons showed severe deficits in trafficking dynamics. To explore putative downstream deficits of htt aggregation, ultrastructural changes of presynaptic motor neurons and muscles were assessed, but no significant effects were observed. However, the force and kinetics of muscle contractions were severely affected in P-htt animals, reminiscent of human chorea. Reduced muscle force production translated to altered locomotory behavior. A novel HD aggregation model was established to track htt aggregation throughout adulthood in the wing, showing similar aggregation patterns with larvae. Expressing P-htt in the adult nervous system resulted in significantly reduced lifespan, which could be partially rescued by feeding flies the mTOR inhibitor rapamycin. These findings advance our understanding of htt aggregate progression as well the downstream physiological impacts on the nervous system and peripheral tissues.

Dynamic and structural insights into allosteric regulation on MKP5 a dual-specificity phosphatase

Skeens, E. — 2024-09-05

Dual-specificity mitogen-activated protein kinase (MAPK) phosphatases (MKPs) directly dephosphorylate and inactivate the MAPKs. Although the catalytic mechanism of dephosphorylation of the MAPKs by the MKPs is established, a complete molecular picture of the regulatory interplay between the MAPKs and MKPs still remains to be fully explored. Here, we sought to define the molecular mechanism of MKP5 regulation through an allosteric site within its catalytic domain. We demonstrate using crystallographic and NMR spectroscopy approaches that residue Y435 is required to maintain the structural integrity of the allosteric pocket. Along with molecular dynamics simulations, these data provide insight into how changes in the allosteric pocket propagate conformational flexibility in the surrounding loops to reorganize catalytically crucial residues in the active site. Furthermore, Y435 contributes to the interaction with p38 MAPK and JNK, thereby promoting dephosphorylation. Collectively, these results highlight the role of Y435 in the allosteric site as a novel mode of MKP5 regulation by p38 MAPK and JNK.

Natural selection acting on complex traits hampers the predictive accuracy of polygenic scores in ancient samples

Anorve-Garibay, V. — 2024-09-11

The prediction of phenotypes from ancient humans has gained interest due to its potential to investigate the evolution of complex traits. These predictions are commonly performed using polygenic scores computed with DNA information from ancient humans along with genome-wide association studies (GWAS) data from present-day humans. However, numerous evolutionary processes could impact the prediction of phenotypes from ancient humans based on polygenic scores. In this work we investigate how natural selection impacts phenotypic predictions on ancient individuals using polygenic scores. We use simulations of an additive trait to analyze how natural selection impacts phenotypic predictions with polygenic scores. We simulate a trait evolving under neutrality, stabilizing selection and directional selection. We find that stabilizing and directional selection have contrasting effects on ancient phenotypic predictions. Stabilizing selection accelerates the loss of large-effect alleles contributing to trait variation. Conversely, directional selection accelerates the loss of small and large-effect alleles that drive individuals farther away from the optimal phenotypic value. These effects result in specific shared genetic variation patterns between ancient and modern populations which hamper the accuracy of polygenic scores to predict phenotypes. Furthermore, we conducted simulations that include realistic strengths of stabilizing selection and heritability estimates to show how natural selection could impact the predictive accuracy of ancient polygenic scores for two widely studied traits: height and body mass index. We emphasize the importance of considering how natural selection can decrease the reliability of ancient polygenic scores to perform phenotypic predictions on an ancient population.

niiv: Fast Self-supervised Neural ImplicitIsotropic Volume Reconstruction

Troidl, J. — 2024-09-13

Three-dimensional (3D) microscopy data often is anisotropic with significantly lower resolution (up to 8x) along the z axis than along the xy axes. Computationally generating plausible isotropic resolution from anisotropic imaging data would benefit the visual analysis of large-scale volumes. This paper proposes niiv, a self-supervised method for isotropic reconstruction of 3D microscopy data that can quickly produce images at arbitrary output resolutions. The representation embeds a learned latent code within a neural field that describes the implicit higher-resolution isotropic image region. We use a novel attention-guided latent interpolation approach, which allows flexible information exchange over a local latent neighborhood. Under isotropic volume assumptions, we self-supervise this representation on low-/high-resolution lateral image pairs to reconstruct an isotropic volume from low-resolution axial images. We evaluate our method on simulated and real anisotropic electron (EM) and light microscopy (LM) data. Compared to a state-of-the- art diffusion-based method, niiv shows improved reconstruction quality (+1 dB PSNR) and is over three orders of magnitude faster (1,000x) to infer. Specifically, niiv reconstructs a 1283 voxel volume in 2/10th of a second, renderable at varying (continuous) high resolutions for display.

Distinct basal ganglia decision dynamics under conflict and uncertainty

Ging-Jehli, N. R. — 2024-09-14

The basal ganglia (BG) play a key role in decision-making, preventing impulsive actions in some contexts while facilitating fast adaptations in others. The specific contributions of different BG structures to this nuanced behavior remain unclear, particularly under varying situations of noisy and conflicting information that necessitate ongoing adjustments in the balance between speed and accuracy. Theoretical accounts suggest that dynamic regulation of the amount of evidence required to commit to a decision (a dynamic "decision boundary") may be necessary to meet these competing demands. Through the application of novel computational modeling tools in tandem with direct neural recordings from human BG areas, we find that neural dynamics in the theta band manifest as variations in a collapsing decision boundary as a function of conflict and uncertainty. We collected intracranial recordings from patients diagnosed with either Parkinsons disease (n=14) or dystonia (n=3) in the subthalamic nucleus (STN), globus pallidus internus (GPi), and externus (GPe) during their performance of a novel perceptual discrimination task in which we independently manipulated uncertainty and conflict. To formally characterize whether these task and neural components influenced decision dynamics, we leveraged modified diffusion decision models (DDMs). Behavioral choices and response time distributions were best characterized by a modified DDM in which the decision boundary collapsed over time, but where the onset and shape of this collapse varied with conflict. Moreover, theta dynamics in BG structures predicted the onset and shape of this collapse but differentially across task conditions. In STN, theta activity was related to a prolonged decision boundary (indexed by slower collapse and therefore more deliberate choices) during high-conflict situations. Conversely, rapid declines in GPe theta during low conflict conditions were related to rapidly collapsing boundaries and expedited choices, with additional complementary decision bound adjustments during high uncertainty situations. Finally, GPi theta effects were uniform across conditions, with increases in theta prolonging the collapse of decision bounds. Together, these findings provide a nuanced understanding of how our brain thwarts impulsive actions while nonetheless enabling behavioral adaptation amidst noisy and conflicting information.

Practice Reshapes the Geometry and Dynamics of Task-tailored Representations

Kikumoto, A. — 2024-09-15

Extensive practice makes task performance more efficient and precise, leading to automaticity. However, theories of automaticity differ on which levels of task representations (e.g., low-level features, stimulus-response mappings, or high-level conjunctive memories of individual events) change with practice, despite predicting the same pattern of improvement (e.g., power law of practice). To resolve this controversy, we built on recent theoretical advances in understanding computations through neural population dynamics. Specifically, we hypothesized that practice optimizes the neural representational geometry of task representations to minimally separate the highest-level task contingencies needed for successful performance. This involves efficiently reaching conjunctive neural states that integrate task-critical features nonlinearly while abstracting over non-critical dimensions. To test this hypothesis, human participants (n = 40) engaged in extensive practice of a simple, context-dependent action selection task over 3 days while recording EEG. During initial rapid improvement in task performance, representations of the highest-level, context-specific conjunctions of task-features were enhanced as a function of the number of successful episodes. Crucially, only enhancement of these conjunctive representations, and not lower-order representations, predicted the power-law improvement in performance. Simultaneously, over sessions, these conjunctive neural states became more stable earlier in time and more aligned, abstracting over redundant task features, which correlated with offline performance gain in reducing switch costs. Thus, practice optimizes the dynamic representational geometry as task-tailored neural states that minimally tesselate the task space, taming their high-dimensionality.

Optimal transport reveals dynamic gene regulatory networks via gene velocity estimation

Zhao, W. — 2024-09-16

Inferring gene regulatory networks from gene expression data is an important and challenging problem in the biology community. We propose OTVelo, a methodology that takes time-stamped single-cell gene expression data as input and predicts gene regulation across two time points. It is known that the rate of change of gene expression, which we will refer to as gene velocity, provides crucial information that enhances such inference; however, this information is not always available due to the limitations in sequencing depth. Our algorithm overcomes this limitation by estimating gene velocities using optimal transport. We then infer gene regulation using time-lagged correlation and Granger causality via regularized linear regression. Instead of providing an aggregated network across all time points, our method uncovers the underlying dynamical mechanism across time points. We validate our algorithm on 13 simulated datasets with both synthetic and curated networks and demonstrate its efficacy on 4 experimental data sets. Author summaryUnderstanding how genes interact to regulate cellular functions is crucial for advancing our knowledge of biology and disease. We present OTVelo, a method that uses single-cell gene expression data collected at different time points to infer gene regulatory networks. OTVelo offers a dynamic view of how gene interactions change over time, providing deeper insights into cellular processes. Unlike traditional methods, OTVelo captures temporal information through ancestor-descendant transitions without assuming a specific underlying regulatory model. We validate our approach using both simulated and real-world data, demonstrating its effectiveness in revealing complex gene regulation patterns. This method could lead to new discoveries in understanding biological systems and developing disease treatments.

A mosaic of whole-body representations in human motor cortex

Deo, D. R. — 2024-09-16

Understanding how the body is represented in motor cortex is key to understanding how the brain controls movement. The precentral gyrus (PCG) has long been thought to contain largely distinct regions for the arm, leg and face (represented by the "motor homunculus"). However, mounting evidence has begun to reveal a more intermixed, interrelated and broadly tuned motor map. Here, we revisit the motor homunculus using microelectrode array recordings from 20 arrays that broadly sample PCG across 8 individuals, creating a comprehensive map of human motor cortex at single neuron resolution. We found whole-body representations throughout all sampled points of PCG, contradicting traditional leg/arm/face boundaries. We also found two speech-preferential areas with a broadly tuned, orofacial-dominant area in between them, previously unaccounted for by the homunculus. Throughout PCG, movement representations of the four limbs were interlinked, with homologous movements of different limbs (e.g., toe curl and hand close) having correlated representations. Our findings indicate that, while the classic homunculus aligns with each areas preferred body region at a coarse level, at a finer scale, PCG may be better described as a mosaic of functional zones, each with its own whole-body representation.

Reduced Hydrogen Sulfide Bioavailability Contributes to Cardiometabolic Heart Failure with Preserved Ejection Fraction

Doiron, J. — 2024-09-19

BackgroundHeart failure with preserved ejection fraction (HFpEF) is a significant public health concern with limited treatment options. Dysregulated nitric oxide-mediated signaling has been implicated in HFpEF pathophysiology, however, little is known about the role of endogenous hydrogen sulfide (H2S) in HFpEF. ObjectivesThis study evaluated H2S bioavailability in patients and two animal models of cardiometabolic HFpEF and assessed the impact of H2S on HFpEF severity through alterations in endogenous H2S production and pharmacological supplementation. We also evaluated the effects of the H2S donor, diallyl trisulfide (DATS) in combination with the GLP-1/glucagon receptor agonist, survodutide, in HFpEF. MethodsHFpEF patients and two rodent models of HFpEF ("two-hit" L-NAME + HFD mouse and ZSF1 obese rat) were evaluated for H2S bioavailability. Two cohorts of two-hit mice were investigated for changes in HFpEF pathophysiology: (1) endothelial cell cystathionine-{gamma}-lyase (EC-CSE) knockout; (2) H2S donor, JK-1, supplementation. DATS and survodutide combination therapy was tested in ZSF1 obese rats. ResultsH2S levels were significantly reduced (i.e., 81%) in human HFpEF patients and in both preclinical HFpEF models. This depletion was associated with reduced CSE expression and activity, and increased SQR expression. Genetic knockout of H2S -generating enzyme, CSE, worsened HFpEF characteristics, including elevated E/e ratio and LVEDP, impaired aortic vasorelaxation and increased mortality. Pharmacologic H2S supplementation restored H2S bioavailability, improved diastolic function and attenuated cardiac fibrosis corroborating an improved HFpEF phenotype. DATS synergized with survodutide to attenuate obesity, improve diastolic function, exercise capacity, and reduce oxidative stress and cardiac fibrosis. ConclusionsH2S deficiency is evident in HFpEF patients and conserved across multiple preclinical HFpEF models. Increasing H2S bioavailability improved cardiovascular function, while knockout of endogenous H2S production exacerbated HFpEF pathology and mortality. These results suggest H2S dysregulation contributes to HFpEF and increasing H2S bioavailability may represent a novel therapeutic strategy for HFpEF. Furthermore, our data demonstrate that combining H2S supplementation with GLP-1/glucagon receptor agonist may provide synergistic benefits in improving HFpEF outcomes. HighlightsO_LIH2S deficiency is evident in both human HFpEF patients and two clinically relevant models. C_LIO_LIReduced H2S production by CSE and increased metabolism by SQR impair H2S bioavailability in HFpEF. C_LIO_LIPharmacological H2S supplementation improves diastolic function and reduces cardiac fibrosis in HFpEF models. C_LIO_LITargeting H2S dysregulation presents a novel therapeutic strategy for managing HFpEF. C_LIO_LIH2S synergizes with GLP-1/glucagon agonist and ameliorates HFpEF C_LI

Is there a ubiquitous spectrolaminar motif of local field potential power across primate neocortex?

Mackey, C. A. — 2024-09-19

Mendoza-Halliday, Major et al.1 (referred to as "Mendoza-Halliday et al." for brevity), advocate for a local field potential (LFP)-based approach to functional identification of cortical layers during "laminar" (simultaneous recordings from all cortical layers) multielectrode recordings in nonhuman primates (NHPs). They describe a "ubiquitous spectrolaminar motif" in the primate neocortex: 1) 75-150 Hz power peaks in the supragranular layers, 2) 10-19 Hz power peaks in the infragranular layers and 3) the crossing point of their laminar power gradients identifies Layer 4 (L4). Identification of L4 is critical in general, but especially for Mendoza-Halliday et al. as the "motif" discovery is couched within a framework whose central hypothesis is that gamma activity originates in the supragranular layers and reflects feedforward activity, while alpha-beta activity originates in the infragranular layers and reflects feedback activity. In an impressive scientific effort, Mendoza-Halliday et al. analyzed laminar data from 14 cortical areas in 2 prior macaque studies and compared them to marmoset, mouse, and human data to further bolster the canonical nature of the motif. Identification of such canonical principles of brain operation is clearly a topic of broad scientific interest. Similarly, a reliable online method for L4 identification would be of broad scientific value for the rapidly increasing use of laminar recordings using numerous evolving technologies. Despite Mendoza-Halliday et al.s papers strengths, and its potential for scientific impact, a series of concerns that are fundamental to the analysis and interpretation of laminar activity profile data in general, and local field potential (LFP) signals in particular, led us to question its conclusions. Here, we address four key questions: Q1) Is the spectrolaminar motif ubiquitous, i.e. "found everywhere" or "universal"? Q2) Do features of the motif reliably identify Layer (L) 4? Q3) Are Mendoza-Halliday et al.s newly introduced methods (FLIP and vFLIP) reliable? And Q4) Are the proposed biophysical mechanisms underlying the motif well justified? We used new sets of data comprised of stimulus-evoked laminar response profiles from primary and higher-order auditory cortices (A1 and belt cortex), and primary visual cortex (V1) to test these questions. The rationale for using these areas as a test bed for new methods is that, in contrast to higher-order cortical areas, their laminar anatomy and physiology have already been extensively characterized by prior studies, and there is general agreement across laboratories on key matters like L4 identification. In short, we find that Mendoza-Halliday et al.s findings do not generalize well to these cortical areas. Specifically, regarding Q1: Though we can find a spectrolaminar gradient that is qualitatively consistent with Mendoza-Halliday et al., it is quantifiable in only 61-64% of our recordings, indicating that the motif is common but by no means universal (see "Evaluation of the FLIP method [...]" and "Evaluation of vFLIP [...]"). Regarding Q2: The motifs high/low frequency gradient cross point identified L4 in only 29-33% of our recordings. Regarding Q3: FLIP and vFLIP exhibit marked variability across studies, across brain areas, and spuriously detect cortical layer inversions (see "FLIPs fitting process [...]" and "Evaluation of vFLIP [...]"). Regarding Q4: the biophysical modeling findings cited to support Mendoza-Hallidays conclusions (see "Going forward - in silico [...]") do not reproduce the LFP data trends. While our findings are in many respects at odds with those of Mendoza-Halliday et al., the paper already has, and will continue to spark debate and further experimentation. Hopefully this countervailing presentation will lead to robust collegial efforts to define optimal strategies for applying laminar recording methods in future studies.

Mammalian Retinal Bipolar Cells: Morphological Identification and Systematic Classification in Rabbit Retina, with a Comparative Perspective

Famiglietti, E. V. — 2024-09-20

Retinal bipolar cells (BCs) convey visual signals from photoreceptors to more than 50 types of rabbit retinal ganglion cells (Famiglietti, 2020). More than 40 years ago, 10-11 types of bipolar cell were recognized in rabbit and cat retinas (Famiglietti, 1981). Twenty years later 10 were identified in mouse, rat, and monkey, while recent molecular genetic studies indicate that there are 15 types of bipolar cell in mouse retina (Shekhar et al., 2016). The present detailed study of more than 800 bipolar cells in ten Golgi-impregnated rabbit retinas indicates that there are 14-16 types of cone bipolar cell and one type of rod bipolar cell in rabbit retina. These have been carefully analyzed in terms of dendritic and axonal morphology, and axon terminal stratification with respect to fiducial starburst amacrine cells. In fortuitous proximity, several types of bipolar cell can be related to identified ganglion cells by stratification and by contacts suggestive of synaptic connection. These results are compared with other studies of rabbit bipolar cells. Homologies with bipolar cells of mouse and monkey are considered in functional terms.

Visitor effect on the behavior of chimpanzees (pan troglodyte) at a primate rescue center

Maurer, S. — 2024-09-23

1.The objective of this study was to examine the impact of visitors on the behavior of chimpanzees at a sanctuary. We hypothesized there wouldnt be an increase in abnormal, agnostic, or self-directed behaviors during visits, nor a decrease in affiliative behaviors when visitors are present. The study examined the effects of visitor presence on chimpanzee behavior at Fundacio MONA, a rescue center. Key findings includeO_LINo significant changes in abnormal or self-directed behaviors or agonistic behaviors were observed with visitor presence. C_LIO_LIAffiliative behaviors (excluding grooming) showed a slight decrease during visits. C_LI These results indicate that guided visitor interactions do not adversely affect chimpanzee behavior and may even enhance their welfare. The study supports the implementation of structured visitor programs for public education and funding, without compromising animal well-being. This contrasts with some previous research in zoo settings, suggesting that controlled visits can be beneficial in primate sanctuaries.

Evolutionary trajectories of β-lactam resistance in Enterococcus faecalis strains

Ugalde-Silva, P. — 2024-09-23

Resistance to ampicillin and imipenem in Enterococcus faecalis is infrequent. However, the evolution of resistance can occur through prolonged antibiotic exposure during the treatment of chronic infections. In this study, we conducted a Long-Term Evolution Experiment (LTEE) using four genetically diverse strains of E. faecalis with varying susceptibilities to ampicillin and imipenem. Each strain was subjected to increasing concentrations of either ampicillin or imipenem over 200 days, with three independent replicates for each strain. Selective pressure from imipenem led to the rapid selection of highly resistant lineages across all genetic backgrounds, compared to ampicillin. In addition to high resistance, we describe, for the first time, the evolution of a {beta}-lactam dependent phenotype observed in lineages from all backgrounds. WGS and bioinformatic analysis revealed mutations in three main functional classes: genes involved in cell wall synthesis and degradation, genes in the walK/R two-component system, and genes in the c-di-AMP pathway. Our analysis identified new mutations in genes known to be involved in resistance as well as novel genes potentially associated with resistance. Furthermore, the newly described {beta}-lactam dependent phenotype was correlated with the inactivation of c-di-AMP degradation, resulting in high levels of this second messenger. Together, these data highlight the diverse genetic mechanisms underlying resistance to ampicillin and imipenem in E. faecalis. The emergence of high resistance levels and {beta}-lactam dependency underscores the importance of understanding evolutionary dynamics in the development of antibiotic resistance. ImportanceE. faecalis is a major human pathogen, and treatment is frequently compromised by poor response to first-line antibiotics such ampicillin. Understanding the factors that play a role in susceptibility/resistance to these drugs will help guide the development of much needed treatments.

Plasticity in the pectoral fin skeleton is induced by altered foraging regime in a South American cichlid

Gilbert, M. — 2024-09-24

The fins of fishes are remarkably diverse, and this variation is tied to the ecology and locomotor mode of a species. While numerous genetic factors are known to pattern fins in development, it is unclear how developmental plasticity shapes the fin skeleton. Here, we analyze the cichlid Satanoperca daemon, raised under three distinct feeding regimes, and show that plasticity is pervasive across the pectoral fin skeleton with foraging mode impacting patterning of both the endoskeleton and dermal skeleton. Radials and fin rays were {micro}CT scanned and analyzed using a combination of linear measures and geometric morphometrics. Anteroposterior patterning of both radials and fin rays are affected by feeding regime. Notably, S. daemon pectoral fin rays show distinct patterns of fin ray branching between treatments, suggesting altered fin stiffness. We argue that the observed changes in the fin likely reflect developmental plasticity resultant from altered swimming behaviors when fishes are challenged to forage in different ways. These data show how non-genetic mechanisms can shape both the endoskeleton and dermal skeleton of fins, and that foraging mode can induce plastic changes in skeletal elements that do not directly interface with food items.

SARS-CoV-2 ORF3a Protein Impairs Syncytiotrophoblast Maturation, Alters ZO-1 Localization, and Shifts Autophagic Pathways in Trophoblast Cells and 3D Organoids

Kumar, D. — 2024-09-25

SARS-CoV-2 infection poses a significant risk to placental physiology, but its impact on placental homeostasis is not well understood. We and others have previously shown that SARS-CoV-2 can colonize maternal and fetal placental cells, yet the specific mechanisms remain unclear. In this study, we investigate ORF3a, a key accessory protein of SARS-CoV-2 that exhibits continuous mutations. Our findings reveal that ORF3a is present in placental tissue from pregnant women infected with SARS-CoV-2 and disrupts autophagic flux in placental cell lines and 3D stem-cell-derived trophoblast organoids (SC-TOs), impairing syncytiotrophoblast differentiation and trophoblast invasion. This disruption leads to protein aggregation in cytotrophoblasts (CTB) and activates secretory autophagy, increasing CD63+ extracellular vesicle secretion, along with ORF3a itself. ORF3a also compromises CTB barrier integrity by disrupting tight junctions via interaction with ZO-1, mediated by its PDZ-binding motif, SVPL. Colocalization of ORF3a and ZO-1 in SARS-CoV-2-infected human placental tissue supports our in vitro findings. Deleting the PDZ binding motif in the ORF3a protein (ORF3a-noPBM mutant) restored proper ZO-1 localization at the cell junctions in an autophagy-independent manner. Lastly, we demonstrate that constitutive ORF3a expression induces SC-TOs to transition towards a secretory autophagy pathway likely via the PBM motif, as the ORF3a-NoPBM mutants showed a significant lack of CD63 expression. This study demonstrates the functional impact of ORF3a on placental autophagy and reveals a new mechanism for the activation of secretory autophagy, which may lead to increased extracellular vesicle secretion. These findings provide a foundation for exploring therapeutic approaches targeting ORF3a, specifically focusing on its PBM region to block its interactions with host cellular proteins and limiting placental impact.

Non-Optical, Label-free Electrical Capacitance Imaging of Microorganisms

Incandela, J. T. — 2024-09-25

Many fundamental insights into microbiology have come from imaging, which is typically synonymous with optical techniques. However, the sample preparation needed for many optical microscopy methods such as labeling, fixing, or genetic modification, limits the range of species and environments we can investigate. Here we demonstrate the use of electrical capacitance measurements as a non-optical method for imaging live microbial samples. In electrical capacitance imaging (ECI), samples are positioned in contact with a semiconductor sensor array, and localized capacitance measurements are made across the array. From these measurements, we generate textured images of a variety of microbial colonies. We determine that capacitance is correlated with local sample thickness by comparing ECI data to 3D confocal scans. We further illustrate with ECI that a difference in capacitance signal allows microbial species to be spatially distinguished in co-culture conditions. In order to highlight the versatility of our system, we capture the cross-sectional development of floating pellicle biofilms in a liquid culture at millimeter length scales during weeks-long time-lapse experiments. These novel results establish a new low-cost and portable platform which can be used for spatially and temporally resolved experiments in diverse environments with a wide variety of microbial species. IMPORTANCEMicrobes live in diverse environments, and occupy biological roles across many timescales. Investigating the full scope of microbial activity requires imaging systems appropriate to each context. Though optical microscopy is powerful, the use of light, lenses, and other hardware limits where it can be applied. At the same time, existing non-optical imaging methods are frequently destructive to samples and require extensive equipment. In this paper we present a non-optical imaging system that is small, cheap, requires no sample labeling, and is compatible with a variety of microbial species. Our system uses semiconductor chips to measure the inherent material properties of a sample with spatial sensitivity, producing images of microbes contrasted against their environment and each other. Our technique captures label-free, micrometer-resolution images with a pocket-sized device, enabling microbiological imaging experiments in new environments with new species.

Combination of Imipridone ONC201 or ONC206 with Temozolomide and Radiotherapy in triple ITR therapy reduces intracranial tumor burden and prolongs survival in an orthotopic wild-type IDH GBM mouse model

Zhou, L. — 2024-09-27

Glioblastoma remains the most lethal common primary brain tumor in adults with limited therapeutic options. TIC10/ONC201, a first-in-class imipridone we discovered, achieved meaningful therapeutic effects in phase I/II trials in patients with diffuse gliomas harboring H3K27M mutations, and currently the drug is in randomized phase III testing (ACTION trial; NCT05580562). ONC201 targets mitochondrial protease ClpP to disrupt oxidative phosphorylation and trigger the integrated stress response (ISR), TRAIL/DR5, and tumor cell death. We hypothesized that ONC201 and its analogue ONC206 synergize with temozolomide (TMZ) and ionizing radiation (IR), standard-of-care glioblastoma therapies. ONC201 enhances TMZ or IR-induced apoptosis, and cytotoxicity. ClpP-silencing suppresses ONC201-induced cytotoxicity but not TMZ or RT. Both ONC201 and ONC206 reduce expression of TMZ-resistance mediator MGMT. Suppression of MGMT protein was observed in H3K27M-mutated DIPG cell lines following treatment with ONC201 or ONC206 with or without TMZ. Cytokine profiling indicates distinct ONC201 alterations relative to TMZ suggesting distinct anti-tumor immune mechanisms. Triple IR+TMZ+ONC201 (ITR) therapy prolongs median survival to 123 days with a tail on survival curve (3-of-7 mice alive beyond 200-days) in an orthotopic U251 GBM model versus ONC201 (44-days; p=0.000197), IR (63-days; p=0.0012), TMZ (78-days; p=0.0354), ONC201+IR (55-days; p=0.0004), ONC201+TMZ (80-days; p=0.0041) and IR+TMZ (103-days; p>0.05). By 231-days, the only surviving mice were in IRT group. Our results support investigation of ONC201/ONC206 in combination with TMZ and IR (ITR) in GBM or H3K27M mutated diffuse glioma therapy.

Cyclooxygenase-2 disruption predisposes to heart failure with preserved ejection fraction

Ricciotti, E. — 2024-09-30

Heart failure (HF) is one of the most strongly associated adverse cardiovascular events linked to the use of cyclooxygenase (COX)-2 selective and non-selective nonsteroidal anti-inflammatory drug (NSAID). Nevertheless, it remains uncertain whether NSAID exposure is more likely to lead to heart failure with reduced ejection fraction (HFrEF) or preserved ejection fraction (HFpEF). In adult mice, postnatal genetic deletion or pharmacological inhibition of COX-2 did not affect cardiac function. In contrast, aged female inducible COX-2 (iCOX-2) knockout (KO) mice displayed diastolic dysfunction, cardiac hypertrophy, pulmonary congestion, and elevated levels of plasma N-terminal pro B-type natriuretic peptide (BNP) when compared to age- and sex- matched controls, while their ejection fraction (EF) remained preserved ([≥] 50%). No such phenotype was observed in aged male iCox-2 KO mice. Aged female iCox-2 KO mice showed a shift from prostanoid to leukotriene biosynthesis, along with changes in the expression of mitochondrial genes and calcium-handling proteins in the myocardium. The ratio of phospholamban to SERCA2a was increased, indicating an inhibitory effect on SERCA2a activity, which may contribute to impaired myocardial relaxation. In larval zebrafish, COX-2 inhibition by celecoxib caused a modest yet significant reduction in heart rate and diastolic function, while EF was preserved. Additionally, celecoxib increased BNP expression and ventricular calcium transient amplitude. Diabetic patients in the Harvard-Partners electronic medical record exposed to NSAIDs selective for COX-2 inhibition were more strongly associated with an increased risk of HFpEF compared to HFrEF. Collectively, these findings indicate that COX-2 deletion or inhibition does not impair systolic cardiac function but instead leads to an HFpEF phenotype in mice, zebrafish, and humans. An imbalance in calcium handling may mediate the impairment of myocardial relaxation following COX-2 suppression. SummaryGenetic deletion or pharmacological inhibition of COX-2 results in heart failure with preserved ejection fraction across zebrafish, mice, and humans.

Attenuated ectopic action potential firing in parvalbumin expressing interneurons in a mouse model of Dravet Syndrome

Hill, S. F. — 2024-10-02

Dravet syndrome is caused by heterozygous loss-of-function variants in SCN1A, which encodes the voltage-gated sodium channel Nav1.1. Our recent work suggests that a primary pathogenic mechanism of Dravet syndrome is impaired action potential propagation along axons of cortical parvalbumin-positive fast-spiking GABAergic interneurons (PVINs). Ectopic action potentials (EAPs) are action potentials that initiate distal to the axon initial segment. We recently demonstrated that a large proportion of PVINs fire EAPs during periods of increased excitation. Although their function remains unknown, EAPs may play a role in amplifying (when occurring in excitatory cells) and/or preventing seizures (when occurring in interneurons). Regardless of function, their generation in distal axons suggests that EAP frequency could be a useful proxy for distal axonal excitability. We hypothesized that EAPs are attenuated in PVINs from Dravet syndrome (Scn1a+/-) mice due to dysfunction of the distal axon. We induced EAPs in PVINs in acute brain slices prepared from male and female wildtype (WT) and Scn1a+/- mice at P18-21 and P35-56, when we have previously identified axonal conduction deficits in Scn1a+/- PVINs. We elicited EAPs in 17/22 (77%) of WT PVINs, including 6 (22%) that exhibited barrages of EAPs. In contrast, Scn1a+/- PVINs never fired barrages (0%), and only 8/23 (34%) exhibited even single EAPs. This finding adds to the body of evidence supporting impaired action potential propagation in Dravet syndrome PVINs, and is the first evidence of impaired EAP firing in a disease model, suggesting that dysregulation of EAPs could be involved in the pathophysiology of human disease.

A novel behavioral apparatus for spontaneous exploration and operant conditioning of social information under spatial conditions in rats

Wise, T. B. — 2024-10-03

BackgroundEvidence from the fields of evolutionary biology and neuroscience supports the theory that spatial cognition and social cognition share neural mechanisms. Rodent models are widely used to study either spatial or social cognition, but few studies have explored the interactions between the spatial and social cognitive domains due to the lack of appropriate paradigms. New methodOur study introduces the Vertical Maze (VM), a novel behavioral apparatus designed to measure multiple aspects of spatial and social cognition. The VM features a standard 3-chamber maze positioned above multilevel columns allowing for the presentation of conspecifics at varying spatial distances and familiarity levels. This arrangement enables conspecifics to serve as discriminative stimuli for both social and spatial spontaneous and goal-oriented assessments. The three-dimensional design of the VM allows rats to use multisensory cues to judge distance, direction, and social identity of conspecifics. ResultsIn the present study, we found that rats can 1) discriminate the spatial distance of conspecifics located below them in an operant conditioning task, and 2) discriminate social novelty when conspecifics are presented below at the near, middle, and far distances in a spontaneous exploration task. Critically, it was necessary for rats to explore all levels of the maze to perform these discriminations. Comparison with existing methodsThis new method advances the field by permitting the presentation of social information (conspecifics) at different spatial distances. The use of conspecifics to serve as stimuli for both social and spatial discriminations allows more direct comparison of behavioral measures across these information domains. Importantly, the presentation of conspecifics as stimuli below the 3-chamber level of the maze engages auditory, visual, and olfactory systems, encouraging a robust multisensory representation of conspecifics presented at a distance. ConclusionsOur results confirm that the VM is an effective tool for studying both spatial and social cognition, facilitating the development of novel automated tasks in these areas. This new method opens new avenues for investigating the neural and cognitive foundations of spatial and social behavior, as well as for exploring the possibility of shared mechanisms across these cognitive domains.

Onco-fetal protein Nogo-A restricts human and mouse glioma vascularization and growth via VEGF-Notch-hippo-metabolic signaling

Schwab, M. — 2024-10-03

Glioblastoma is one of the most deadly human cancers characterized by high degrees of vascularization, but targeting its vasculature has resulted in very limited success so far. Angiogenesis, the growth of new blood vessels, is highly dynamic during brain development, enters a mostly quiescent state in the adult homeostatic brain, and is reactivated in vascular-dependent CNS diseases including brain tumors. In consequence, a better understanding of the relevance of the onco-fetal axis - describing the reactivation of fetal signaling programs in tumors - in endothelial- and perivascular cells of the human brain tumor vasculature harbors great translational potential, yet remains poorly defined. In development, neurovascular link (NVL) molecules guide both neuronal growth cones as well as capillary endothelial tip cells. Nogo-A is an NVL molecule known to inhibit axonal growth in the developing and adult CNS and to restrict angiogenesis during brain development, but its role in the mouse and human brain tumor vasculature along the onco-fetal axis remains unknown. Here, we characterize Nogo-A as an onco-fetal protein expressed in the neurovascular unit (NVU) in human fetal brains and human gliomas in vivo that negatively regulates sprouting angiogenesis and endothelial tip cells in glioma vascularization. The Nogo-A-specific Delta 20 domain restricts angiogenic sprouting and branching and promotes vascular normalization while inhibiting glioma growth in experimental gliomas. Moreover, Nogo-A expression in tumor cells negatively correlate/s with glioma malignancy in vivo. In vitro, Nogo-A Delta 20 reduced human brain- and brain tumor endothelial cell (HBMVEC, HBTMVEC) and human umbilical vein endothelial cell (HUVEC) spreading, migration, and sprouting, in a dose-dependent manner and inhibited filopodia extension glucose metabolism. Mechanistically, RNA sequencing of Nogo-A Delta 20-treated HBMVECs and HBTMVECs revealed Nogo-A Delta 20-induced positive regulation of the angiogenesis-inhibiting Dll4-Notch-pathway and inhibition of the angiogenesis-promoting VEGF-VEGFR and Hippo-YAP-TAZ pathways, whereas metabolomics and functional metabolic assays revealed Nogo-A Delta 20-induced negative regulation of endothelial glycolysis in HBMVECs and HBTMVECs. These findings characterize Nogo-A as an onco-fetal protein in the human glial brain tumor vasculature and identify Nogo-A Delta 20 signaling as an important negative regulator of human glioma vascularization and growth. Enhancing Nogo-A signaling may be an attractive alternative or combinatorial anti-angiogenic therapy to restrict human glioma/glioblastoma vascularization and growth.

cGAS deficient mice display premature aging associated with de-repression of LINE1 elements and inflammation

Martinez, J. C. — 2024-10-11

Aging-associated inflammation, or inflammaging" is a driver of multiple age-associated diseases. Cyclic GMP-AMP Synthase (cGAS) is a cytosolic DNA sensor that functions to activate interferon response upon detecting viral DNA in the cytoplasm. cGAS contributes to inflammaging by responding to endogenous signals such as damaged DNA or LINE1 (L1) cDNA which forms in aged cells. While cGAS knockout mice are viable their aging has not been examined. Unexpectedly, we found that cGAS knockout mice exhibit accelerated aging phenotype associated with induction of inflammation. Transcription of L1 elements was increased in both cGAS knockout mice and in cGAS siRNA knockdown cells associated with high levels of cytoplasmic L1 DNA and expression of ORF1 protein. Cells from cGAS knockout mice showed increased chromatin accessibility and decreased DNA methylation on L1 transposons. Stimulated emission depletion microscopy (STED) showed that cGAS forms nuclear condensates that co-localize with H3K9me3 heterochromatin marks, and H3K9me3 pattern is disrupted in cGAS knockout cells. Taken together these results suggest a previously undescribed role for cGAS in maintaining heterochromatin on transposable elements. We propose that loss of cGAS leads to loss of chromatin organization, de-repression of transposable elements and induction of inflammation resulting in accelerated aging.

Ramping dissociates motor and cognitive sequences in the parietal and prefrontal cortices

Doyle, H. — 2024-10-12

Humans complete different types of sequences as a part of everyday life. These sequences can be divided into two important categories: those that are abstract, in which the steps unfold according to a rule at super-second to minute time scale, and those that are motor, defined solely by individual movements and their order which unfold at the sub-second to second timescale. For example, the sequence of making spaghetti consists of abstract tasks (preparing the sauce and cooking the noodles) and nested motor actions (stir pasta water). Previous work shows neural activity increases (ramps) in the rostrolateral prefrontal (RLPFC) during abstract sequence execution (Desrochers et al., 2015, 2019). During motor sequence production, activity occurs in regions of the prefrontal cortex (Yewbrey et al., 2023). However, it remains unknown if ramping is a signature of motor sequence production as well or solely an attribute of abstract sequence monitoring and execution. We tested the hypothesis that significant ramping activity occurs during motor sequence production in the RLPFC. Contrary to our hypothesis, we did not observe significant ramping activity in the RLPFC during motor sequence production, but we found significant ramping activity in bilateral inferior parietal cortex, in regions distinct from those observed during an abstract sequence task. Our results suggest different prefrontal-parietal circuitry may underlie abstract versus motor sequence execution.

TE-Seq: A Transposable Element Annotation and RNA-Seq Pipeline

Kelsey, M. M. G. — 2024-10-15

The recognition that transposable elements (TEs) play important roles in many biological processes has elicited growing interest in analyzing sequencing data derived from this mobile dark genome. The TE-Seq pipeline conducts an end-to-end analysis of RNA-sequencing data, examining both genes and TEs. It implements the most current computational methods tailor-made for TEs, enabling a comprehensive analysis of TE expression at both the individual element level and at the TE clade level. If supplied with long-read DNA sequencing data, it creates a TE-complete genome incorporating non-reference (polymorphic) TE-loci, enabling the functional characterization of the evolutionarily youngest mobile elements in the genome.

Information transfer from spatial to social distance in rats: implications for the role of the posterior parietal cortex in spatial-social integration

Wise, T. B. — 2024-10-16

Humans and other social animals can represent and navigate complex networks of social relationships in ways that are suggestive of representation and navigation in space. There is some evidence that cortical regions initially required for processing space have been adapted to include processing of social information. One candidate region for supporting both spatial and social information processing is the posterior parietal cortex (PPC). We examined the hypothesis that rats can transfer or generalize distance information across spatial and social domains and that this phenomenon requires the PPC. In a novel apparatus, rats learned to discriminate two conspecifics positioned at different spatial distances (near vs. far) in a goal-driven paradigm. Following spatial learning, subjects were tested on probe trials in which spatial distance was replaced with social distance (cagemate vs. less familiar conspecific). The PPC was chemogenetically inactivated during a subset of probe sessions. We predicted that, in control probe trials, subjects would select conspecifics whose social distance matched the previously learned spatial distance. That is, if trained on the near distance, the rat would choose the highly familiar cagemate, and if trained on the far distance, the rat would choose the less familiar conspecific. Subjects learned to discriminate conspecifics based on spatial distance in our goal-driven paradigm. Moreover, choice for the appropriate social distance in the first probe session was significantly higher than chance. This result suggests that rats transferred learned spatial information to social contexts. Contrary to our predictions, PPC inactivation did not impair spatial to social information transfer. Possible reasons are discussed. To our knowledge, this is the first study to provide evidence that spatial and social distance are processed by shared cognitive mechanisms in the rat model.

Replicating retroviral delivery of an IL-15 superagonist improves antitumor immunity and long-term survival in poorly immunogenic glioblastoma models

Haddad, A. F. — 2024-10-17

Glioblastoma (GBM) is the most lethal primary brain neoplasm due to its highly immunosuppressive microenvironment and resistance to conventional therapies. To overcome this challenge, we engineered a replicating retrovirus (RRV) to deliver a superagonist interleukin-15 receptor-linked fusion protein (RLI) directly to tumor cells, engineering them into local immunotherapy biofactories. This strategy leverages the tumor-selective replication of RRV to achieve localized and sustained RLI expression within the tumor microenvironment. In two orthotopic poorly immunogenic GBM mouse models, intratumoral administration of RRV RLI significantly reduced tumor growth and prolonged survival compared to controls, with some mice achieving long-term remission and demonstrating immunologic memory upon rechallenge. Transcriptomic and flow cytometric analyses revealed that RRV RLI treatment enhanced infiltration and activation of CD8 T cells, NK cells, and upregulated antigen presentation pathways within the tumor microenvironment. Depletion studies indicated that the therapeutic efficacy of RRV RLI is dependent on both CD4 and CD8 T cells. Notably, combining RRV RLI with the GBM standard of care chemotherapeutic agent temozolomide (TMZ) synergistically improved survival outcomes. Subsequent single-cell RNA and T cell receptor sequencing identified enhanced effector cell activation, antigen presentation, and clonal T cell expansion in the combination therapy group. Further T cell receptor analysis and clustering implied a tumor-specific immune response rather than one targeting the viral delivery vehicle, suggesting that this therapeutic approach could be reapplied without eliciting anti-vector immunity. Our findings suggest that RRV-mediated delivery of RLI effectively transforms GBM tumors into immunostimulatory hubs, eliciting a potent anti-tumor immune response. This novel viral immunotherapy holds significant promise for clinical translation in the treatment of GBM and other difficult-to-treat solid tumors.

A Cortical Microcircuit for Region-Specific Credit Assignment in Reinforcement Learning

Chevy, Q. — 2024-10-17

The distributed architecture of the cortex poses a fundamental challenge for reinforcement learning: how to assign credit specifically to regions that contribute to successful behavior? Cortical neurons can be driven by both global reinforcers, like rewards, and local sensory features, making it difficult to disentangle these influences. To address this, we investigated cortical reinforcement learning by manipulating the reward-predictive sensory modality during learning tasks, while monitoring key regulators of cortical activity--local inhibitory neurons, and cholinergic inputs. We found that VIP interneurons are broadly recruited by reward-predictive cues via a modality-independent cholinergic signal. However, when task demands aligned with local computation, SST interneurons suppressed VIP recruitment through an inhibitory feedback loop. A computational model demonstrates that this cholinergic-VIP-SST interneuron circuit motif enables targeted reinforcement learning and region-specific credit assignment in the cortex. These results offer a neurobiologically-grounded framework for how the cortex uses global reinforcement signals to direct plasticity to task-relevant regions, enabling those regions to adapt and fine-tune their responses.

THRESHOLD: A Comprehensive Transcriptomic Analysis Tool for Evaluating Gene Saturation and Impact in Disease Progression

Gammell, F. M. — 2024-10-22

Gene expression studies serve as a foundational tool in molecular biology, providing insights into developmental, physiological, and pathological processes. Variations in gene expression can indicate disease states, which are vital in understanding disease progression, subtype manifestations, and identifying therapeutic targets based on detailed expression patterns. To effectively investigate gene expression patterns, especially in large datasets, a robust and precise analysis tool is crucial. In response to this critical analytical need, we developed THRESHOLD, a novel tool that goes beyond traditional gene expression analysis by introducing the concept of gene saturation. Unlike conventional methods that focus on absolute expression levels or binary differential expression, THRESHOLD quantifies the consistency of gene expression across patients, revealing co-regulation patterns that may otherwise be overlooked. This novel metric offers a unique perspective on gene expression patterns by highlighting consistent trends across patient samples, which are critical for understanding disease mechanisms and stratifying patients based on molecular signatures. The tool offers several features, including user-defined parameters, statistical comparisons, and interactive data visualization. THRESHOLD has uncovered compelling insights into disease progression using TCGA Cancer Datasets. For instance, bladder urothelial carcinoma demonstrated increasing upregulated gene saturation in progressive cancer stages (p < 0.00001). Moreover, THRESHOLD identified heightened gene saturation in patients with earlier onset of prostate adenocarcinoma (p < 0.0001) and revealed a critical fusion transcript, SLC45A2-AMACR, implicated in prostate adenocarcinoma progression, recurrence, and metastasis. Additionally, novel biomarkers and potential candidates for drug therapies were identified through protein-protein interaction networks and functional analyses of saturation data in colon adenocarcinoma and breast invasive carcinoma. Collectively, THRESHOLD advances our understanding of patient stratification and molecular signatures by offering a more detailed view of gene expression dynamics. The THRESHOLD tool is publicly available at: https://github.com/alperuzun/THRESHOLD.

Spatial memory in Alzheimer's disease 5XFAD mice is enhanced by XPO1 inhibitor KPT-330

Wong, S. Q. — 2024-10-22

The proteostatic decline in Alzheimers disease is well established and improvement in proteostasis could potentially delay cognitive impairment. One emerging entry point to modulate proteostasis is the regulation of nucleo-cytoplasmic partitioning of proteins across the nuclear pore via karyopherins. The nuclear exportin XPO1 is a key regulator of proteostasis by driving the assembly of ribosomes and by modulating the process of autophagy. We recently found that XPO1 inhibitor KPT-330 (Selinexor), an FDA approved drug against multiple myelomas, enhances proteostasis, leading to benefits in models of neurodegenerative diseases in C. elegans and Drosophila. Here, we find that KPT-330 increases autophagy in murine neuronal cells and improves spatial memory performance in a murine model of Alzheimers disease (5XFAD). Unexpectedly, general amyloid deposition in several brain regions was significantly increased by KPT-330, but specific regions, especially the thalamus, displayed significantly lower deposition, suggesting that XPO1 inhibition has regional-specific effects on proteostasis and amyloid plaque formation. Altogether, we conclude that XPO1 inhibition can improve cognition via spatially-specific reductions in amyloid deposition.

Convergent olfactory circuits for courtship in Drosophila revealed by ds-Tango

Fisher, J. D. — 2024-10-23

Animals exhibit sex-specific behaviors that are governed by sexually dimorphic circuits. One such behavior in male Drosophila melanogaster, courtship, is regulated by various sensory modalities, including olfaction. Here, we reveal how sexually dimorphic olfactory pathways in male flies converge at the third-order, onto lateral horn output neurons, to regulate courtship. To achieve this, we developed ds-Tango, a modified version of the monosynaptic tracing and manipulation tool trans-Tango. In ds-Tango, two distinct configurations of trans-Tango are positioned in series, thus providing selective genetic access not only to the monosynaptic partners of starter neurons but also to their disynaptic connections. Using ds-Tango, we identified a node of convergence for three sexually dimorphic olfactory pathways. Silencing this node results in deficits in sex recognition of potential partners. Our results identify lateral horn output neurons required for proper courtship behavior in male flies and establish ds-Tango as a tool for disynaptic circuit tracing.

Mobility of Four Structural Regions Drives Isoform-Specific Properties of Plant LPOR

Gabruk, M. — 2024-10-24

Light-dependent protochlorophyllide oxidoreductase (LPOR) is a photocatalytic enzyme in the chlorophyll (Chl) biosynthetic pathway that underwent duplications in angiosperms, resulting in the emergence of multiple isoforms across various plant species. The physiological roles of these LPOR homologs remained unclear, so we selected six plant species with different number of isoforms of the enzyme and characterized their properties in vitro. Our findings revealed that these isoforms vary in their affinity for the reaction product, chlorophyllide (Chlide), as well as for NADPH under lipid-free conditions and in reaction mixtures supplemented with plant lipids. Additionally, we observed differences in their oligomerization behavior. Our experimental approach generated a dataset comprising several hundred pairs of spectra, recorded before and after reaction-triggering illumination. This data was used to analyze the correlation between fluorescence emission maxima before and after photoconversion. The analysis showed that some isoforms rapidly release Chlide after the reaction, while others retain the pigment in the binding pocket, especially at high NADPH concentrations. These results suggest that LPOR isoforms differ in their rates of Chlide release and complex disassembly, potentially influencing the overall rate of the Chl biosynthetic pathway, even in mature leaves. We further analyzed the flexibility of these isoforms using AlphaFold2 predictions, identifying four regions of the enzyme that are particularly mobile. Two of these regions are involved in pigment binding, while the other two play a role in oligomerization. Based on these findings, we propose a model of conformational changes that drive the formation of LPOR oligomers.

LUNAR: A Deep Learning Model to Predict Glioma Recurrence Using Integrated Genomic and Clinical Data

Patricoski-Chavez, J. A. — 2024-10-24

Gliomas account for approximately 25.5% of all primary brain and central nervous system tumors, with a striking 80.8% of these being malignant. The prognosis varies significantly; low-grade gliomas (LGGs) can exhibit 5-year survival rates of up to 80%, while higher-grade gliomas (HGGs) often see rates below 5%. Recurrence is a common challenge, occurring in 52%-62% of LGGs and 90% of HGGs, complicating clinical management and treatment planning. Currently, no widely available models exist for predicting glioma recurrence, which is critical for optimizing patient outcomes. Machine learning (ML) and deep learning (DL) techniques have shown promise in predicting recurrence for various cancers, most using Electronic Health Records (EHR). This study introduces gLioma recUrreNce Attention-based classifieR (LUNAR), a DL-based model to predict early versus late glioma recurrence by integrating clinical, genomic, and mRNA expression data from patients with primary grade II-IV gliomas. The data was obtained from The Cancer Genome Atlas and the Glioma Longitudinal Analysis Consortium (GLASS).

Abl kinases regulate FGF signaling independent of Crk phosphorylation to prevent Peters anomaly

Wu, H. — 2024-10-26

Peters anomaly, the most common cause of congenital corneal opacity, stems from corneal-lenticular adhesion. Despite numerous identified mutations, a cohesive molecular framework of the diseases etiology remains elusive. Here, we identified Abl kinases as pivotal regulators of FGF signaling, as genetic ablation of Abl kinases restores lens induction even in the absence of FGF signaling. Intriguingly, both Abl kinase deficiency and increased FGF-Ras activity result in Peters anomaly independent of ERK signaling, which can be rescued by allelic deletion of Abl substrate, Crk. However, contrary to the prevailing belief that Abl kinases regulate Crk proteins by direct phosphorylation, mutations at Abl kinase phosphorylation sites on Crk and CrkL did not yield any observable effects. Instead, our findings reveal that Abl kinases phosphorylate Ptpn12, which in turn inhibits p130Cas phosphorylation and Crk recruitment, crucial for Rho GTPases activation and cytoskeletal dynamics. Consequently, Abl kinase deficiency reduces actomyosin contractility within the lens vesicle and genetically interacts with RhoA inhibition. Conversely, Rac1 deletion mitigates Peters anomaly in models with aberrant FGF, Abl kinase and RhoA signaling. Our results demonstrate that Abl kinases regulate FGF signaling to balance RhoA and Rac1 activity via the Ptpn12-p130Cas pathway, suggesting that targeting tension-mediated lens vesicle separation could be a therapeutic strategy for Peters anomaly.

Continuous integration of heading and goal directions guides steering

Crown, A. M. — 2024-10-26

Navigating animals must integrate a diverse array of sensory cues into a single locomotor decision. Insects perform intricate navigational feats using a brain region termed the central complex in which an animals heading direction is transformed through several layers of circuitry to elicit goal-directed locomotion. These transformations occur mostly in the fan-shaped body (FB), a major locus of multi-sensory integration in the central complex. Key aspects of these sensorimotor computations have been extensively characterized by functional studies, leveraging the genetic tools available in the fruit fly. However, our understanding of how neuronal activity in the FB dictates locomotor behaviors during navigation remains enigmatic. Here, we manipulate the activity of two key neuronal populations that input into the FB-the PFNa and PFNd neurons-used to encode the direction of two complex navigational cues: wind plumes and optic flow, respectively. We find that flies presented with unidirectional optic flow steer along curved walking trajectories, but silencing PFNd neurons abolishes this curvature. We next use optogenetic activation to introduce a fictive heading signal in the PFNs to establish the causal relationship between their activity and steering behavior. Our studies reveal that the central complex guides locomotion by summing the PFN-borne directional signals and shifting movement trajectories left or right accordingly. Based on these results, we propose a model of central complex-mediated locomotion wherein the fly achieves fine-grained control of sensory-guided steering by continuously integrating its heading and goal directions over time.

EEG-VLM Toolbox: Extending voxel-based lesion mapping to multi-dimensional EEG data

Hardstone, R. — 2024-10-26

Focal brain lesions (such as with stroke) cause functional changes in local and distributed neural systems. While there is a long history of post-stroke neurophysiological assessment using electroencephalography (EEG), the observed neurophysiological changes have rarely been related to specific lesion locations. Therefore, the relationships between anatomical injury and physiological changes after stroke remain unclear. Voxel-based lesion symptom mapping (VLSM) is a tool for statistically relating stroke lesion locations to "symptoms", but current VLSM methods are restricted to symptoms that can be defined by a single value. Therefore, current VLSM techniques are unable to map the relationships between anatomical injury and multidimensional neurophysiological data such as EEG, which contains rich spatio-temporal information across different channels and frequency bands. Here we present a novel algorithm, EEG Voxel-based Lesion Mapping (EEG-VLM), that produces the set of significant relationships between precise neuroanatomical injury locations and neurophysiology (defined by a cluster of adjacent EEG channels and frequency bands). Further, the algorithm provides statistical analyses to define the overall significance of each neural structure-function relationship by correcting for multiple comparisons using a permutation test. Applying EEG-VLM to a dataset of recordings from chronic stroke patients performing a cued upper extremity movement task, we found that subjects with lesions in frontal subcortical white matter have reduced ipsilesional parietal cue-evoked EEG responses. These results are consistent with damage to a frontal-parietal network that has been associated with impairments in attention. EEG-VLM is a novel and unbiased method for relating neurophysiologic changes after stroke with neuroanatomic lesions. In the context of focal brain lesions associated with neurological impairments, we propose that this method will enable improved mechanistic understanding, facilitate biomarker development, and guide neurorehabilitation strategies.

A comparative network analysis of high-risk MYCN amplified and high-risk MYCN non-amplified neuroblastoma patients

Chang, E. — 2024-10-29

Neuroblastoma is the most common extracranial solid tumor in children. Under contemporary staging guidelines, amplification of MYCN, defined as possessing 10 or more copies of the MYCN gene, is sufficient to classify a patient as high-risk. We aim to investigate the complex genetic landscape of neuroblastoma through a network biology perspective, focusing on complex protein-protein interaction (PPI) networks. Reanalyzing three historical neuroblastoma RNA-Seq datasets, we conducted a comparative network analysis of high-risk MYCN amplified patients and high-risk MYCN non-amplified patients to identify PPI networks that serve as predictive biomarkers for developing high-risk disease. We used R scripts to extract the top 100 most highly expressed genes from each dataset and then analyzed the expression profiles using Proteinarium, a network analysis tool that identifies clusters of patients with shared PPI networks. Statistically significant clusters were annotated and analyzed for network similarity between datasets. Our study isolated four significant clusters with majority high-risk MYCN amplified patients. We annotated the genes unique to high-risk MYCN amplified patients and identified several potential network biomarkers for this particular genotype of neuroblastoma, including ribosome biogenesis (RPL7A, RPL30, RPL35, RPS8), heat shock protein (HSPA8, HSP90AA1, HSP90AB1), matrix metallopeptidase (MMP2, MMP9, MMP13), and collagen (COL2A1, COL5A3, COL6A2, COL11A1, COL16A1) gene networks. The results of this study provide potential therapeutic targets for high-risk MYCN amplified neuroblastoma.

scMultiNODE: Integrative Model for Multi-Modal Temporal Single-Cell Data

Zhang, J. — 2024-10-29

Measuring single-cell genomic profiles at different timepoints enables our understanding of cell development. This understanding is more comprehensive when we perform an integrative analysis of multiple measurements (or modalities) across various developmental stages. However, obtaining such measurements from the same set of single cells is resource-intensive, restricting our ability to study them jointly. We introduce scMultiNODE, an unsupervised integration model that combines gene expression and chromatin accessibility measurements in developing single cells, while preserving cell type variations and cellular dynamics. First, scMultiNODE uses a scalable, Quantized Gromov-Wasserstein optimal transport to align a large number of cells across different measurements. Next, it utilizes neural ordinary differential equations to explicitly model cell development with a regularization term to learn a dynamic latent space. Experiments on six real-world developmental single-cell datasets demonstrate that scMultiNODE can integrate temporally profiled multi-modal single-cell measurements more effectively than existing methods that focus on cell type variations and often overlook cellular dynamics. We also demonstrate that scMultiNODEs joint latent space facilitates several insightful downstream analyses of single-cell development, including the investigation of complex cell trajectories and the enabling of cross-modal label transfer. The data and code are publicly available at https://github.com/rsinghlab/scMultiNODE.

Diffusion-based artificial genomes and their usefulness for local ancestry inference

Szatkownik, A. — 2024-10-31

The creation of synthetic data through generative modeling has emerged as a significant area of research in genomics, offering versatile applications from tailoring functional sequences with specific attributes to generating high-quality, privacy-preserving in silico genomes. Notwithstanding these advancements, a key challenge remains: while some methods exist to evaluate artificially generated genomic data, comprehensive tools to assess its usefulness are still limited. To tackle this issue and present a promising use case, we test artificial genomes within the framework of population genetics and local ancestry inference (LAI). Building on previous work in deep generative modeling for genomics, we introduce a novel, frugal diffusion model and show that it produces high-quality genomic data. We then assess the performance of a downstream machine learning LAI model trained on composite datasets comprising both real and/or synthetic data. Our findings reveal that the LAI model achieves comparable performance when trained exclusively on real data versus high-quality synthetic data. Moreover, we highlight how data augmentation using high-quality artificial genomes significantly benefits the LAI model, particularly when real data is limited. Finally, we compare the conventional use of a single synthetic dataset to a robust ensemble approach, wherein multiple LAI models are trained on diverse synthetic datasets, and their predictions are aggregated. Our study highlights the potential of frugal diffusion-based generative models and synthetic data integration in genomics. This approach could improve fair representation across populations by overcoming data accessibility challenges, while ensuring the reliability of genomic analyses conducted on artificial data.

Viewing geometry drives flexible perception of object motion and depth

Xu, Z.-X. — 2024-10-31

1Vision is an active process. We move our eyes and head to acquire useful information and to track objects of interest. While these movements are essential for many behaviors, they greatly complicate the analysis of retinal image motion--the image motion of an object reflects both how that object moves in the world and how the eye moves relative to the scene. Our brain must account for the visual consequences of self-motion to accurately perceive the 3D layout and motion of objects in the scene. Traditionally, compensation for eye movements (e.g., smooth pursuit) has been modeled as a simple vector subtraction process. While these models are effective for pure eye rotations and 2D scenes, we show that they fail to apply to more natural viewing geometries involving combinations of eye rotation and translation. We develop theoretical predictions for how perception of object motion and depth should depend on the observers inferred viewing geometry. Through psychophysical experiments, we demonstrate novel perceptual biases that manifest when different viewing geometries are simulated by optic flow, in the absence of physical eye movements. Remarkably, these biases occur automatically, without training or feedback, and are well predicted by our theoretical framework. A neural network model trained to perform the same tasks exhibits neural response patterns similar to those observed in macaque area MT, suggesting a possible neural basis for these adaptive computations. Our findings demonstrate that the visual system automatically infers viewing geometry from optic flow and flexibly attributes components of image motion to either self-motion or depth structure according to the inferred geometry. Our findings unify previously separate bodies of work by showing that the visual consequences of self-motion play a crucial role in computing object motion and depth, thus enabling the visual system to adaptively perceive a dynamic 3D environment.

Synaptic gene expression is coordinated across development by conserved chromatin regulators

Kentro, J. A. — 2024-10-31

Chemical synapses are the primary sites of communication in the nervous system. Synapse formation is a complex process involving hundreds of proteins that must be expressed in two cells at the same time. We find that synaptic genes are broadly and specifically coordinated at the level of transcription across developing nervous systems. How this spatiotemporal coordination is achieved remains an open question. Through genomic and functional studies in Drosophila, we demonstrate corresponding coordination of chromatin accessibility and identify chromatin regulators DEAF1 and CLAMP as broad repressors of synaptic gene expression outside windows of peak synaptogenesis. Disruption of either factor temporally dysregulates synaptic gene expression across neuronal subtypes, leading to excess synapse formation. We further find that DEAF1, which is linked to syndromic intellectual disability, is both necessary and sufficient to constrain synapse formation. Our findings reveal the critical importance of broad temporally coordinated repression of synaptic gene expression in regulating neuronal connectivity and identify two key repressors.

Vinculin Y822 phosphorylation regulates cardiomyocyte adhesion dynamics and adherens junction maturation in the heart

Li, X. — 2024-11-03

In the heart, cell-matrix and cell-cell adhesions reorganize in response to increased cardiac demand and growth to promote cardiomyocyte maturation. Vinculin, a mechanosensitive adaptor protein, links filamentous actin to cell-matrix and cell-cell adhesions and is thus positioned to regulate adhesion reorganization. However, how the two adhesion systems are coordinated in the heart, and the role of vinculin in this process is poorly understood. Here, we define the role of vinculin phosphorylation at tyrosine residue 822 (pY822) in cardiomyocyte adhesion and heart function. We found that pY822 correlated with dynamic junction remodeling in the developing heart but was lost as junctions matured postnatally. We then mutated Y822 to phenylalanine (Y822F) in the mouse to determine pY822 function in vivo. Homozygous mutant Vcl Y822F mice were viable and exhibited normal cardiac function at ten weeks of age; however, cardiac dysfunction was observed at 28 weeks. Vinculin and adherens junction proteins were reduced at cardiomyocyte junctions in Y822F hearts. In contrast, 5/{beta}1 integrin and fibronectin increased along the lateral border of Y822F cardiomyocytes. Our results demonstrate that vinculin Y822 phosphorylation regulates the balance between cadherin and integrin adhesion organization, highlighting the importance of post-translational modification in modulating vinculin function in heart physiology.

Inhibition of Yes-Associated Protein (YAP) with Verteporfin Enhances Radiosensitivity in Chordoma by Inducing G2M Arrest and Inhibiting the DNA Damage Response

Akinduro, O. O. — 2024-11-04

Chordomas are locally invasive cancers that are highly resistant to radiotherapy. The Brachyury and Yes-Associated Protein (YAP) regulatory axis has been implicated as the primary driver of tumorigenicity in chordoma. Here, we aimed to enhance chordoma radiosensitivity by repurposing the FDA-approved YAP inhibitor, Verteporfin. We used five patient-derived chordoma cell lines and generated two YAP1 knockdown cell lines to validate the YAP-targeting phenotype in chordoma. Verteporfin treatment reduced the expression of DNA damage repair proteins and genes. YAP inhibition with either verteporfin or YAP knockdown resulted in enhanced DNA double-stranded breaks after radiation via inhibition of the DNA damage repair pathway and accumulation of cells in the G2M phase. Verteporfin inhibited chordoma tumor growth alone and in combination with radiation in a xenograft mouse model treated with verteporfin loaded microparticles, resulting in sensitization of chordoma tumors to radiation. YAP inhibition with verteporfin renders chordoma more sensitive to radiation via inhibition of the DNA damage repair cascade and accumulation of cells in G2M when they are most susceptible to radiation damage.

Loss of mitochondrial enzyme GPT2 leads to reprogramming of synaptic glutamate metabolism

Baytas, O. — 2024-11-07

Recessive loss-of-function mutations in the mitochondrial enzyme Glutamate Pyruvate Transaminase 2 (GPT2) cause intellectual disability in children. Given this cognitive disorder, and because glutamate metabolism is tightly regulated to sustain excitatory neurotransmission, here we investigate the role of GPT2 in synaptic function. GPT2 catalyzes a reversible reaction interconverting glutamate and pyruvate with alanine and alpha-ketoglutarate, a TCA cycle intermediate; thereby, GPT2 may play an important role in linking mitochondrial tricarboxylic acid (TCA) cycle with synaptic transmission. In mouse brain, we find that GPT2 is enriched in mitochondria of synaptosomes (isolated synaptic terminals). Loss of Gpt2 in mouse appears to lead to reprogramming of glutamate and glutamine metabolism, and to decreased glutamatergic synaptic transmission. Whole-cell patch-clamp recordings in pyramidal neurons of CA1 hippocampal slices from Gpt2-null mice reveal decreased excitatory post-synaptic currents (mEPSCs) without changes in mEPSC frequency, or importantly, changes in inhibitory post-synaptic currents (mIPSCs). Additional evidence of defective glutamate release included reduced levels of glutamate released from Gpt2-null synaptosomes measured biochemically. Glutamate release from synaptosomes was rescued to wild-type levels by alpha-ketoglutarate supplementation. Additionally, we observed evidence of altered metabolism in isolated Gpt2-null synaptosomes: decreased TCA cycle intermediates, and increased glutamate dehydrogenase activity. Notably, alterations in the TCA cycle and the glutamine pool were alleviated by alpha-ketoglutarate supplementation. In conclusion, our data support a model whereby GPT2 mitochondrial activity may contribute to glutamate availability in pre-synaptic terminals, thereby highlighting potential interactions between pre-synaptic mitochondrial metabolism and synaptic transmission.

Blood-tumor barrier organoids recapitulate glioblastoma microenvironment and enable high-throughput modeling of therapeutic delivery

Cho, C.-F. — 2024-11-12

The blood-brain barrier (BBB) is a highly specialized system that is critical for regulating transport between the blood and the central nervous system. In brain tumors, the vasculature system is compromised, and is referred to as the blood-tumor barrier (BTB). The ability to precisely model the unique physiological properties of the BTB is essential to decipher its role in tumor pathophysiology and for the rational design of efficacious therapeutics. Here, we introduce a robust and high-throughput in vitro 3D human BTB organoid model that recapitulates various key features of the BTB observed in vivo and in clinical GBM samples. The organoids are composed of patient-derived glioblastoma stem cells (GSCs), human brain endothelial cells (EC), astrocytes and pericytes, which are formed through self-assembly. Transcriptomic and functional analyses reveal that the GSCs in the BTB organoids exhibit enhanced level of stemness, mesenchymal signature, invasiveness and angiogenesis, and this is further confirmed in in vivo studies. We demonstrate the ability of the BTB organoids to model therapeutic delivery and drug efficacy on brain tumor cells. Collectively, our findings show that the BTB organoid model has broad utility as a clinically representative system for studying the BTB and evaluating brain tumor therapies.

Aberrant expression of collagen type X in solid tumor stroma is associated with EMT, immunosuppressive and pro-metastatic pathways, bone marrow stromal cell signatures, and poor survival prognosis

Famili-Youth, E. H. H. — 2024-11-14

BackgroundCollagen type X (ColX1, encoded by COL10A1) is expressed specifically in the cartilage-to-bone transition, in bone marrow cells, and in osteoarthritic (OA) cartilage. We have previously shown that ColX1 is expressed in breast tumor stroma, correlates with tumor-infiltrating lymphocytes, and predicts poor adjuvant therapy outcomes in ER+/HER2+ breast cancer. However, the underlying molecular mechanisms for these effects are unknown. In this study, we performed bioinformatic analysis of COL10A1-associated gene modules in breast and pancreatic cancer as well as in cells from bone marrow and OA cartilage. These findings provide important insights into the mechanisms of transcriptional and extracellular matrix changes which impact the local stromal microenvironment and tumor progression. MethodsImmunohistochemistry was performed to examine collagen type X expression in solid tumors. WGCNA was used to generate COL10A1-associated gene networks in breast and pancreatic tumor cohorts using RNA-Seq data from The Cancer Genome Atlas. Computational analysis was employed to assess the impact of these gene networks on development and progression of cancer and OA. Data processing and statistical analysis was performed using R and various publicly-available computational tools. ResultsExpression of COL10A1 and its associated gene networks highlights inflammatory and immunosuppressive microenvironments, which identify aggressive breast and pancreatic tumors and contribute to metastatic potential in a sex-dependent manner. Both cancer types are enriched in stroma, and COL10A1 implicates bone marrow-derived fibroblasts as drivers of the epithelial-to-mesenchymal transition (EMT) in these tumors. Heightened expression of COL10A1 and its associated gene networks is correlated with poorer patient outcomes in both breast and pancreatic cancer. Common transcriptional changes and chondrogenic activity are shared between cancer and OA cartilage, suggesting that similar microenvironmental alterations may underlie both diseases. ConclusionsCOL10A1-associated gene networks may hold substantial value as regulators and biomarkers of aggressive tumor phenotypes with implications for therapy development and clinical outcomes. Identification of tumors which exhibit high expression of COL10A1 and its associated genes may reveal the presence of bone marrow-derived stromal microenvironments with heightened EMT capacity and metastatic potential. Our analysis may enable more effective risk assessment and more precise treatment of patients with breast and pancreatic cancer. Research HighlightsO_LIColX highlights features of EMT in breast and pancreatic cancer C_LIO_LIColX gene modules are immunosuppressive and pro-metastatic C_LIO_LIColX-associated gene networks contribute to sex differences in pancreatic cancer C_LIO_LIColX-positive fibroblasts define more aggressive tumors with poorer survival C_LIO_LIColX is emerging as a biomarker for bone marrow-derived cells in cancer C_LI

A novel rhodopsin-based voltage indicator for simultaneous two-photon optical recording with GCaMP in vivo

Villette, V. — 2024-11-17

Genetically encoded voltage indicators (GEVIs) allow optical recording of membrane potential from targeted cells in vivo. However, red GEVIs that are compatible with two-photon microscopy and that can be multiplexed in vivo with green reporters like GCaMP, are currently lacking. To address this gap, we explored diverse rhodopsin proteins as GEVIs and engineered a novel GEVI, 2Photron, based on a rhodopsin from the green algae Klebsormidium nitens. 2Photron, combined with two photon ultrafast local volume excitation (ULoVE), enabled multiplexed readout of spiking and subthreshold voltage simultaneously with GCaMP calcium signals in visual cortical neurons of awake, behaving mice. These recordings revealed the cell-specific relationship of spiking and subthreshold voltage dynamics with GCaMP responses, highlighting the challenges of extracting underlying spike trains from calcium imaging.

Phospholipid Scramblase 1 (PLSCR1) Regulates Interferon-Lambda Receptor 1 (IFN-lambdaR1) and IFN-lambda Signaling in Influenza A Virus (IAV) Infection

Yang, A. X. — 2024-11-21

Phospholipid scramblase 1 (PLSCR1) is an antiviral interferon-stimulated gene (ISG) that has several known anti-influenza functions. However, the mechanisms in relation to its expression compartment and enzymatic activity have not been completely explored. Moreover, only limited animal models have been studied to delineate its role at the tissue level in influenza infections. Our results showed that Plscr1 expression was highly induced by influenza A virus (IAV) infection in vivo and in airway epithelial cells treated with IFN-{lambda}. We found that infected Plscr1-/-mice exhibited exacerbated body weight loss, decreased survival rates, heightened viral replication, and increased lung damage. Interestingly, transcriptomic analyses demonstrated that Plscr1 was required for the expression of type 3 interferon receptor (Ifn-{lambda}r1) and a large subset of ISGs upon IAV infection. The impaired expression of Ifn-{lambda}r1 and downstream ISGs may be responsible for delayed viral clearance and worse lung inflammation in Plscr1-/- mice. PLSCR1 acts as a transcriptional activator of IFN-{lambda}R1 by directly binding to its promotor after IAV infection. In addition, PLSCR1 interacted with IFN-{lambda}R1 on the cell surface of pulmonary epithelial cells following IAV infection, suggesting it also modulated IFN-{lambda} signaling via protein-protein interactions. The lipid scramblase activity of PLSCR1 was found to be dispensable for its anti-flu activity. Finally, single-cell RNA sequencing data indicated that Plscr1 expression was significantly upregulated in ciliated airway epithelial cells in mice following IAV infection. Consistently, Plscr1floxStopFoxj1-Cre+ mice with ciliated epithelial cell-specific Plscr1 overexpression showed reduced susceptibility to IAV infection, less inflammation and enhanced Ifn-{lambda}r1 expression, suggesting that Plscr1 primarily regulates type 3 IFN signaling as a cell intrinsic defense factor against IAV in ciliated airway epithelial cells. Our research will elucidate virus-host interactions and pave the way for the development of novel anti-influenza drugs that target human elements like PLSCR1, thereby mitigating the emergence of drug-resistant IAV strains.

Molecular and genetic characterization of sex-linked orange coat color in the domestic cat

Kaelin, C. B. — 2024-11-22

The Sex-linked orange mutation in domestic cats causes variegated patches of reddish/yellow hair and is a defining signature of random X-inactivation in female tortoiseshell and calico cats. Unlike the situation for most coat color genes, there is no apparent homolog for Sex-linked orange in other mammals. We show that the Sex-linked orange is caused by a 5 kb deletion that leads to ectopic and melanocyte-specific expression of the Rho GTPase Activating Protein 36 (Arhgap36) gene. Single cell RNA-seq studies from fetal cat skin reveal that red/yellow hair color is caused by reduced expression of melanogenic genes that are normally activated by the Melanocortin 1 receptor (Mc1r)--cyclic adenosine monophosphate (cAMP)--protein kinase A (PKA) pathway, but the Mc1r gene and its ability to stimulate cAMP accumulation is intact. Instead, we show that increased expression of Arhgap36 in melanocytes leads to reduced levels of the PKA catalytic subunit (PKAC); thus, Sex-linked orange is genetically and biochemically downstream of Mc1r. Our findings solve a comparative genomic conundrum, provide in vivo evidence for the ability of Arhgap36 to inhibit PKA, and reveal a molecular explanation for a charismatic color pattern with a rich genetic history.

Semi-automated high content analysis of pollen performance using TubeTracker

Yimga Ouonkap, S. V. — 2024-11-22

Pollen function is critical for successful plant reproduction and crop productivity and it is important to develop accessible methods to quantitatively analyze pollen performance to enhance reproductive resilience. Here we introduce TubeTracker as a method to quantify key parameters of pollen performance such as, time to pollen grain germination, pollen tube tip velocity and pollen tube survival. TubeTracker integrates manual and automatic image processing routines and the graphical user interface allows the user to interact with the software to make manual corrections of automated steps. TubeTracker does not depend on training data sets required to implement machine learning approaches and thus can be immediately implemented using readily available imaging systems. Furthermore, TubeTracker is an excellent tool to produce the pollen performance data sets necessary to take advantage of emerging AI-based methods to fully automate analysis. We tested TubeTracker and found it to be accurate in measuring pollen tube germination and pollen tube tip elongation across multiple cultivars of tomato. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/624782v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1fc2a63org.highwire.dtl.DTLVardef@42f3a2org.highwire.dtl.DTLVardef@18911d6org.highwire.dtl.DTLVardef@1f236f0_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract Graphical user interface of TubeTracker showing all supported functionalities. C_FIG

Host specificity of gastrointestinal parasites in free-ranging sloths from Costa Rica

Vanderheoven, E. — 2024-11-23

The diversity and host specificity of gastrointestinal parasites infecting free-ranging sloths is poorly known. We characterized gastrointestinal parasites of two sloth species from Costa Rica-- three-fingered sloth (Bradypus variegatus) and two-fingered sloth (Choloepus hoffmanni)--for the first time in both a primary forest and an urban habitat. We asked whether host-parasite interactions were predominantly structured by host identity, the habitats in which hosts occurred, or both. Protozoa and nematode eggs were present in both species, but cestode eggs were recorded only in C. hoffmanni. We found eight parasitic morphotypes, which matches the total number of parasite taxa described in sloths over the past 100 years, and these first reports of gastrointestinal parasites of sloths from primary forest radically expand our knowledge of the general diversity and host specificity of sloth parasites. We found no significant difference in overall parasite richness between sloth species or habitats, but the parasite richness of C. hoffmanni was 2-fold greater in the primary forest vs. urban habitat. As no parasite sharing was observed between sloth species, we found strong and significant differences in parasite composition between host species regardless of habitat. In B. variegatus, we observed eggs of four nematode taxa (Spirocercidae, Subuluroidea, Spirurida, Ascaridida) and cysts of Eimeriidae (Apicomplexa). By contrast, in C. hoffmanni, we observed cestodes (Anoplocephalidae), a different nematode from the family Spirocercidae and also cysts of Eimeriidae (Apicomplexa). Many rare taxa were recorded only in samples from the primary forest, and these did not match any sloth parasites that had been previously described in the literature, suggesting that at least some could be undescribed species. Together, these results highlight the need for further research in comparative wildlife parasitology, the characterization of host-parasite transmission networks, and the identification of any intermediate hosts that may be relevant to sloth health.

TimeFlies: an snRNA-seq aging clock for the fruit fly head sheds light on sex-biased aging

Tennant, N. — 2024-11-27

Although multiple high-performing epigenetic aging clocks exist, few are based directly on gene expression. Such transcriptomic aging clocks allow us to identify potential age-associated genes directly. However, most existing transcriptomic clocks model a subset of genes and are limited in their ability to predict novel biomarkers. With the growing application of single-cell sequencing, there is a need for robust single-cell transcriptomic aging clocks. Moreover, aging clocks have yet to be applied to investigate the elusive phenomenon of sex differences in aging. We introduce TimeFlies, a pan-cell-type snRNA-seq aging clock for the Drosophila melanogaster head. TimeFlies uses deep learning to classify the donor age of cells based on genome-wide gene expression profiles. Using explainability methods, we identified key marker genes contributing to the classification, with lncRNAs showing up as highly enriched among predicted biomarkers. lncRNA:roX1 and lncRNA:roX2 are top clock genes across cell types. Both are regulators of X chromosome dosage compensation, a pathway previously found to be significantly affected by aging in the mouse brain. We validated these findings experimentally in Drosophila, showing a decrease in survival when dosage compensation is inhibited in vivo. Furthermore, we trained sex-specific TimeFlies clocks and noted significant differences in model predictions and explanations between male and female clocks, suggesting that different pathways drive aging in males and females.

Dynamic Regulation Of The Chromatin Environment By ASH1L Modulates Human Neuronal Structure And Function

Jhanji, M. — 2024-12-02

Precise regulation of the chromatin environment through post-translational histone modification modulates transcription and controls brain development. Not surprisingly, mutations in a large number of histone-modifying enzymes underlie complex brain disorders. In particular, the histone methyltransferase ASH1L modifies histone marks linked to transcriptional activation and has been implicated in multiple neuropsychiatric disorders. However, the mechanisms underlying the pathobiology of ASH1L-asociated disease remain underexplored. We generated human isogenic stem cells with a mutation in ASH1Ls catalytic domain. We find that ASH1L dysfunction results in reduced neurite outgrowth, which correlates with alterations in the chromatin profile of activating and repressive histone marks, as well as the dysregulation of gene programs important for neuronal structure and function implicated in neuropsychiatric disease. We also identified a novel regulatory node implicating both the SP and Kruppel-like families of transcription factors and ASH1L relevant to human neuronal development. Finally, we rescue cellular defects linked to ASH1L dysfunction by leveraging two independent epigenetic mechanisms that promote transcriptional activation. In summary, we identified an ASH1L-driven epigenetic and transcriptional axis essential for human brain development and complex brain disorders that provide insights into future therapeutic strategies for ASH1L-related disorders.

The oncogene SLC35F2 is a high-specificity transporter for the micronutrients queuine and queuosine

Burtnyak, L. — 2024-12-04

The nucleobase queuine (q) and its nucleoside queuosine (Q) are micronutrients derived from bacteria that are acquired from the gut microbiome and/or diet in humans. Following cellular uptake, Q is incorporated at the wobble base (position 34) of tRNAs with a GUN anticodon, which is important for efficient translation. Early studies suggested that cytosolic uptake of queuine is mediated by a selective transporter that is regulated by mitogenic signals, but the identity of this transporter has remained elusive. Here, through a cross-species bioinformatic search and genetic validation, we have identified the solute carrier family member SLC35F2 as a unique transporter for both queuine and queuosine in Schizosaccharomyces pombe and Trypanosoma brucei. Furthermore, gene disruption in HeLa cells revealed that SLC35F2 is the sole transporter for queuosine in HeLa cells (Km 174 nM) and a high-affinity transporter for the queuine nucleobase (Km 67 nM), with the presence of another low-affinity transporter (Km 259 nM) in these cells. Competition uptake studies show that SLC35F2 is not a general transporter for other canonical ribonucleobases or ribonucleosides, but selectively imports q and Q. The identification of SLC35F2, an oncogene, as the transporter of both q and Q advances our understanding of how intracellular levels of queuine and queuosine are regulated and how their deficiency contributes to a variety of pathophysiological conditions, including neurological disorders and cancer. Significance StatementThe discovery of SLC35F2 as the eukaryotic transporter of queuine and queuosine is key to understanding how these micronutrients are salvaged from the human gut and distributed to different body tissues. Queuosine modification of tRNAs enhances the accuracy and efficiency of codon-anticodon pairing and regulates a range of biological and pathophysiological states, including oxidative stress responses, cancer, learning, memory, and gut homeostasis.

Universal Single Copy Knock-In System in Caenorhabditis elegans: One Plasmid to Target All Chromosomes

Dinneen, E. — 2024-12-10

Successful transgenesis in model organisms has dramatically helped us understand gene function, regulation, genetic networks, and potential applications. Here, we introduce the universal single-copy knock-in system (Universal SKI System or U-SKI), designed for inserting any transgene by CRISPR/Cas9 in the Caenorhabditis elegans genome. The Universal SKI System takes advantage of a plasmid (pSKI), which can also be used for extrachromosomal arrays, to facilitate the insertion of a transgene at specific safe harbor loci on each autosomal chromosome. The pSKI plasmid contains multiple restriction sites for easy cloning and serves as a CRISPR/Cas9-based insertion repair template because it has two synthetic and long homology arms that recombine with the SKI cassettes. This system also uses a single crRNA guide, which acts as a Co-CRISPR enrichment marker. Overall, the Universal SKI System is highly flexible; with the same Universal SKI cassette on each autosome, researchers can select the insertion site and streamline tracking while reducing the complexity of expressing single-copy transgenes in C. elegans.

Dysfunctional oscillatory bursting patterns underlie working memory deficits in adolescents with ADHD

Kavanaugh, B. — 2024-12-10

Identifying neural markers of clinical symptom fluctuations is prerequisite to developing more precise brain-targeted treatments in psychiatry. We have recently shown that working memory (WM) in healthy adults is dependent on the rise and fall interplay between alpha/beta and gamma bursts within frontoparietal regions, and that deviations in these patterns lead to WM performance errors. However, it is not known whether such bursting deviations underlie clinically relevant WM-related symptoms or clinical status in individuals with WM deficits. In adolescents (n=27) with attention deficit hyperactivity disorder (ADHD), we investigated WM-related dynamics between alpha/beta and gamma bursts in relation to clinical status fluctuations. Participants repeatedly completed a visual Sternberg spatial working memory task during EEG recording as part of their participation in two research studies (n=224 person-sessions). Source localizing EEG data to each participants structural MRI, the rate and volume of alpha, beta, and gamma bursts were examined within the dorsolateral prefrontal cortex (DLPFC) and posterior parietal cortex (PPC). Alpha/beta and gamma bursts at the DLPFC and PPC displayed complimentary roles in WM processes. Alpha/beta bursting decreased during stimuli encoding and increased during the delay, while gamma bursting was elevated during encoding and decreased during the delay. Deviations in bursting patterns were associated with WM errors and clinical symptoms. We conclude that dysfunctional alpha/beta and gamma burst dynamics within the frontoparietal region underlie both intra-individual WM performance and WM symptom fluctuations in adolescents with ADHD. Such burst dynamics reflect a novel target and biomarker for WM-related treatment development.

The intersection of inflammation and DNA damage as a novel axis underlying the pathogenesis of autism spectrum disorders

lizarraga, s. B. — 2024-12-11

Autism spectrum disorders (ASD) affects 1 in 36 children and is characterized by repetitive behaviors and difficulties in social interactions and social communication. The etiology of ASD is extremely heterogeneous, with a large number of ASD cases that are of unknown or complex etiology, which suggests the potential contribution of epigenetic risk factors. In particular, epidemiological and animal model studies suggest that inflammation during pregnancy could lead to an increased risk of ASD in the offspring. However, the molecular mechanisms that contribute to ASD pathogenesis in relation to maternal inflammation during pregnancy in humans are underexplored. Several pro-inflammatory cytokines have been associated with increased autistic-like behaviors in animal models of maternal immune activation, including IL-17A. Using a combination of ASD patient lymphocytes and stem cell-derived human neurons exposed to IL-17A we discovered a shared molecular signature that highlights a metabolic and translational node that could lead to altered neuronal excitability. Further, our work on human neurons brings forward the possibility that defects in the DNA damage response could be underlying the effect of IL-17A on human excitatory neurons, linking exacerbated unrepaired DNA damage to the pathogenicity of maternal inflammation in connection to ASD.

Efficient High-Throughput DNA Breathing Features Generation Using Jax-EPBD

Inan, T. T. — 2024-12-12

DNA breathing dynamics--transient base-pair opening and closing due to thermal fluctuations--are vital for processes like transcription, replication, and repair. Traditional models, such as the Extended Peyrard-Bishop-Dauxois (EPBD), provide insights into these dynamics but are computationally limited for long sequences. We present JAX-EPBD, a high-throughput Langevin molecular dynamics framework leveraging JAX for GPU-accelerated simulations, achieving up to 30x speedup and superior scalability compared to the original C-based EPBD implementation. JAX-EPBD efficiently captures time-dependent behaviors, including bubble lifetimes and base flipping kinetics, enabling genome-scale analyses. Applying it to transcription factor (TF) binding affinity prediction using SELEX datasets, we observed consistent improvements in R2 values when incorporating breathing features with sequence data. Validating on the 77-bp AAV P5 promoter, JAX-EPBD revealed sequence-specific differences in bubble dynamics correlating with transcriptional activity. These findings establish JAX-EPBD as a powerful and scalable tool for understanding DNA breathing dynamics and their role in gene regulation and transcription factor binding.

Endosome maturation is orchestrated by inside-out proton signaling through a Na+/H+ exchanger and pH-dependent Rab GTPase cycling

Lee, Y. — 2024-12-12

Endosome maturation requires progressive lumen acidification. To what extent is lumen acidification sensed by cytosolic-side molecules that drive endosome maturation? We show here that "inside-out" proton signaling through the endosomal Na+/H+ Exchanger 6 (NHE6) activates the late endosome master regulator Rab7. The mechanism involves potent inactivation of the Rab7 GTPase-activating protein (GAP) TBC1D5 with decreasing pH. NHE6 interacts with TBC1D5 in a complex with Rab7. Neurons from NHE6-null mice or mice engineered with a selective defect in NHE6 proton efflux exhibit blocked endosome maturation and decreased active Rab7, consistent with an overactive Rab7 GAP. Finally, epistatic knock-down of TBC1D5, thereby reducing Rab7 GAP activity, in NHE6-null neurons rescues Rab7 GTPase cycling and endosome maturation. Importantly, NHE6 is mutated in Christianson Syndrome underscoring the significance of these mechanisms to neurodegeneration. We conclude that lumen acidification regulates pH-dependent Rab GTPase cycling to coordinate late endosome maturation by a process involving proton signaling.

In silico biophysics and rheology of blood and red blood cells in Gaucher Disease

Chai, Z. — 2024-12-12

Gaucher Disease (GD) is a rare genetic disorder characterized by a deficiency in the enzyme glucocerebrosidase, leading to the accumulation of glucosylceramide in various cells, including red blood cells (RBCs). This accumulation results in altered biomechanical properties and rheological behavior of RBCs, which may play an important role in blood rheology and the development of bone infarcts, avascular necrosis (AVN) and other bone diseases associated with GD. In this study, dissipative particle dynamics (DPD) simulations are employed to investigate the biomechanics and rheology of blood and RBCs in GD under various flow conditions. The model incorporates the unique characteristics of GD RBCs, such as decreased deformability and increased aggregation properties, and aims to capture the resulting changes in RBC biophysics and blood viscosity. This study is the first to explore the Youngs modulus and aggregation parameters of GD RBCs by validating simulations with confocal imaging and experimental RBC disaggregation thresholds. Through in silico simulations, we examine the impact of hematocrit, RBC disaggregation threshold, and cell stiffness on blood viscosity in GD. The results reveal three distinct domains of GD blood viscosity based on shear rate: the aggregation domain, where the RBC disaggregation threshold predominantly influences blood viscosity; the transition area, where both RBC aggregation and stiffness impact on blood viscosity; and the stiffness domain, where the stiffness of RBCs emerges as the primary determinant of blood viscosity. By quantitatively assessing RBC deformability, RBC disaggregation threshold, and blood viscosity in relation to bone disease, we find that the RBC aggregation properties, as well as their deformability and blood viscosity, may contribute to its onset. These findings enhance our understanding of how changes in RBC properties impact on blood viscosity and may affect bone health, offering a partial explanation for the bone complications observed in GD patients. Author summaryIn Gaucher Disease (GD), a genetic deficiency in the enzyme glucocerebrosidase leads to the accumulation of glucosylceramide in red blood cells (RBCs), resulting in altered biomechanical properties. These changes affect blood flow characteristics, particularly blood viscosity, and may contribute to bone health issues seen in GD patients, including bone infarcts, avascular necrosis (AVN), and other bone diseases. In our study, we apply dissipative particle dynamics (DPD) simulations to explore how GD impacts RBC behavior under various flow conditions. We model GD RBCs with decreased deformability and increased aggregation, examining how these properties influence blood viscosity across three distinct shear rate domains: aggregation, transition, and stiffness. By validating our simulations with confocal imaging data and experimental RBC disaggregation thresholds, we quantitatively assess the effects of RBC stiffness, aggregation, and hematocrit levels on blood flow in GD. We find that the RBC aggregation properties, deformability and blood viscosity, may contribute to the onset of bone disease. These findings improve our understanding of how changes in RBC properties influence blood viscosity and may contribute to bone health issues, providing a partial explanation for the bone complications observed in GD patients.

CSF1R-CAR T cells induce CSF1R signalling and promote cancer cell growth

Callahan, A. — 2024-12-20

Chimeric antigen receptor (CAR) T cells have transformed the landscape of cancer therapy and demonstrate unprecedented success in treating relapsed/refractory blood cancers. The mechanism underlaying the interactions and responses of CAR T cells and their targets remain incompletely understood. Previous studies focus on the activation of CAR T cells and attempt to optimise CAR design to increase efficacy, meanwhile ignoring tumours and their responses to CAR ligation. Here, we evaluate the signalling capacity of a second generation CSF1-tageted CSF1R CAR compared with a scFv-targeted CD19-CAR using a SILAC co- culture approach coupled with phosphotyrosine (pY) enrichment and LC-MS/MS. We show that ligation of CSF1R-expressing THP1 cells with CSF1R-CAR T cells induces CSF1R-like signalling in THP1 cells, whereas no target cell signalling response is observed after CD19- CAR/Raji B cell ligation. Using small molecule inhibitors of Lck, actin polymerisation, and CSF1R, we find that CAR-induced CSF1R signalling in THP1 cells depends exclusively on CSF1R kinase activity with no participation from T cell activation. Consistently, CSF1R- CAR T cells promote THP1 growth at low effector-to-target (E:T) ratios but prevent THP1 growth at high E:T ratios. Our data provide evidence for an unintended consequence of CARs; CAR-induced signalling in cancer cells. These data may have broad implications for the choice of CAR antigen for optimal clinical efficacy. One Sentence Summary: CSF1R-CAR activates intracellular signalling cascades in THP1 cells, which promote THP1 cell growth.

Genomic insights into Plasmodium vivax and Plasmodium simium host shifts in Latin America

Lefebvre, M. J. M. — 2024-12-22

Malaria in Latin America is largely caused by Plasmodium vivax, but its lesser-known sister species, Plasmodium simium, has recently emerged from monkeys to infect humans, thus raising new public health concerns. By analyzing 719 monkey samples and whole genome variations for 19 P. simium and 408 P. vivax isolates, we investigated the evolutionary history and population genetics of the two species. P. vivax, typically restricted to humans, was identified in three Colombian and one Brazilian monkeys, suggesting host niche expansion. Genetic analysis reveals recent genetic exchanges between both species and indicates that P. simium originated from a host jump approximately a century ago, possibly linked to P. vivax migration from Mexico to Brazil. Genome-wide scans revealed signals of positive selection in P. simium genes involved in interactions with primate hosts and mosquito vectors. These findings highlight P. simium evolutionary history and zoonotic malaria risks, and underscore the need to include monkeys in malaria prevention measures while ensuring human-wildlife coexistence.

C. elegans LET-381/FoxF and DMD-4/DMRT control development of the mesodermal HMC endothelial cell

Stefanakis, N. — 2024-12-24

Endothelial cells form the inner layer of blood vessels and play key roles in circulatory system development and function. A variety of endothelial cell types have been described through gene expression and transcriptome studies; nonetheless, the transcriptional programs that specify endothelial cell fate and maintenance are not well understood. To uncover such regulatory programs, we studied the C. elegans Head Mesodermal Cell (HMC), a non-contractile mesodermal cell bearing molecular and functional similarities to vertebrate endothelial cells. Here, we demonstrate that a Forkhead transcription factor, LET-381/FoxF, is required for HMC fate specification and maintenance of HMC gene expression. DMD-4, a Dmrt transcription factor, acts downstream of and in conjunction with LET-381 to mediate HMC fate specification and gene expression maintenance. DMD-4, independently of LET-381, also represses the expression of genes associated with a different, non-HMC, mesodermal fate. Our studies uncover essential roles for FoxF transcriptional regulators in endothelial cell development, and suggest that the identity of FoxF co-functioning target transcription factors promotes specific non-contractile mesodermal fates.

Rethinking cancer drug synergy prediction: a call for standardization in machine learning applications

Wong, A. M. — 2024-12-24

Drug resistance poses a significant challenge to cancer treatment, often caused by intratumor heterogeneity. Combination therapies have been shown to be an effective strategy to prevent resistant cancer cells from escaping single-drug treatments. However, discovering new drug combinations through traditional molecular assays can be costly and time-consuming. In silico approaches can overcome this limitation by exploring many candidate combinations at scale. This study systematically evaluates the utility of various machine learning algorithms, input features, and drug synergy prediction tasks. Our findings indicate a pressing need for establishing a standardized framework to measure and develop algorithms capable of predicting synergy.

Transcriptional Integration of Meiotic Prophase I Progression and Early Oocyte Differentiation

Abt, K. M. — 2025-01-06

Female reproductive senescence results from the regulated depletion of a finite pool of oocytes called the ovarian reserve. This pool of oocytes is initially established during fetal development, but the oocytes that comprise it must remain quiescent for decades until they are activated during maturation in adulthood. In order for developmentally competent oocytes to populate the ovarian reserve they must successfully initiate both meiosis and oogenesis. As the factors that regulate the timing and fidelity of these early events remain elusive, we assessed the precise function and timing of the transcriptional regulator TAF4b during meiotic prophase I progression in mouse fetal oocytes. Compared to matched controls, E14.5 Taf4b-deficient oocytes enter meiosis I in a timely manner however, their subsequent progression through the pachytene-to-diplotene transition of meiotic prophase I is compromised. Moreover, this disruption of meiotic progression is associated with the reduced ability of Taf4b-deficient oocytes to repair double-strand DNA breaks. Transcriptional profiling of Taf4b-deficient oocytes reveals that between E16.5 and E18.5 these oocytes fail to coordinate the reduction of meiotic gene expression and the induction of oocyte differentiation genes. These studies reveal that TAF4b promotes the formation of the ovarian reserve in part by orchestrating the timely transition to meiosis I arrest and oocyte differentiation, which are often perceived as separate events.

Ensemble priming via competitive inhibition: local mechanisms of sensory context storage and deviance detection in the neocortical column

Thorpe, R. V. — 2025-01-09

The process by which neocortical neurons and circuits amplify their response to an unexpected change in stimulus, typically referred to as deviance detection (DD), has traditionally been thought to be the product of specialized cell types and/or routing from distinct brain areas. Here, we explore a different theory, whereby DD emerges intrinsically from local network-level interactions within a neocortical column. We propose that deviance-driven neural dynamics are generated by ensembles of excitatory and inhibitory neurons that have a fundamental inhibitory connectivity motif: competitive inhibition between reciprocally connected neural representations under modulation from feed-forward selective (dis)inhibition. Implementing this motif in two computational models with different levels of biophysical abstraction, we were able to simulate a variety of phenomena pertaining to the experimentally observed shifts in neural tuning during DD across neurons, time, and stimulus history. We further tested hypotheses related to our theory and examined the robustness of emergent phenomena consistent with prior experimental observations. Our results show that ensemble priming via competitive inhibition under modulation from selective (dis)inhibition can serve as a local mechanism for encoding short-term stimulus memory, enabling deviance-driven shifts in stimulus representation. This work establishes a novel theoretical paradigm that resolves previously confounding aspects of predictive sensory processing in Neocortex, and we provide a number of corollary predictions that can be tested in future in vivo studies.

DNA polymerase beta expression in head & neck cancer modulates the poly(ADP-ribose)-mediated replication checkpoint

Khan, M. M. — 2025-01-20

Head and Neck Squamous Cell Carcinoma (HNSCC) imposes a significant health burden, necessitating innovative therapeutic strategies to enhance treatment efficacy. Current treatments such as surgery, radiation, and chemotherapy have limited effectiveness and yield severe side effects, emphasizing the need for targeted therapies. We have focused on DNA polymerase beta (Pol{beta}) and its roles in replication stress, cellular responses to DNA damaging therapies, and DNA damage response modifiers. Our investigations reveal a regulatory role for base excision repair (BER) proteins, including Pol{beta}, in the cellular response to inhibitors of poly(ADP-ribose) glycohydrolase (PARG), an enzyme involved in poly(ADP-ribose) (PAR) degradation. The inhibition of PARG, in HNSCC cells, elicits replication stress. Further, this activates the PAR-induced S-phase/ATR checkpoint, leading to a block to replication, cell cycle arrest, and the onset of apoptosis. However, Pol{beta} overexpression mitigates this response, reducing replication-stress induced PAR foci formation, suggesting a modulation of replication checkpoint activation. We found that PARG inhibitor treatment is ineffective on HNSCC cells that overexpress Pol{beta}, implying that the PARG inhibitor-induced PAR and apoptotic response is dependent on the level of Pol{beta}. Further, our in vitro experiments demonstrate that combining PARG and ATR/CHK1 inhibitors overcomes Pol{beta}-mediated treatment resistance in HNSCC cells, producing enhanced effects as compared to the individual treatment conditions. Our findings suggest a possible treatment paradigm for HNSCC, employing ATR or CHK1 inhibitors in combination with PARG inhibitors. This strategy offers a promising path for more effective HNSCC treatments, potentially overcoming Pol{beta}-related resistance.

Sensorimotor adaptation reveals systematic biases in 3D perception.

Lim, C. — 2025-01-22

The existence of biases in visual perception and their impact on visually guided actions has long been a fundamental yet unresolved question. Evidence revealing perceptual or visuomotor biases has typically been disregarded because such biases in spatial judgments can often be attributed to experimental measurement confounds. To resolve this controversy, we leveraged the visuomotor systems adaptation mechanism -- triggered only by a discrepancy between visual estimates and sensory feedback -- to directly indicate whether systematic errors in perceptual and visuomotor spatial judgments exist. To resolve this controversy, we leveraged the adaptive mechanisms of the visuomotor system to directly reveal whether systematic biases or errors in perceptual and visuomotor spatial judgments exist. In a within-subject study (N=24), participants grasped a virtual 3D object with varying numbers of depth cues (single vs. multiple) while receiving haptic feedback. The resulting visuomotor adaptations and aftereffects demonstrated that the planned grip size, determined by the visually perceived depth of the object, was consistently overestimated. This overestimation intensified when multiple cues were present, despite no actual change in physical depth. These findings conclusively confirm the presence of inherent biases in visual estimates for both perception and action, and highlight the potential use of visuomotor adaptation as a novel tool for understanding perceptual biases.

Target the Heart: a new axis of Alzheimer's disease prevention

Heller, L. — 2025-01-28

Cyclosporine A and other calcineurin inhibitors have been identified as prospective treatments for preventing Alzheimers disease. Utilizing a neural network model, Z-LaP Tracker, we previously found that calcineurin inhibitors elicit a unique behavioral profile in zebrafish larvae characterized by increased activity, acoustic hyperexcitability, and reduced visually guided behaviors. Screening a large library of FDA-approved drugs using Z-LaP Tracker revealed a cluster of 65 drugs demonstrating a cyclosporine A-like behavioral profile. 14 of these drugs were heart medications, including angiotensin receptor blockers, beta-blockers, alpha-adrenergic receptor antagonists, and a statin. This suggests some heart medications may be effective in preventing or ameliorating Alzheimers disease pathology. Other studies have shown that many of these 14 drugs directly or indirectly inhibit the calcineurin-NFAT pathway, alike cyclosporine A. Dual administration of the heart medications with cyclosporine A in Z-LaP Tracker revealed synergistic effects: lower doses of each heart medication could be delivered in conjunction with a lower dose of cyclosporine A to evoke a similar or larger behavioral effect than higher doses of each drug independently. This indicates that co-administering a low dose of cyclosporine A with select cardiac drugs could be a potentially effective treatment strategy for Alzheimers disease and cardiovascular dysfunction, while mitigating side effects associated with higher doses of cyclosporine A. Given that heart disease precedes Alzheimers disease in many patients, physicians may be able to create a treatment regimen that simultaneously addresses both conditions. Our results suggest that cyclosporine A combined with simvastatin, irbesartan, cilostazol, doxazosin, or nebivolol are the most promising candidates for future exploration.

Distinctive associations between plasma p-tau181 levels and hippocampal subfield volume across the Alzheimer's disease continuum

Rich, A. — 2025-01-28

BackgroundPlasma p-tau181 is a promising diagnostic marker of Alzheimers disease (AD) pathology, reflecting amyloid accumulation, tau deposition, and downstream neurodegeneration that leads to cognitive impairment. However, the specificity of plasma p-tau181 to AD-related tau pathology remains unclear. ObjectiveTo assess whether plasma p-tau181 is differentially associated with volumetric changes in distinct hippocampal subfields and whether they mediate the relationship between plasma p-tau181 and cognition across the AD continuum. Methods213 participants with normal cognition (N=57), mild cognitive impairment (N=109), and AD (N=47) from the Alzheimers Disease Neuroimaging Initiative (ADNI) were included for cross-sectional analyses of hippocampal subfield volume that was quantified using the Automatic Segmentation of Hippocampal Subfields (ASHS) software. A subset (n=89) was evaluated for one-year longitudinal changes in hippocampal subfield volume. ResultsHigher plasma p-tau181 levels (pg/mL) were associated with decreased volumes in the CA1 and dentate gyrus, bilaterally, and right entorhinal cortex (ps < 0.05). Additionally, volumes of these subfields partially mediated the relationship between plasma p-tau181 and ADNI memory and executive function composite scores. Baseline plasma p-tau181, however, did not predict longitudinal atrophy of hippocampal subfields across diagnostic groups. ConclusionsPlasma p-tau181 is differentially associated with hippocampal subfields that are closely related to both age- and AD-related neurodegeneration. Elevated plasma p-tau181 levels may reflect tau accumulation, and volumetric changes in CA1 and DG may mediate the detrimental effect of tau pathology on cognition.

Visual Influence Networks in Walking Crowds

Yoshida, K. — 2025-01-30

Collective motion in human crowds has been understood as a self-organizing phenomenon that is generated from local visual interactions between neighboring pedestrians. To analyze these interactions, we introduce an approach that estimates local influences in observational data on moving human crowds and represents them as spatially-embedded dynamic networks (visual influence networks). We analyzed data from a human "swarm" experiment (N = 10, 16, 20) in which participants were instructed to walk about the tracking area while staying together as a group. We reconstructed the network every 0.5 seconds using Time-Dependent Delayed Correlation (TDDC). Using novel network measures of local and global leadership (direct influence and branching influence), we find that both measures strongly depend on an individuals spatial position within the group, yielding similar but distinctive leadership gradients from the front to the back. There was also a strong linear relationship between individual influence and front-back position in the crowd. The results reveal that influence is concentrated in specific positions in a crowd, a fact that could be exploited by individuals seeking to lead collective crowd motion.

In-silico molecular enrichment and clearance of the human intracranial space

Causemann, M. — 2025-01-30

The mechanisms of intracranial solute transport are fundamental to human brain health, with alterations often linked to disease and functional impairment, and with distinct opportunities for personalized diagnostics and treatment. However, our understanding of these mechanisms and their interplay remains incomplete, in part due to the complexity of integrating insights across scales, between species and from different modalities. Here, we combine mixed-dimensional modelling, multi-modal magnetic resonance images, and high performance computing to construct and explore a high-fidelity in-silico model of human intracranial molecular enrichment. This model predicts the temporo-spatial spreading of a solute within an image-derived geometric representation of the subarachnoid space, ventricular system and brain parenchyma, including networks of surface perivascular spaces (PVSs). Our findings highlight the significant impact of cerebrospinal fluid (CSF) production and intracranial pulsatility on molecular enrichment following intrathecal tracer injection. We demonstrate that low-frequency vasomotion induces moderate CSF flow in surface PVS networks which substantially enhances tracer enrichment, and that impaired enrichment is a direct natural consequence of enlarged PVSs. This openly available technology platform thus provides an opportunity for integrating separate observations on diffusion in neuropil, vascular dynamics, intracranial pulsatility, CSF production, and efflux, and for exploring drug delivery and clearance in the human brain.

Improved Spatial Transcriptomics Clustering with Nested Graph Neural Networks

Jain, A. — 2025-02-08

We introduce a novel approach, STING, for spatial transcriptomic clustering analysis. Unlike existing state-of-the-art techniques that use graph-based neural networks (GNNs) trained on graphs generated from the spatial proximity of tissue locations (or spots), STING incorporates spot-specific related genes. This feature allows STING to better distinguish between clusters and identify meaningful gene-gene relations for knowledge discovery. It is a nested GNN framework that simultaneously models gene-gene and spatial relations. Using the gene expression, we generate a spot-specific gene-gene co-expression graph. We implement an inner GNN for these graphs to generate embeddings for each location. Next, we utilize these embeddings as features in a sample-wide graph generated using spatial information. We implement an outer GNN for this graph to reconstruct the original gene expression data. Finally, STING is trained end-to-end to generate embeddings that capture gene-gene and spatial information, which we input to a clustering algorithm to produce the spatial clusters. Experiments for 26 samples across 7 datasets and 5 spatial sequencing technologies show that STING outperforms the existing state-of-the-art techniques with a 1.58% to 4.07% improvement in the clustering evaluation metric, thus confirming that integrating gene-gene relation information with the clustering task leads to more informative embeddings and better clusters. Furthermore, experiments on a human breast cancer dataset show that STING identifies relevant genes and gene-gene relations, enabling biological hypothesis generation.

Collective Transitions from Orbiting to Matrix Invasion in 3D Multicellular Spheroids

Kim, J. — 2025-02-11

Coordinated cell rotation along a curved matrix interface can sculpt epithelial tissues into spherical morphologies. Subsequently, radially-oriented invasion of multicellular strands or branches can occur by local remodeling of the confining matrix. These symmetry-breaking transitions emerge from the dynamic reciprocity between cells and matrix, but remain poorly understood. Here, we show that epithelial cell spheroids collectively transition from circumferential orbiting to radial invasion via bi-directional interactions with the surrounding matrix curvature. Initially, spheroids exhibit an ellipsoidal shape but become rounded as orbiting occurs. However, cells gradually reorient from coordinated rotation towards outward strand invasion due to the accumulation of contractile tractions at discrete sites. Remarkably, the initial ellipsoid morphology predicts subsequent invasion of 2-4 strands roughly aligned with the major axis. We then perturb collective migration using osmotic pressure, showing that orbiting can be arrested and invasion can be reversed. We also investigate coordinated orbiting in "mosaic" spheroids, showing a small fraction of "leader" cells with weakened cell-cell adhesions can impede collective orbiting but still invade into the matrix. Finally, we establish a minimal self-propelled particle model to elucidate how collective orbiting is mediated by the crosstalk of cell-cell and cell-matrix adhesion along a curved boundary. Altogether, this work elucidates how tissue morphogenesis is governed by the interplay of collective behaviors and the local curvature of the cell-matrix, with relevance for embryonic development and tumor progression.

Mutations in TAC1B drive CDR1 and MDR1 expression and azole resistance in C. auris

Barker, K. S. — 2025-02-11

ObjectiveCandida auris has emerged as a fungal pathogen of particular concern owing in part to its propensity to exhibit antifungal resistance, especially to the commonly prescribed antifungal fluconazole. In this work we aimed to determine how mutations in the transcription factor gene TAC1B, which are common among resistant isolates and confer fluconazole resistance, exert this effect. MethodsSelected TAC1B mutations from clinical isolates were introduced into a susceptible isolate and reverted to the wild-type sequence in select clinical isolates using CRISPR Cas9 gene editing. Disruption mutants were likewise generated for select genes of interest. TAC1B mutants were subjected to transcriptional profiling by RNA-seq, and relative expression of specific genes of interest was determined by qRT-PCR. Antifungal susceptibilities were determined by modified CLSI broth microdilution. ResultsTAC1B mutations leading to A640V, A657V, and F862_N866del conferred fluconazole resistance, as well as increased resistance to other triazoles, when introduced into a susceptible isolate. RNA-seq revealed that the ATP-Binding Cassette (ABC) transporter gene CDR1 as well as the Major Facilitator Superfamily (MFS) transporter gene MDR1 were both upregulated by these TAC1B mutations. Disruption of CDR1 greatly abrogated resistance in strains with TAC1B mutations whereas disruption of MDR1 had little to no effect. However, disruption of both CDR1 and MDR1 resulted in an additional reduction in resistance as compared to disruption of either gene alone. ConclusionTAC1B mutations leading to A640V, A657V, and F862_N866del all result in increased resistance to fluconazole and other triazole antifungals, and increased expression of both CDR1 and MDR1 in C. auris. CDR1 is the primary driver of resistance conferred by these TAC1B mutations.

ML-MAGES: A machine learning framework for multivariate genetic association analyses with genes and effect size shrinkage

Liu, X. — 2025-02-12

A fundamental goal of genetics is to identify which and how genetic variants are associated with a trait, often using the regression results from genome-wide association (GWA) studies. Important methodological challenges are accounting for inflation in GWA effect estimates as well as investigating more than one trait simultaneously. We leverage machine learning approaches for these two challenges, developing a computationally efficient method called ML-MAGES. First, we shrink the inflation in GWA effect sizes caused by non-independence among variants using neural networks. We then cluster variant associations among multiple traits via variational inference. We compare the performance of shrinkage via neural networks to regularized regression and fine-mapping, two approaches used for addressing inflated effects but dealing with variants in focal regions of different sizes. Our neural network shrinkage outperforms both methods in approximating the true effect sizes in simulated data. Our infinite mixture clustering approach offers a flexible, data-driven way to distinguish different types of associations--trait-specific, shared across traits, or spurious--among multiple traits based on their regularized effects. Clustering applied to our neural network shrinkage results also produces consistently higher precision and recall for distinguishing gene-level associations in simulations. We demonstrate the application of ML-MAGES on association analyses of two quantitative traits and two binary traits in the UK Biobank (genetic and phenotypic data from 500,000 residents of the UK). Our identified associated genes from single-trait enrichment tests overlap with those having known relevant biological processes to the traits. Besides trait-specific associations, ML-MAGES identifies several variants with shared multi-trait associations, suggesting putative shared genetic architecture.

Disruption of Mitochondrial Dynamics and Stasis Leads to Liver Injury and Tumorigenesis

Ma, X. — 2025-02-12

Background & AimsMitochondrial dysfunction has been implicated in aging and various cancer development. As highly dynamic organelles, mitochondria constantly undergo fission, mediated by dynamin-related protein 1 (DRP1, gene name Dnm1l), and fusion, regulated by mitofusin 1 (MFN1), MFN2, and optic atrophy 1 (OPA1). However, whether and how dysregulation of mitochondria dynamics would be involved in liver pathogenesis and tumorigenesis is unknown. MethodsDnm1l Flox/Flox (Dnm1lF/F), Mfn1F/F and Mfn2F/F mice were crossed with albumin-Cre mice to generate liver-specific Dnm1l knockout (L-Dnm1l KO), L-Mfn1 KO, L-Mfn2 KO, L-Mfn1, Mfn2 double KO (DKO), and L-Mfn1, Mfn2, Dnm1l triple KO (TKO) mice. These mice were housed for various periods up to 18 months. Some mice also received hydrodynamic tail vein injections of a Sleeping Beauty transposon-transposase plasmid system with c-MYC and YAP. Blood and liver tissues were harvested for biochemical and histological analysis. ResultsL-Dnm1l KO mice had elevated serum alanine aminotransferase levels and increased hepatic fibrosis as early as two months of age. By 12 to 18 months, male L-Dnm1l KO mice developed spontaneous liver tumors, primarily hepatocellular adenomas. While female L-Dnm1l KO mice also developed liver tumors, their incidence was much lower. In contrast, neither L-Mfn1 KO nor L-Mfn2 KO mice had notable liver injury or tumorigenesis. However, a small portion of DKO mice developed tumors at 15-18 month-old. Increased DNA damage, senescence and compensatory proliferation were observed in L-Dnm1l KO mice but were less evident in L-Mfn1 KO, L-Mfn2 KO or DKO mice, indicating that mitochondrial fission is more important to maintain hepatocyte homeostasis and prevent liver tumorigenesis. Interestingly, further deletion of Mfn1 and Mfn2 in L-Dnm1l KO mice markedly abolished liver injury, fibrosis, and both spontaneous and oncogene-induced tumorigenesis. RNA sequencing and metabolomics analysis revealed significant activation of the cGAS-STING-interferon pathway and alterations in the tumor microenvironment pathways, alongside increased pyrimidine synthesis and metabolism in the livers of L-Dnm1l KO mice. Notably, the changes in gene expression and pyrimidine metabolism were considerably corrected in the TKO mice. ConclusionsMitochondrial dynamics and stability are essential for maintaining hepatic mitochondrial homeostasis and hepatocyte functions. Loss of hepatic DRP1 promotes liver tumorigenesis by increasing pyrimidine metabolism and activating the cGAS-STING-mediated innate immune response.

HKU5 bat merbecoviruses use divergent mechanisms to engage bat and mink ACE2 as entry receptors

Alfajaro, M. M. — 2025-02-13

Identifying receptors for bat coronaviruses is critical for spillover risk assessment, countermeasure development, and pandemic preparedness. While Middle East respiratory syndrome coronavirus (MERS-CoV) uses DPP4 for entry, the receptors of many MERS-related betacoronaviruses remain unknown. The bat merbecovirus HKU5 was previously shown to have an entry restriction in human cells. Using both pseudotyped and full-length virus, we show that HKU5 uses Pipistrellus abramus bat ACE2 but not human ACE2 or DPP4 as a receptor. Cryo-electron microscopy (cryo-EM) analysis of the virus-receptor complex and structure-guided mutagenesis reveal a spike and ACE2 interaction that is distinct from other ACE2-using coronaviruses. MERS-CoV vaccine sera poorly neutralize HKU5 informing pan-merbecovirus vaccine design. Notably, HKU5 can also engage American mink and stoat ACE2, revealing mustelids as potential intermediate hosts. These findings highlight the versatility of merbecovirus receptor use and underscore the need for continued surveillance of bat and mustelid species.

Mutations in ASH1L cause a neurodevelopmental disorder with sex differences in epilepsy and autism

Papendorp, C. M. — 2025-02-23

To understand brain phenotypes associated with ASH1L, we performed both studies in mouse models and clinical phenotyping of human subjects. We found in mice that ASH1L mutations result in seizures, microcephaly, and also less complex dendritic morphology. When we analyzed human subjects based for epilepsy, intellectual disability, and ASD, we found sex differences in epilepsy and autism, with epilepsy predominantly in female and ASD in male subjects. To understand the cellular and molecular mechanisms of the sex-difference, we performed whole cell patch clamp electrophysiology in mice and found hyperexcitability in female compared with male hippocampal CA1 neurons. We report the identification of sex-specific transcriptomic signatures resulting from ASH1L haploinsufficiency. Differentially expressed genes in female mice showed distinct association with epileptic encephalopathy, postnatal microcephaly and autistic behaviors. Thus, the role of ASH1L in specific circuits may be sex-dependent leading to sexual dimorphic effects from disruption of this gene.

3D spatial transcriptomics reveals the molecular domain structure of the mouse olfactory bulb

Klimpert, N. — 2025-02-24

A core organizing principle of the vertebrate brain is its symmetry along multiple axes. However, the precision with which neurons, circuit modules, and brain regions align to these axes remains poorly understood. Here, we used 3D spatial transcriptomics to reconstruct the anatomical and molecular organization of the mouse olfactory bulb. We mapped the positions of nearly one thousand molecularly distinct glomeruli, the structural and functional units of odor processing, revealing highly symmetric organization across hemispheres. Within each bulb, we delineated a curved axis of symmetry that divides pairs of sister glomeruli. Gene expression in the olfactory epithelium predicted glomerular position with near-glomerular resolution. However, glomerular symmetry did not extend to deeper layer mitral and granule cells, suggesting a reorganization from sensory input to cortical output pathways. Our findings provide the first comprehensive map of the olfactory bulb and reveal how its molecular structure is instructed by epithelial gene expression programs.

Attractor-based models for sequences and pattern generation in neural circuits

Londono Alvarez, J. — 2025-03-10

Neural circuits in the brain perform a variety of essential functions, including input classification, pattern completion, and the generation of rhythms and oscillations that support functions such as breathing and locomotion. There is also substantial evidence that the brain encodes memories and processes information via sequences of neural activity. Traditionally, rhythmic activity and pattern generation have been modeled using coupled oscillators, whereas input classification and pattern completion have been modeled using attractor neural networks. Here, we present a theoretical framework that demonstrates how attractor-based networks can also generate diverse rhythmic patterns, such as those of central pattern generator circuits (CPGs). Additionally, we propose a mechanism for transitioning between patterns. Specifically, we construct a network that can step through a sequence of five different quadruped gaits. It is composed of two dynamically distinct modules: a "counter" network, that can count the number of external inputs it receives via a sequence of fixed points; and a locomotion network, that encodes five different quadruped gaits as limit cycles. A sequence of locomotive gaits is obtained by connecting the counter network with the locomotion network. Specifically, we introduce a new architecture for layering networks that produces "fusion" attractors, binding pairs of attractors from individual layers. All of this is accomplished within a unified framework of attractor-based models using threshold-linear networks.

Microbiome functional gene pathways predict cognitive performance in older adults with Alzheimers disease

Zeamer, A. L. — 2025-03-11

Disturbances in the gut microbiome is increasing correlated with neurodegenerative disorders, including Alzheimers Disease. The microbiome may in fact influence disease pathology in AD by triggering or potentiating systemic and neuroinflammation, thereby driving disease pathology along the "microbiota-gut-brain-axis". Currently, drivers of cognitive decline and symptomatic progression in AD remain unknown and understudied. Changes in gut microbiome composition may offer clues to potential systemic physiologic and neuropathologic changes that contribute to cognitive decline. Here, we recruited a cohort of 260 older adults (age 60+) living in the community and followed them over time, tracking objective measures of cognition, clinical information, and gut microbiomes. Subjects were classified as healthy controls or as having mild cognitive impairment based on cognitive performance. Those with a diagnosis of Alzheimers Diseases with confirmed using serum biomarkers. Using metagenomic sequencing, we found that relative species abundances correlated well with cognition status (MCI or AD). Furthermore, gene pathways analyses suggest certain microbial metabolic pathways to either be correlated with cognitive decline or maintaining cognitive function. Specifically, genes involved in the urea cycle or production of methionine and cysteine predicted worse cognitive performance. Our study suggests that gut microbiome composition may predict AD cognitive performance.

A Brain-Computer Interface for Improving Auditory Attention in Multi-Talker Environments

Haro, S. — 2025-03-13

ObjectiveThere is significant research in accurately determining the focus of a listeners attention in a multi-talker environment using auditory attention decoding (AAD) algorithms. These algorithms rely on neural signals to identify the intended speaker, assuming that these signals consistently reflect the listeners focus. However, some listeners struggle with this competing talkers task, leading to suboptimal tracking of the desired speaker due to potential interference from distractors. The goal of this study was to enhance a listeners attention to the target speaker in real time and investigate the underlying neural bases of this improvement. ApproachThis paper describes a closed-loop neurofeedback system that decodes the auditory attention of the listener in real time, utilizing data from a non-invasive, wet electroencephalography (EEG) brain-computer interface (BCI). Fluctuations in the listeners real-time attention decoding accuracy was used to provide acoustic feedback. As accuracy improved, the ignored talker in the two-talker listening scenario was attenuated; making the desired talker easier to attend to due to the improved attended talker signal-to-noise ratio (SNR). A one-hour session was divided into a 10-minute decoder training phase, with the rest of the session allocated to observing changes in neural decoding. ResultsIn this study, we found evidence of suppression of (i.e., reduction in) neural tracking of the unattended talker when comparing the first and second half of the neurofeedback session (p = 0.012). We did not find a statistically significant increase in the neural tracking of the attended talker. SignificanceThese results establish a single session performance benchmark for a time-invariant, non-adaptive attended talker linear decoder utilized to extract attention from a listener integrated within a closed-loop neurofeedback system. This research lays the engineering and scientific foundation for prospective multi-session clinical trials of an auditory attention training paradigm.

Adaptive learning via BG-thalamo-cortical circuitry

He, Q. — 2025-03-13

1People adjust their use of feedback over time through a process referred to as adaptive learning. We have recently proposed that the underlying mechanisms of adaptive learning are rooted in how the brain organizes time into similarly credited units, which we refer to as latent states. Here we develop a BG-thalamo-cortical circuit model of this process and show that it captures both the commonalities and heterogeneity in human adaptive learning behavior. Our model learns incrementally through synaptic plasticity in PFC-BG connections, but upon observing discordant information, produces thalamocortical reset signals that alter PFC connectivity, driving attractor state transitions that facilitate rapid updating of behavioral policy. We demonstrate that this mechanism can give rise to optimized learning dynamics in the context of either changepoints or reversals, and that under reasonable biological assumptions the model is able to generalize efficiently across these conditions, adjusting behavior in a context-appropriate manner. Taken together, our results provide a biologically plausible mechanistic model for adaptive learning that explains existing behavioral data and makes testable predictions about the computational roles of different brain regions in complex learning behaviors.

An archaic reference-free method to jointly infer Neanderthal and Denisovan introgressed segments in modern human genomes

Planche, L. — 2025-03-17

Admixture between populations is a common feature of human history. Admixture events introduce new genetic variation that can fuel evolution. Characterizing the significance of admixture events on the evolution of a population across various species is of great interest to evolutionary geneticists. Local Ancestry Inference (LAI) methods infer genetic ancestry of an individual at a particular chromosomal location. Certain methods specialize in detecting archaic introgression, which consists of interbreeding between modern and archaic humans like Neanderthals and Denisovans. Most current LAI methods allow the detection of a single archaic ancestry, and post-processing may distinguish between multiple waves of introgression. These methods vary in how they choose archaic or modern reference genomes for the inference. Here, we present a new HMM-based method (DAIseg), which has the advantage of simultaneously distinguishing between multiple waves of ancient and recent admixture, using only modern human reference genomes. Simulations demonstrate that DAIseg achieves higher overall performance than state-of-the-art methods. We also apply DAIseg to Papuan populations to jointly detect Denisovan and Neanderthal introgressed segments, and identify a higher number of archaic segments than previous methods. Analysis of inferred introgressed segments, shows that we can identify evidence for two Denisovan introgression events in Papuans without having any post-processing and filtering. Overall, on top of being able to deal with both Archaic and recent admixture, DAIseg provides a more principled approach for detecting and classifying Denisovan and Neanderthal segments which will improve downstream analysis of introgressed segments to infer the impact of archaic introgression in humans.

Multi-omic Evaluations Nominate an ER-Mitochondrial Axis and Inflammatory Macrophage as Drivers of Right Atrial Dysfunction

Mendelson, J. B. — 2025-03-25

BackgroundRight atrial (RA) dysfunction is an emerging risk factor for poor outcomes in pulmonary arterial hypertension, however the mechanisms underlying compromised RA function are understudied. ObjectivesMulti-omic analyses defined the cellular and molecular mediators associated with RA dysfunction in pulmonary artery banded (PAB) swine. Methods4-week-old castrated male Yorkshire pigs were subjected to PAB and aged six weeks to induce right heart failure. Cardiac MRI evaluated RA size and function. snRNAseq defined the cell-specific alterations in RA tissue. Mitochondrial proteomics and metabolomics analyses examined the metabolic alterations in RA samples. Inducible pluripotent stem cell-derived atrial cardiomyocytes (iPSC-ACM) were treated with tunicamycin to induce endoplasmic reticulum (ER) stress and mitochondrial structure and function were probed. ResultsPAB induced RA dilation/dysfunction and atrial cardiomyocyte hypertrophy. snRNAseq demonstrated PAB altered the cellular composition of the RA defined by increased inflammatory macrophages and an alteration of cardiomyocyte subpopulations. RA cardiomyocytes exhibited ER stress and mitochondrial metabolic enzyme dysregulation. PAB RAs, but not PAB right ventricles, had downregulation of branched chain amino acid degrading enzymes. Metabolomics profiling revealed BCAA and fatty acid metabolism were impaired in the dysfunctional RA. Tunicamycin-induced ER stress disrupted mitochondrial structure/function in iPSC-ACMs. ConclusionsMulti-omic evaluations demonstrate RA dysfunction is characterized by cardiomyocyte metabolic derangements due to ER dysregulation and an accumulation of pro-inflammatory macrophages.

Global Metagenomics Reveals Hidden Protist Diversity

Gutierrez, M. F. R. — 2025-03-26

Protists, defined as eukaryotes distinct from animals, plants, and fungi, are a polyphyletic group that dominates the eukaryotic tree of life, exhibiting significant phylogenetic diversity and fulfilling critical ecological roles. Historically, research has prioritized protists associated with animals and plants, particularly those of medical significance, thereby overlooking the majority of protist diversity. Conventional molecular methods, such as 18S rRNA gene amplicon sequencing, frequently encounter limitations, including primer binding bias and PCR bias caused by gene length variations, resulting in a biased representation of protistan diversity. Further, most protist lineages are notoriously difficult to cultivate. Here, we analyzed over 30,000 metagenome assemblies and protist single-cell genomes, together with 21 long-amplicon 18S datasets, spanning marine, freshwater, and soil ecosystems. We recovered 157,956 18S rRNA gene sequences ([≥]800 bp), which clustered into 103,338 operational taxonomic units (OTUs) at 97% sequence identity and 24,438 OTUs at 85% identity. Notably, 69% of 13,238 non-singleton clusters at 85% identity consisted exclusively of environmental sequences, uncovering a wealth of novel, uncultivated, and unclassified protist diversity. A comprehensive taxonomic framework of eukaryotes based on concatenated 18S and 28S rRNA genes that incorporated most novel lineages revealed substantial underrepresentation of Amoebozoa, Discoba, and Rhizaria in reference databases, with many lacking isolate or genome sequence representation. Further, we identified 13 lineages that likely represent deeply branching diversity, including candidates at approximately class- to phylum-level depth, that lack representation in public databases. The corresponding 85% OTUs were primarily affiliated with Excavata, with some branching deeply in the eukaryotic tree. Comprehensive analysis of the global distribution of eukaryotes revealed uneven microbial eukaryotic diversity across supergroups and ecosystems, with notable novelty particularly in soil and marine environments. We then examined co-occurrence between protists and prokaryotes, predicting putative symbiotic or predator-prey relationships, particularly among understudied protist groups with bacteria such as Verrucomicrobia and Rickettsiales. Our results extend current reference coverage and provide a global, contig-based framework for protistan diversity and distribution, highlighting major gaps in curated databases and metabarcoding coverage and guiding targeted studies of these organisms ecological roles.

Neutrophil progenitor cell therapy rescues host defense against Staphylococcus aureus in murine chronic granulomatous disease

Hinman, K. D. — 2025-03-30

Despite advances in engineered adaptive immune cell therapies, current options for innate immune cell therapies are sparse. In this work, we demonstrate the utility of a neutrophil progenitor-based cell therapy. Murine conditionally-immortalized neutrophil progenitors (NPs) overcome the hurdles of alternative cell therapies by engrafting in the unconditioned host and substantially contributing to the host neutrophil population. Here we demonstrate the therapeutic value of NPs using a murine model of the primary immunodeficiency chronic granulomatous disease (CGD). Those with CGD are highly susceptible to infection with Staphylococcus aureus because of genetic mutations that impair neutrophil antimicrobial function. We find that the prophylactic treatment of CGD mice with transfused NPs rescue them from an otherwise lethal S. aureus pulmonary infection. In investigating the mechanisms behind the improved clearance of S. aureus and survival of CGD mice, our data suggests that the antimicrobial function of host CGD neutrophils is rescued by the presence of donor-derived wild-type neutrophils. We also observe that survival is improved to >50% in the CGD model when mice receive NPs post-infection. This work highlights the application of NPs to improving outcomes to acute bacterial infection in CGD, demonstrating the translational potential of conditionally-immortalized myeloid progenitors as a cellular therapy.

Relative contributions of the ERG11VF125AL and MRR1AN647T mutations to fluconazole resistance in Clade III Candidozyma (Candida) auris clinical isolates

Barker, K. S. — 2025-03-31

ObjectivesCandidozyma (Candida) auris is an emerging fungal pathogen of global concern that often exhibits multi-drug resistance. Over 90% of isolates are resistant to fluconazole. Of the six described clades of C. auris, Clade III has been found to be nearly universally fluconazole resistant and almost every Clade III isolate described carries a mutation in the gene encoding the fluconazole target sterol demethylase (ERG11) leading to a VF125AL substitution and a mutation leading to a N647T substitution in the gene encoding Mrr1a, a transcriptional regulator of the Mdr1 transporter. Both mutations have been shown to contribute to fluconazole resistance in C. auris. MethodsIn the present study we introduced the Clade III MRR1A mutation into a Clade I background using CRISPR-Cas9 gene editing. In two Clade III clinical isolates we corrected the native MRR1A and ERG11 mutations to their wild-type sequences as well as disrupted MDR1. Triazole susceptibilities and MDR1 gene expression were measured in all strains. ResultsIntroduction of the N647T substitution in a Clade I background confers a modest reduction in fluconazole and voriconazole susceptibility. Similarly, correction of MRR1A or disruption of MDR1 in each Clade III background resulted in a one-dilution decrease in fluconazole and voriconazole MIC while the ERG11 correction resulted in a three-dilution decrease in fluconazole and voriconazole MIC. ConclusionsOur findings show that while the MRR1A mutation makes a modest contribution, the ERG11 mutation is responsible for most of the fluconazole resistance observed in Clade III isolates. We also show that while these mutations likewise affect voriconazole susceptibility, they have no effect on susceptibility to itraconazole, isavuconazole, or posaconazole suggesting the potential therapeutic utility of these antifungals for infections due to Clade III isolates of C. auris.

A transcriptional biosensor reveals mechanisms of α-ketoglutarate signaling to chromatin

Sternisha, A. C. — 2025-04-07

Alpha-ketoglutarate (KG) is required for chromatin demethylation but mechanisms controlling KG abundance in the nucleus are poorly defined. Therefore, we designed a biosensor system to monitor this metabolite pool in human cells using an KG-responsive cyanobacterial transcription factor, NtcA. We then coupled this system with a genetic screen to identify genes that regulate KG in the nucleus, defining an inter-organelle pathway in which sequential mitochondrial activities of the GPT2 transaminase and SLC25A11 transporter supply nuclear KG. Using a mouse model of GPT2 deficiency, a human inborn error of metabolism, we found that this pathway controls chromatin methylation in the developing brain. Our work provides a tool to assess KG signaling to chromatin and a framework for leveraging forward genetics to study nuclear metabolite pools.

DILP8 serves as a mature follicle sensor to prevent excessive accumulation of maturefollicles in Drosophila ovaries and oocyte aging.

Oramas, R. — 2025-04-08

Excessive mature follicle accumulation in ovaries harms oocyte health and offspring viability. As such, the number of mature follicles in ovaries is tightly controlled. In Drosophila, each ovary is comprised of [~]16 ovarioles, each containing 1-2 mature follicles, regardless of the females mating status. The mechanism by which the female flies count the number of mature follicles to coordinate egg release and oogenesis remains a mystery. Previous work, along with our RNAseq and antibody analysis, demonstrated that Drosophila insulin-like peptide 8 (DILP8) is expressed in somatic follicle cells of mature follicles but not in younger follicles. Contrary to previous findings, we found that DILP8 is not essential for mating-induced ovulation/egg laying. In contrast, global depletion or follicle-cell-specific knockdown of dilp8 leads to defective ovulation/egg laying and a significant increase of mature follicles in virgin females. In addition, we found that excessive accumulation of mature follicles in dilp8-knockdown females leads to poor oocyte quality. Furthermore, knockdown of Lgr3 (Leucine-rich repeat-containing G protein-coupled receptor 3), encoding a previously identified DILP8 receptor, showed similar ovulation/egg laying defects, accumulation of mature follicles, and poor oocyte quality in virgin females. Therefore, DILP8 functions as a mature follicle sensor to prevent excessive accumulation of mature follicles and maintain oocyte quality through the Lgr3 receptor in virgin females. Because mating is not always available in the wild and the DILP8/Lgr3 pathway is highly conserved across multiple species, our findings suggest that DILP8/Lgr3 is likely critical for maintaining the optimal reproductive fitness of virgin females and for species survival in the wild.

scContrast: A contrastive learning based approach for encoding single-cell gene expression data

Li, W. Y. — 2025-04-14

Single-cell RNA sequencing (scRNA-seq) captures gene expression at a individual cell resolution, which reveals critical insights into cellular diversity, disease processes, and developmental biology. However, a key challenge in scRNA-seq analysis is clustering similar cells across multiple batches, particularly when distinct sequencing protocols are used. In this work, we present scContrast, a semi-supervised contrastive learning method tailored for embedding scRNA-seq data from both plate- and droplet-based protocols into a universal representation space. By leveraging five simple augmentations, scContrast extracts biologically relevant signals from gene expression data while filtering out batch effects and technical artifacts. We trained scContrast on a subset of Tabula Muris tissues and evaluated its zero-shot performance on unseen tissues. Our results demonstrate that scContrast generalizes effectively to new tissues and outperforms the leading UCE approach in integrating scRNA-seq data from droplet- and plate-based sequencing protocols.

Sex differentiation, divergence, and introgression in the leishmaniasis vector Lutzomyia longipalpis

Stuckert, A. M. — 2025-04-14

An estimated one billion people are at risk of acquiring the disease leishmaniasis annually. This disease, among others, is transmitted to humans via sandfly vectors of the family Psychodidae. In the Neotropics, Lutzomyia longipalpis is one of the most common vectors of leishmaniasis, but little is known about its genetics and population structure, hampering control efforts. We generated sex-specific genome assemblies of Lu. longipalpis from Colombia, and examined population genomics of this species. We found that sex accounts for [~]11% of genome-wide variation, primarily driven by differentiation between the sexes in an XY sex chromosome. Analyses of transposable elements indicate that the size difference in the sex chromosome is driven by more transposable elements in the male chromosome. Further, we examined population genomics and structure across the South American continent. We identified significant genetic divergence between Colombian and Brazilian sandflies and population structure within Lu. longipalpis sensu lato samples collected from the same sites in Brazil. These results strongly suggest the presence of multiple, sympatric species within a cryptic species complex, concordant with other published phenotypic evidence. SIGNIFICANCELeishmaniasis has emerged as an important public health issue. Transmission occurs exclusively through Phlebotomine sandflies of the genera Lutzomyia and Phlebotomus, yet our ability to prevent its spread is constrained by limited knowledge of the biological and ecological factors that make these insects effective vectors. In this manuscript, we leverage whole-genome sequencing to investigate the evolutionary forces shaping genetic diversity in Lutzomyia longipalpis, a key vector of leishmaniasis.

Scaling Large Language Models for Next-Generation Single-Cell Analysis

Rizvi, S. A. — 2025-04-17

AO_SCPLOWBSTRACTC_SCPLOWSingle-cell RNA sequencing has transformed our understanding of cellular diversity, yet current single-cell foundation models (scFMs) remain limited in their scalability, flexibility across diverse tasks, and ability to natively integrate textual information. In this work, we build upon the Cell2Sentence (C2S) framework, which represents scRNA-seq profiles as textual "cell sentences," to train Large Language Models (LLMs) on a corpus comprising over one billion tokens of transcriptomic data, biological text, and metadata. Scaling the model to 27 billion parameters yields consistent improvements in predictive and generative capabilities and supports advanced downstream tasks that require synthesis of information across multi-cellular contexts. Targeted fine-tuning with modern reinforcement learning techniques produces strong performance in perturbation response prediction, natural language interpretation, and complex biological reasoning. This predictive strength enabled a dual-context virtual screen that nominated the kinase inhibitor silmitasertib (CX-4945) as a candidate for context-selective upregulation of antigen presentation. Experimental assessment in human cell models unseen during training supported this prediction, demonstrating that C2S-Scale can effectively guide the discovery of context-conditioned biology. C2S-Scale unifies transcriptomic and textual data at unprecedented scales, surpassing both specialized single-cell models and general-purpose LLMs to provide a platform for next-generation single-cell analysis and the development of "virtual cells."

Characterizing Clinical Toxicity in Cancer Combination Therapies

Wong, A. M. — 2025-04-19

AO_SCPLOWBSTRACTC_SCPLOWPredicting synergistic cancer drug combinations through computational methods offers a scalable approach to creating therapies that are more effective and less toxic. However, most algorithms focus solely on synergy without considering toxicity when selecting optimal drug combinations. In the absence of combinatorial toxicity assays, a few models use toxicity penalties to balance high synergy with lower toxicity. However, these penalties have not been explicitly validated against known drug-drug interactions. In this study, we examine whether synergy scores and toxicity metrics correlate with known adverse drug interactions. While some metrics show trends with toxicity levels, our results reveal significant limitations in using them as penalties. These findings highlight the challenges of incorporating toxicity into synergy prediction frameworks and suggest that advancing the field requires more comprehensive combination toxicity data.

Sleep is associated with reduction of excitatory signaling in medial prefrontal cortex

Yamada, T. — 2025-05-01

Although many sleep medications enhance inhibitory signaling, it remains unclear whether inhibitory or excitatory neurotransmitters contribute to the natural transition from wakefulness to sleep in humans. Here, we show that changes in excitatory, rather than inhibitory, neurotransmitter levels are associated with this transition. Young, healthy participants underwent two nap sessions with polysomnography, during which glutamate and GABA concentrations in the medial prefrontal cortex were measured using magnetic resonance spectroscopy. Glutamate gradually decreased during deeper sleep stages compared to wakefulness in the second session, with better sleep quality. No such change occurred in the first session with poorer sleep, likely due to the first-night effect. Furthermore, reduced glutamate significantly mediated sleep-onset latency in both sessions. Conversely, GABA concentration did not change from wakefulness to sleep in either session. These findings provide the first evidence that reduced excitatory signaling is a key feature of natural good sleep onset in the human brain.

Anxiety state-related task disengagement varies with trait anxiety

Sayali, C. — 2025-05-07

Cognitively demanding tasks are often perceived as costly due to the cognitive control resources they require, leading to effort avoidance, particularly in psychiatric populations with motivational impairments. Research on anxiety and cognitive effort are mixed: some studies suggest anxiety increases the perceived effort cost and avoidance, while others indicate that cognitive effort engagement can serve as an adaptive coping strategy. To reconcile these perspectives, we examined the interaction between state and trait anxiety on cognitive effort evaluation and engagement in two experiments. We hypothesized that state anxiety enhances task engagement as difficulty increases, and that this effect is diminished in individuals with high trait anxiety. Experiment 1 assessed self-reported anxiety in an online sample, while Experiment 2 manipulated state anxiety through autobiographical recall. Both experiments employed flow induction and effort discounting paradigms. Across both studies, the effect of state anxiety on task engagement depended on trait anxiety, but the direction of the state anxiety effect was opposite to the effect we predicted. In Experiment 1, participants with low trait anxiety reported reduced task engagement, as indexed by lower flow scores, when state anxiety was higher, but only in easy tasks. This effect was attenuated in participants with higher trait anxiety. The same pattern was observed in Experiment 2, but this time the interaction between trait and state anxiety was present regardless of task difficulty. These findings suggest that trait anxiety may reflect reduced impact of state anxiety on task disengagement. Public significance statementThis study demonstrated that effects of state anxiety on task disengagement depend on individual differences in trait anxiety. People with higher trait anxiety reported reduced effects of state anxiety on task disengagement.

Neural and behavioral signatures of policy compression in cognitive control

Liu, S. — 2025-05-07

Making context-dependent decisions incurs cognitive costs. Cognitive control studies have investigated the nature of such costs from both computational and neural perspectives. In this paper, we offer an information-theoretic account of the costs associated with context-dependent decisions. According to this account, the brains limited capacity to store context-dependent policies necessitates "compression" of policies into internal representations with an upper bound on code length, quantified by an information-theoretic measure (policy complexity). These representations are decoded into actions by sequentially inspecting each bit, such that longer codes take more time to decode. When a response deadline is imposed, the account predicts that policy complexity should increase with the deadline. Higher policy complexity is associated with several behavioral signatures: (i) higher accuracy; (ii) lower variability; and (iii) lower perseveration. Analyzing data from a rule-based action selection task, we found evidence supporting all of these predictions. We further hypothesized that complex policies require higher neural dimensionality (which constrains the code space). Consistent with this hypothesis, we found that policy complexity correlates with a measure of neural dimensionality in a rule-based decision task. This finding brings us a step closer to understanding the neural implementation of policy compression and its implications for cognitive control.

A spatial code governs olfactory receptor choice and aligns sensory maps in the nose and brain

Brann, D. H. — 2025-05-08

Although topographical maps organize many peripheral sensory systems, it remains unclear whether olfactory sensory neurons (OSNs) choose which of the [~]1100 odor receptors (ORs) to express based upon their spatial location in the olfactory epithelium (OE) or instead ORs are scattered randomly. Here we reveal that each OR is expressed at a precise mean position along the OE dorsoventral axis, thereby instantiating a receptor map. This patterning reflects the differential use, by precursors and mature OSNs, of a coherent gene expression program controlled by a spatially-varying retinoic acid gradient; this program -- which includes key transcription factors and axon guidance genes -- translates position into a spatially appropriate distribution of OR choices and aligns the epithelial map of OR identity with the glomerular map present in the olfactory bulb. These results identify a transcriptional code that distinguishes and spatially organizes the vast array of sensory channels that comprise the olfactory system.

Multiomic profiling of hypoxic glioblastoma stem cells reveals expansion of subpopulations with distinct epigenetic and CNV profiles

Corseri, A. — 2025-05-09

Glioblastoma is characterized by extensive intratumoral heterogeneity driven by dynamic mi-croenvironmental cues such as hypoxia. While transcriptional and epigenetic variability have been separately linked to hypoxia responses, the integrated impact of hypoxia on gene regulation and clonal architecture in glioblastoma stem cells (GSCs) remains poorly defined. We applied singlenucleus multi-omics--integrating RNA-seq and ATAC-seq--to patient-derived GSCs cultured under normoxic or hypoxic conditions. This enabled simultaneous profiling of gene expression and chromatin accessibility within the same cells. Transcription factor (TF) regulatory networks were inferred using Dictys, while RNA-chromatin dynamics were modeled with MultiVelo. Clonal structure and copy number variations (CNVs) were resolved at single-cell resolution using RIDDLER on snATAC-seq data. Hypoxia induced the emergence of four distinct GSC subpopulations with unique transcriptomic and epigenetic profiles enriched for mesenchymal, angiogenic, and proliferative signatures. Regulatory network modeling revealed novel hypoxia-associated TFs--SP2, CREM, and ETV3--that modulate downstream oncogenic pathways. Trajectory analysis uncovered hypoxia-driven reversals in RNA-chromatin coupling, revealing dysregulated future transcriptional states of key genes such as MMP16 and SVIL. CNV profiling identified 13 clonal substructures, with specific clones (e.g., 5, 6, 9) selectively enriched under hypoxia and harboring distinct chromosomal alterations. These results demonstrate coordinated remodeling of GSC gene regulation and clonal fitness in response to hypoxic stress. Our findings reveal that hypoxia drives concurrent epigenetic, transcriptomic, and clonal selection in glioblastoma stem cells. This integrated model of hypoxia-induced plasticity provides mechanistic insights into tumor adaptation and identifies novel regulators that may serve as targets for therapeutic intervention in the hypoxic niche of glioblastoma.

The phosphatases TCPTP, PTPN22, and SHP1 play unique roles in T cell phosphotyrosine maintenance and feedback regulation of the TCR

Callahan, A. — 2025-05-09

The protein tyrosine phosphatases (PTPs) TCPTP, PTPN22, and SHP1 are critical regulators of the activating phosphotyrosine (pY) site on the initiating T cell kinase, LckY394. Still, the broader implications of these phosphatases in T cell receptor (TCR) signalling and T cell biology remain unclear. By combining CRISPR/Cas9 gene editing and mass spectrometry, we evaluate the protein- and pY-level effects of TCPTP, PTPN22, and SHP1 in the Jurkat T cell model system. We find that deletion of each phosphatase corresponds to unique changes in the proteome of T cells, with few large-scale changes to TCR signalling proteins. Notably, PTPN22 and SHP1 deletions have opposing effects on pY abundance globally, while TCPTP deletion modestly elevates pY levels. Finally, we show that TCPTP is indirectly involved in Erk1/2 positive feedback to the TCR. Overall, our work provides evidence for alternative functions of three T cell phosphatases long thought to be redundant.

Mutations in transcription factors that confer fluconazole resistance also confer reduced susceptibility to manogepix in Candida auris, Candida albicans, Candida parapsilosis, and Candida glabrata (Nakaseomyces glabratus)

Barker, K. S. — 2025-05-09

The fungal pathogen Candida auris is of global concern due to high levels of multidrug resistance and its propensity to cause infectious outbreaks. Over 90% of isolates are resistant to fluconazole, the most commonly prescribed antifungal world-wide. Fluconazole resistance is multifactorial with many isolates carrying mutations in the gene encoding the transcriptional regulator Tac1B, leading to increased expression of the gene encoding the ATP Binding Cassette (ABC) transporter Cdr1. Recently, a study examining C. auris in vitro resistance mechanisms to manogepix, a promising antifungal agent currently in clinical trials, found a TAC1B mutation that confers reduced manogepix and fluconazole susceptibility. We hypothesized that mutations in C. auris TAC1B and similar transcription factors in other Candida species that confer fluconazole resistance might also confer reduced susceptibility to manogepix. We measured manogepix susceptibilities for selected isolates and strains and found C. auris TAC1B, C. albicans and C. parapsilosis TAC1, and C. glabrata PDR1 confer reduced manogepix susceptibility in a manner dependent on ABC transporters similar to Cdr1. Our findings raise the possibility of fluconazole and manogepix cross resistance for clinical isolates harboring mutations in these genes.

Characterization of hyperoxia-induced senescent lung macrophages in neonatal mice

Lin, F. — 2025-05-10

Bronchopulmonary dysplasia (BPD), a chronic lung disease in premature infants, results from mechanical ventilation and hyperoxia amongst other factors. We and others have shown that neonatal hyperoxia, known to lead to a BPD-like phenotype in rodent models, causes lung cellular senescence. In our 3-day hyperoxia model, the majority of senescent cells were lung macrophages, and these peaked at postnatal day (pnd) 7. The features of these senescent macrophages are not characterized. Here, we reanalyzed scRNA-seq datasets (GSE207866) of senescent lung cells from mice exposed to hyperoxia as neonates at pnd7 (SD7), and characterized their gene express profiling and compared them with air control (AirD7) and hyperoxia-exposed group without isolation of senescent cells at pnd7 (O2D7). We first classified the cells into epithelial, endothelial, immune, and mesenchymal cells to extract immune cells. By employing the workflow to the immune clusters, mixed populations of macrophages, monocytes, and dendritic cells and pure population of macrophages, we finally identified seven clusters of macrophages. In SD7 group, 65.9% senescent cells were macrophages, and comprised M1 (64%) and alveolar (62.8%) macrophages. Clusters 0 and 1 were M1 and alveolar macrophages, which were composed of 49.2% of senescent macrophages. These two clusters highly expressed genes involved in innate immunity, inflammation, DNA repair response and phagocytosis. Metabolic switch from mitochondrial respiration to glycolysis and pentose phosphate pathway was observed in these two clusters. Conclusively, senescent macrophages are heterogenous with distinct tissue compartments and metabolic dysregulation.

Trpv1-dependent Cacna1b gene inactivation reveals cell-specific functions of CaV2.2 channels in vivo

Meir, R. Y. — 2025-05-10

Voltage-gated CaV2.2 channels underlie the N-type current, and they regulate calcium entry at many presynaptic nerve endings to control transmitter release. A role for CaV2.2 channels has been well-established in the transmission of pain information using pharmacological and global gene inactivated mouse models. However, investigation of the cell-specific actions of CaV2.2 channels would benefit from the availability of cell-restricted knockout mouse models and particularly in dissecting behavioral responses that depend on CaV2.2 channel activity. Here, we show the importance of CaV2.2 channels in Trpv1-lineage neurons in behavioral responses to sensory stimuli using Cre-dependent inactivation of the Cacna1b gene. Our work shows the cell- type specificity of CaV2.2 channels in mediating rapidly developing heat hypersensitivity and the utility of Cre-dependent inactivation of Cacna1b to discern cell-specific CaV2.2 channel functions.

FastConformation: A Standalone ML-Based Toolkit for Modeling and Analyzing Protein Conformational Ensembles at Scale

Galeazzi, F. M. — 2025-05-14

Deep learning approaches like AlphaFold 2 (AF2) have revolutionized structural biology by accurately predicting the ground state structures of proteins. Recently, clustering and subsampling techniques that manipulate multiple sequence alignment (MSA) inputs into AlphaFold to generate conformational ensembles of proteins have also been proposed. Although many of these techniques have been made open source, they often require integrating multiple packages and can be challenging for researchers who have a limited programming background to employ. This is especially true when researchers are interested in subsampling to produce predictions of protein conformational ensembles, which require multiple computational steps. This manuscript introduces FastConformation, a Python-based application that integrates MSA generation, structure prediction via AF2, and interactive analysis of protein conformations and their distributions, all in one place. FastConformation is accessible through a user-friendly GUI suitable for non-programmers, allowing users to iteratively refine subsampling parameters based on their analyses to achieve diverse conformational ensembles. Starting from an amino acid sequence, users can make protein conformation predictions and analyze results in just a few hours on their local machines, which is significantly faster than traditional molecular dynamics (MD) simulations. Uniquely, by leveraging the subsampling of MSAs, our tool enables the generation of alternative protein conformations. We demonstrate the utility of FastConformation on proteins including the Abl1 kinase, LAT1 transporter, and CCR5 receptor, showcasing its ability to predict and analyze the protein conformational ensembles and effects of mutations on a variety of proteins. This tool enables a wide range of high-throughput applications in protein biochemistry, drug discovery, and protein engineering.

Absence of Systematic Effects of Internalizing Psychopathology on Learning Under Uncertainty

Satti, M. H. — 2025-05-15

Difficulties in adapting learning to meet the challenges of uncertain and changing environments are widely thought to play a central role in internalizing psychopathology, including anxiety and depression. This view stems from findings linking trait anxiety and transdiagnostic internalizing symptoms to learning impairments in laboratory tasks often used as proxies for real-world behavioral flexibility. These tasks typically require learners to adjust learning rates dynamically in response to uncertainty, for instance, increasing learning from prediction errors in volatile environments. However, prior studies have produced inconsistent and sometimes contradictory findings regarding the nature and extent of learning impairments in populations with internalizing disorders. To address this, we conducted eight experiments (N = 820) using predictive inference and reversal learning tasks, and applied a bi-factor analysis to capture internalizing symptom variance shared across and differentiated between anxiety and depression. While we observed robust evidence for adaptive learning-rate modulation across participants, we found no convincing evidence of a systematic relationship between internalizing symptoms and either learning rates or task performance. These findings challenge prominent claims that learning difficulties are a hallmark feature of internalizing psychopathology and suggest that the relationship between these traits and adaptive behavior under uncertainty may be more subtle than previously thought.

Sex differences in DNA methylation in bats

Rayner, J. G. — 2025-05-15

Sex-biased longevity is observed across a wide range of animal taxa, including bats, for reasons not well understood. Patterns of cytosine methylation vary predictably with age in many organisms, offering a valuable means to investigate differences in patterns of aging at the molecular level. We tested sex differences in cytosine methylation across 14 bat species and compared patterns of age-associated variation. Sex differences were overrepresented on the X chromosome, showing a strong pattern of female hypermethylation within promoter regions. Sex and age-associated differences in methylation were non-randomly distributed with respect to proximity to putative sex hormone receptor binding sites, with sites hypermethylated in males and females tending to be underrepresented near androgen and estrogen receptor binding sites, respectively. Across species, we observed the relative steepness of male versus female slopes of age-associated variation was associated with the strength of precopulatory sexual selection, with especially strong trends towards male-biased age-associated slopes in two harem-polygynous species that exhibit female-biased longevity. Our results offer insights into how patterns of methylation differ across sexes and ages, and raise intriguing questions for future research, such as whether sex differences in molecular aging reflect sex-biased longevity, for which records in bats are sparse.

The temporal and perceptual characteristics of emotion-induced blindness

Alarie, M. E. — 2025-05-17

Attentional capture by emotionally salient stimuli is adaptive, permitting identification of possible threats; however, an excessive bias towards emotional stimuli can interrupt goal-directed behavior. This is especially relevant in psychiatric disease, where severe emotional distress can interfere with daily function. As such, understanding the mechanisms by which emotional stimuli compete for attentional resources is a critical area of investigation. Previous studies using rapid serial visual presentation (RSVP) paradigms observe that emotional distractors disrupt the detection of subsequent stimuli, referred to as emotion-induced blindness (EIB). Our study expands upon this work, characterizing how temporal and perceptual factors shape the emergence and intensity of EIB. Contrary to previous assumptions regarding temporal dynamics of EIB, we found that effects of emotional distractors persisted across prolonged image presentation durations. Further, we investigated the extent to which the depth of distractor processing influences EIB using a distractor recall task. While recall was predictive of EIB magnitude, a significant effect of emotional distractors on target detection was nonetheless present even without conscious recall of the distractor. These findings demonstrate the robustness of the EIB effect in RSVP in the context of temporal and perceptual manipulations.

When good guides go bad: empirical evaluation of all unique Cas9 targets in E. coli reveal widespread functionality and rules for gRNA biological design

Phillips, E. K. — 2025-05-23

The Cas9 nuclease has become central to modern methods and technologies in synthetic biology, largely due to the ease in which it can be targeted to specific DNA loci via guide RNAs (gRNAs). Reports vary widely on the actual specificity of this targeting, with some studies observing 60% of gRNAs possessing no activity against the genome, yet an assumption that inactive gRNAs are rare persists in the E. coli community. To resolve these contradictions, we evaluated the activity of nearly 500,000 unique gRNAs in the E. coli K12 MG1655 genome. We show that the overwhelming majority (at least 93%) of unique gRNAs are functional while only 0.3% are nonfunctional.These nonfunctional gRNAs exhibit strong spacer self-interaction, which can be either excluded using a simple design rule or "repaired" during library design. Finally, this work provides the greater microbial synthetic biology community both a set of nearly half a million E. coli gRNAs that have been empirically evaluated in vivo as well as a thoroughly evaluated experimental procedure, complete with appropriate controls for Cas9 activity, for conducting Cas9 assays in E. coli specifically and bacteria more generally. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/651106v3_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@1038ddcorg.highwire.dtl.DTLVardef@566e37org.highwire.dtl.DTLVardef@5df6b2org.highwire.dtl.DTLVardef@ef453c_HPS_FORMAT_FIGEXP M_FIG C_FIG

SCOT+: A Comprehensive Software Suite for Single-Cell alignment Using Optimal Transport

Baker, C. D. — 2025-05-27

SummaryNew advances in single-cell multi-omics experiments have allowed biologists to examine how various biological factors regulate processes in concert on the cellular level. However, measuring multiple cellular features for a single cell can be quite resource-intensive or impossible with the current technology. By using optimal transport (OT) to align cells and features across disparate datasets produced by separate assays, Single Cell alignment using Optimal Transport+ (SCOT+), our unsupervised single-cell alignment software suite, allows biologists to align their data without the need for any correspondence. SCOT+ has a generic optimal transport solution that can be reduced to multiple different OT optimization procedures, each of which provide state-of-the-art single-cell alignment performance. With our user-friendly website and tutorials, this new package will help improve biological analyses by allowing for more accurate downstream analyses on multi-omics single-cell measurements. Implementation and AvailabilityOur algorithm is implemented in Pytorch and available on PyPI and GitHub (https://github.com/scotplus/scotplus). Additionally, we have many tutorials available in a separate GitHub repository (https://github.com/scotplus/book_source) and on our website (https://scotplus.github.io/).

Brain network dynamics reflect psychiatric illness status and transdiagnostic symptom profiles across health and disease

Cocuzza, C. V. — 2025-05-28

The network organization of the human brain dynamically reconfigures in response to changing environmental demands, an adaptive process that may be disrupted in a symptom-relevant manner across psychiatric illnesses. Here, in a transdiagnostic sample of participants with (n=134) and without (n=85) psychiatric diagnoses, functional connectomes from intrinsic (resting-state) and task-evoked fMRI were decomposed to identify constraints on brain network dynamics across six cognitive states. Hierarchical clustering of 110 clinical, behavioral, and cognitive measures identified participant-specific symptom profiles, revealing four core dimensions of functioning: internalizing, externalizing, cognitive, and social/reward. Brain network dynamics were flattened across cognitive states in individuals with psychiatric illness and could be used to accurately separate dimensional symptom profiles more robustly than both case-control status and primary diagnostic grouping. A key role of inhibitory cognitive control and frontoparietal network interactions was uncovered through systematic model comparison. We provide novel evidence that brain network dynamics can accurately differentiate the extent that psychiatrically-relevant dimensions of functioning are exhibited across health and disease.

Trainable subnetworks reveal insights into structure knowledge organization in protein language models

Vinod, R. — 2025-06-01

Protein language models (PLMs) pretrained via a masked language modeling objective have proven effective across a range of structure-related tasks, including high-resolution structure prediction. However, it remains unclear to what extent these models factorize protein structural categories among their learned parameters. In this work, we introduce trainable subnetworks, which mask out the PLM weights responsible for language modeling performance on a structural category of proteins. We systematically trained 39 PLM subnetworks targeting both sequence- and residue-level features at varying degrees of resolution using annotations defined by the CATH taxonomy and secondary structure elements. Using these PLM subnetworks, we assessed how structural factorization in PLMs influences downstream structure prediction. Our results show that PLMs are highly sensitive to sequence-level features and can predominantly disentangle extremely coarse or fine-grained information. Furthermore, we observe that structure prediction is highly responsive to factorized PLM representations and that small changes in language modeling performance can significantly impair PLM-based structure prediction capabilities. Our work presents a framework for studying feature entanglement within pretrained PLMs and can be leveraged to improve the alignment of learned PLM representations with known biological concepts.

Encoding of speech modes and loudness in ventral precentral gyrus

Srinivasan, A. — 2025-06-01

The ability to vary the mode and loudness of speech is an important part of the expressive range of human vocal communication. However, the encoding of these behaviors in the ventral precentral gyrus (vPCG) has not been studied at the resolution of neuronal firing rates. We investigated this in two participants who had intracortical microelectrode arrays implanted in their vPCG as part of a speech neuroprosthesis clinical trial. Neuronal firing rates modulated strongly in vPCG as a function of attempted mimed, whispered, normal or loud speech. At the neural ensemble level, mode/loudness and phonemic content were encoded in distinct neural subspaces. Attempted mode/loudness could be decoded from vPCG with an accuracy of 94% and 89% for two participants respectively, and corresponding neural preparatory activity could be detected hundreds of milliseconds before speech onset. We then developed a closed-loop loudness decoder that achieved 94% online accuracy in modulating a brain-to-text speech neuroprosthesis output based on attempted loudness. These findings demonstrate the feasibility of decoding mode and loudness from vPCG, paving the way for speech neuroprostheses capable of synthesizing more expressive speech.

Evaluating First-Pass, High Protein Capacity Desalting Techniques For Phosphoproteomics Applications

Callahan, A. — 2025-06-03

Many commercial desalting products exist for pre-MS peptide cleanup, although few exist that can handle the high protein input ([≥] 4 mg) required for phosphotyrosine enrichment. For these desalting products, the technical aptitude required for effective and organized desalting is often a barrier to entry for new users. Here, we evaluate four commercially available desalting techniques with varying degrees of automation, operational organization, and chemistries to determine the most cost-effective, user-friendly, and sensitive technique for protein profiling and phosphotyrosine (pY) enrichment. We find that TECAN Narrow Bore Extraction (NBE) products are the most cost effective per sample and least difficult to use, whereas ProtiFis S-Trap are the most expensive per sample and Pierce C18 spin columns have the worst operational organization. ProtiFi S-Trap vastly outperforms other desalting methods for peptide sequencing and protein profiling applications, uniquely identifying 25,654 unique peptide sequences and 375 unique proteins. Consistently, ProtiFi S-Trap samples show the deepest pY sequencing after Src SH2 superbinder enrichment, leading to the highest identification of significantly changing, biologically relevant pY sites in a Jurkat T cell signalling model. Our data show that ProtiFi S-Trap columns provide high peptide recovery, thus increasing meaningful pY site identification. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/657744v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@13b77e2org.highwire.dtl.DTLVardef@13bb4cforg.highwire.dtl.DTLVardef@dba171org.highwire.dtl.DTLVardef@1440f7d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Heightened subcortical reactivity to uncertain-threat is associated with future internalizing symptoms, conditional on stress exposure

Grogans, S. E. — 2025-06-12

BackgroundAnxiety, depression, and related internalizing illnesses are a leading burden on global public health, and often emerge during times of stress. Yet the underlying neurobiology has remained enigmatic, hindering treatment development. MethodsHere we used a combination of tools--including a well-established threat-anticipation fMRI paradigm and longitudinal assessments of internalizing symptoms and negative life events (NLEs)--to identify the neural systems associated with future internalizing illness in a risk-enriched sample of 224 emerging adults followed for 2.5 years. We performed parallel analyses in an overlapping sample of 209 participants who completed a popular threat-related faces paradigm. ResultsHere we show that heightened reactivity to uncertain-threat anticipation in the bed nucleus of the stria terminalis and the periaqueductal gray is associated with a worsening longitudinal course of broadband internalizing symptoms among individuals with low levels of NLE exposure. These associations were specific to uncertain threat and generally remained significant when controlling for concurrent measures of threat-elicited distress or psychophysiological arousal, highlighting the added value of the neuroimaging measures. Symptom trajectories were unrelated to amygdala and frontocortical reactivity to anticipated threat. Contrary to past research, amygdala reactivity to threat-related faces was unrelated to future symptoms. ConclusionsThese observations provide a novel neurobiological framework for conceptualizing transdiagnostic internalizing risk and lay the groundwork for mechanistic and therapeutics research. A racially diverse, risk-enriched sample and pre-registered, best-practices approach enhance confidence in the robustness and translational relevance of these results.

A Thiopurine-like Mutagenic Process Defines TGCT Subtypes

Brown, K. M. — 2025-06-12

Testicular germ cell tumors (TGCTs) are the most common malignancy in young men, exhibit a unique developmental origin and exceptional chemosensitivity. However, the molecular distinctions between TGCT subtypes remain poorly understood. Here we present a comprehensive genomic analysis of 252 treatment-naive primary TGCTs, integrating deep whole-genome sequencing with matched transcriptomic and epigenomic data. We identify new driver genes and uncover defining features of TGCTs, including pervasive chromosome X amplification with subtype-specific X chromosome inactivation, and a germ cell-like transcriptional program. Although previously reported, whole genome doubling (WGD) in TGCTs is further characterized here as ubiquitous, developmentally early, and associated with age at onset. Seminomas are enriched for early driver mutations, secondary WGD events, sustained XIST expression and replication stress-associated indel mutational signatures, while non-seminomas show greater structural complexity, subclonal diversity, relatively earlier-onset WGD, extended tumor latency, and telomere elongation. Moreover, we identify a mutational signature, SBS87, that is exceptionally rare across cancers with exception of thiopurine-treated leukemia, but strikingly prevalent in TGCT, especially non-seminomas. SBS87 is linked to extended tumor latency and telomere elongation, implicating possible environmental or endogenous processes that mimic thiopurine-induced DNA damage in TGCT pathogenesis. Collectively, our findings define TGCTs as molecularly distinct tumors shaped by early genomic instability and highlight SBS87 as a novel mutational footprint with potential etiologic and clinical relevance.

Senescence-Associated Chromatin Rewiring Promotes Inflammation and Transposable Element Activation

Dalgarno, A. — 2025-06-17

Cellular senescence is a stable form of cell cycle arrest that contributes to aging and age-associated diseases through the secretion of inflammatory factors collectively known as the senescence-associated secretory phenotype (SASP). While senescence is driven by transcriptional and epigenetic changes, the contribution of higher-order genome organization remains poorly defined. Here, we present the highest-resolution Hi-C maps ([~]3 kb) to date of proliferating, quiescent, and replicative senescent (RS) human fibroblasts, enabling a comprehensive analysis of 3D genome architecture during senescence. Our analyses reveal widespread senescence-associated remodeling of chromatin architecture, including extensive compartment and subcompartment switching toward transcriptionally active states, and a dramatic increase in unique chromatin loops. These structural features correlate with local DNA hypomethylation and are largely independent of canonical CTCF binding. The altered 3D genome landscape supports expression of SASP genes, inflammation-related pathways, and neuronal gene signatures consistent with age-associated epigenetic drift. We further demonstrate that architectural changes at multiple levels, including compartments, subcompartments, and loops, facilitate the derepression of LINE-1 retrotransposons, linking 3D chromatin structure to activation of proinflammatory transposable elements. Interestingly, quiescent cells, commonly used as senescence controls, exhibited substantial overlap in inflammatory gene expression with senescent cells, raising important considerations for experimental design. Structural analysis of cell cycle genes showed distinct chromatin configurations in senescence versus quiescence, despite similar transcriptional repression. Together, our results establish a high-resolution framework for understanding how genome architecture contributes to the senescent state.

Lubricin's Mucin Domain Has Strong Polyproline Type-II Helical Character

Feng, B. N. — 2025-06-18

Lubricin is a glycoprotein that is crucial for maintaining joint health by preventing joint wear by reducing joint friction in the boundary mode. Lubricin was recently observed to hinder the formation of uric acid crystals in the joint and prevent a form of gouty arthritis. However, despite lubricins great physiological importance, our current understanding of the molecular origins of lubricins beneficial properties is limited by a lack of detailed structural information regarding its central mucin domain: lubricins large size (227.5 kDa) and numerous glycosylations pose a substantial obstacle to conventional experimental methods for solving protein structures. In this work, we employ a combination of physics-based replica exchange molecular dynamics (REMD) simulations and circular dichroism (CD) experiments to shed light on the structure of lubricins central mucin domain. Using REMD, we model [KEPAPTTP]2, an amino acid repeat found throughout the mucin domain, and find that the mucin domain is likely to exhibit polyproline type II (PPII) helices, which are further stabilized by O-linked oligosaccharide chains. Motivated by these simulation results, we performed circular dichroism spectroscopy on fragments of the mucin domain that also show clear polyproline-II helical character, corroborating our computational findings. Altogether, this work provides strong evidence of a lubricin mucin domain with significant polyproline type II content. As polyproline helices are often also found in other glycoproteins with antifreeze properties, this work may also explain the atomistic underpinnings of their interfacial functions, including lubrication and competition with crystal formation. SIGNIFICANCELubricin is a mucinous glycoprotein containing a central heavily glycosylated domain that plays a crucial role in the lubrication of joints. However, little is known about the structure of this large central mucin domain, which makes the development of related therapeutics or biomedical devices challenging. In this work, we provide strong evidence that lubricins mucin domain possesses polyproline type II (PPII) character that is enhanced by glycosylation upon the basis of a combination of molecular dynamics simulations and circular dichroism experiments. Lubricins PPII character provides a molecular basis for its lubricating properties, which may provide insights into related antifreeze glycoproteins (AFGP) and the development of new biocompatible lubricants and ryo-preservatives.

A Chemostat-Based Model for Growing Bacterial Biofilms

Spagnolo, F. — 2025-06-24

Biofilms are groups of microbes that live together in dense communities, often attached to a surface. They play an outsized role in all aspects of microbial life, from chronic infections to biofouling to dental decay. In recent decades, appreciation for the diversity of roles that biofilms play in the environment has grown. Yet, most bacterial studies still rely upon approaches developed in the 19th century and center on planktonic populations alone. Here we present a chemostat-based experimental platform to investigate not only biofilms themselves, but how they interact with their surrounding environments. Our results show that biofilms grow to larger sizes in chemostats as opposed to flasks. In addition, we show that biofilms may be a consistent source of migrants into planktonic populations. We also show that secondary biofilms rapidly develop, although these may be more susceptible to environmental conditions. Taken together, our data suggest that chemostats may be a flexible and insightful platform for the study of biofilms in vitro. IMPORTANCEBiofilms are the predominant way that bacteria live in natural environments and are characterized by three emergent properties: ubiquity, resilience, and impact. They can be found across all environments, both natural and human-made, and across all of recorded time, dating back at least 3.5 billion years. Biofilms also represent a major economic impact of over $5 trillion annually. Yet, most of what is known about bacteria is the result of studies using planktonically growing liquid cultures under laboratory conditions. Here, we propose a comprehensive experimental platform that allows for study of biofilms on the molecular, organismal, and community levels using chemostats.

An atlas of cell type specific regulatory effects in cattle

Li, H. — 2025-06-24

Understanding the genetic and molecular architecture of complex traits and artificial selection is crucial for advancing sustainable precision breeding in cattle and other livestock. Yet, how genetic variation affects cellular gene expression remains elusive in cattle. Here, by integrating 8,866 bulk RNA-seq samples and 999,192 single cells of 81 cell types in 22 bovine tissues, we presented a comprehensive atlas of regulatory variants at the cell type resolution in cattle. By colocalizing with bulk-tissue expression quantitative trait loci (beQTL), we detected 57,043 novel cell-type stratified eQTL and cell-type/state interaction eQTL in 18,153 genes, which also exhibited a stronger tissue/cell-type specificity than beQTL. By examining genome-wide associations (GWAS) of 44 complex traits, these cell-resolved eQTL were colocalized with 505 (24%) additional GWAS loci compared to beQTL. Through integrating this resource with selection signatures between dairy and beef cattle, we provided tissue/cell-specific regulatory insights into cattle breeding. Overall, the current atlas of cell-type-specific regulatory variants will serve as an invaluable resource for cattle genomics and selective breeding.

EEG-GAN: A Generative EEG Augmentation Toolkit for Enhancing Neural Classification

Williams, C. C. — 2025-06-25

Electroencephalography (EEG) is a widely applied method for decoding neural activity, offering insights into cognitive function and driving advancements in neurotechnology. However, decoding EEG data remains challenging, as classification algorithms typically require large datasets that are expensive and time-consuming to collect. Recent advances in generative artificial intelligence have enabled the creation of realistic synthetic EEG data, yet no method has consistently demonstrated that such synthetic data can lead to improvements in EEG decodability across diverse datasets. Here, we introduce EEG-GAN, an open-source generative adversarial network (GAN) designed to augment EEG data. In the most comprehensive evaluation study to date, we assessed its capacity to generate realistic EEG samples and enhance classification performance across four datasets, five classifiers, and seven sample sizes, while benchmarking it against six established augmentation techniques. We found that EEG-GAN, when trained to generate raw single-trial EEG signals, produced signals that reproduce grand-averaged waveforms and time-frequency patterns of the original data. Furthermore, training classifiers on additional synthetic data improved their ability to decode held-out empirical data. EEG-GAN achieved up to a 16% improvement in decoding accuracy, with enhancements consistent across datasets but varying among classifiers. Data augmentations were particularly effective for smaller sample sizes (30 and below), significantly improving 70% of these classification analyses and only significantly impairing 4% of analyses. Moreover, EEG-GAN significantly outperformed all benchmark techniques in 69% of the comparisons across datasets, classifiers, and sample sizes and was only significantly outperformed in 3% of comparisons. These findings establish EEG-GAN as a robust toolkit for generating realistic EEG data, which can effectively reduce the costs associated with real-world EEG data collection for neural decoding tasks.

Inclusion of the ζ-chain drives phosphotyrosine signalling in CD19-CAR T cells

Callahan, A. — 2025-07-07

Although chimeric antigen receptor (CAR) T cell therapy has revolutionised individualised cancer therapies for relapsed/refractory lymphomas, signalling mechanisms underlying CAR T activation remain incompletely understood, especially among the three generations of CAR T exploiting different signalling domains. Here, using Jurkat T cell as a model, we investigate how costimulation influences tyrosine phosphorylation cascades using LC-MS/MS based phosphotyrosine (pY) proteomics and CD69 expression in the presence of small molecule inhibitors of key TCR signalling regulators. We find that including the {zeta}-chain in first ({zeta}-CAR), second (28{zeta}-CAR and BB{zeta}-CAR), and third (28BB{zeta}-CAR) generation CARs largely determines pY signalling, irrespective of costimulation. Further, we show that the phosphatase activity of PTPN22 and SHP-1 are largely negligible for activation of CARs, but indiscriminate inhibition of phosphatases using Pervanadate (PV) selectively activates BB{zeta}-CARs without antigen encounter. Finally, we find that selective, partial inhibition of Itk using Soquelitinib reduces basal CD69 expression in Jurkat CAR T cells while maintaining their ability to activate in response to antigen. Our data suggest that the {zeta}-chain determines the pY signalling profile of CD19-CAR Jurkat T cells and that Itk may drive antigen-independent CD19-CAR activation.

Musculoskeletal architecture of the shoulder: A comparative anatomy study in bats and mice informing human rotator cuff function

Kurtaliaj, I. — 2025-07-08

Overhead motion in humans often leads to shoulder injuries, a consequence of the evolutionary trade-off in glenohumeral joint anatomy that balances stability with mobility. Bats consistently engage in overhead motion during flight, subjecting their shoulders to substantial loading throughout their relatively long lifespan. Remarkably, despite the demands placed on a bats shoulder, instability and rotator cuff tears, which could be fatal to bats in short order, are not observed in nature. We were thus inspired to study functional adaptations in the shoulders of bats that enable this overhead motion. Comparative anatomical studies of the shoulders of bats and mice, similarly-sized quadrupeds, were performed and interpreted using a mathematical model. Scapular anatomy indicated a more prominent role for the infraspinatus muscle in the bat compared to the mouse. Measurements of bat and mice shoulders revealed that the bat glenoid had a larger curvature and arc length than that of mice, providing a larger articulating surface area with and deeper enclosing surface of the humeral head. Modeling results predicted that the bat shoulder is stable over a dramatically larger range of angles compared to the mouse shoulder. These results suggested that adaptations to constraints imposed by the bony anatomy and rotator cuff tendons of the shoulder may contribute to the ability of bats to sustain overhead motion in a high stress, repeated loading environment without injury. Results suggest that bats have evolved unique adaptations in their glenohumeral bony anatomy that reduce stress on the supraspinatus, enhance joint stability, and optimize strength across a broad range of motion.

Programmable microparticles rewire CAR signaling to enable super-physiological expansion of human T cells in vitro

Zeng, Q. — 2025-07-22

T cell proliferative capacity and persistence critically determine the therapeutic success of chimeric antigen receptor (CAR) T cells. However, it remains unknown if and how human CAR-T cells can be externally programmed to reach maximal proliferative capacity. Here, we use programmable PLGA microparticles functionalized with CAR-antigens and CD28-costimulatory antibodies (CAREp) to repeatedly stimulate human CD8+ CAR-T cells in vitro. CAREp-stimulated CAR-T cells expanded continuously for over 100 days--versus [~]30 days with tumor cell stimulation--and achieved up to 1018-fold cumulative expansion, greatly surpassing CD3/28-Dynabeads. Early-phase transcriptomic responses-- upregulation of DNA repair, cell cycle, telomere maintenance, and mitochondrial pathways--aligned with long-term outcomes: massive proliferation, telomere stability, robust respiration, and preserved progenitor phenotype by single-cell sequencing. Differentiation and exhaustion signals were broadly suppressed. Transient telomerase activity further supported physiologic expansion. These findings demonstrate that nanoscale-controlled extracellular cues can rewire intracellular signaling to drive durable, super-physiological expansion of functional CAR-T cells.

TGF-β Coordinates Alanine Synthesis and Import for Myofibroblast Differentiation in Pulmonary Fibrosis

Li, F. — 2025-07-24

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease marked by aberrant fibroblast-to-myofibroblast differentiation, a process that requires metabolic reprogramming. We identify alanine as a critical metabolite that confers metabolic flexibility to support differentiation. TGF-{beta} increases alanine by activating both its synthesis and import in normal and IPF lung fibroblasts. Alanine is synthesized primarily by GPT2, which is regulated by a glutamine-glutamate--ketoglutarate axis. Inhibiting GPT2 depletes alanine and suppresses TGF-{beta}-induced expression of -SMA and COL1A1, an effect reversed by alanine supplementation. We also identify SLC38A2 as a key transporter of both alanine and glutamine that is upregulated by TGF-{beta} and alanine deprivation. Together, SLC38A2 and GPT2 activities converge to maintain intracellular alanine levels to support myofibroblast differentiation. Mechanistically, alanine deficiency suppresses glycolysis and depletes tricarboxylic acid cycle intermediates, while supplementation provides carbon and nitrogen for intracellular glutamate and proline biosynthesis, particularly in the absence of glutamine. Combined inhibition of GPT2 and SLC38A2 suppresses fibrogenic responses in fibroblasts and in human precision-cut lung slices, highlighting a potential therapeutic strategy for fibrotic lung disease.

A tumorigenesis threshold for endogenous Myc revealed by dosage-compensation for Myc haploinsufficiency in the absence of p53.

Bao, X. — 2025-07-31

The MYC proto-oncogene is crucial for neoplasia in most tumors. Overexpressed, oncogenic MYC amplifies the flux through most major processes but does not specify a unique carcinogenic pathway. This "amplifier" model suggests that MYC must exceed an expression threshold to become oncogenic. We designed a genetic test of this model, using the mouse Trp53 null mutant (p53KO) as a highly robust tumor generator to examine the effect of a modest change in the endogenous Myc level (Myc+/-). Strikingly, tumor-free survival is greatly extended in p53KO mice with haploid Myc gene-dosage, yet in the tumors that do develop (mainly hemangiosarcomas and thymic lymphomas), their Myc deficit has been invariably compensated either by increasing Myc genomic dosage (hemangiosarcomas) or expression (lymphomas). Furthermore, acutely halving the endogenous Myc gene-dosage in established tumor allografts curtails growth rates. These results indicate that even an incremental reduction of MYC activity can be salutary in cancer and that one of the major tumor suppressor functions of p53 derives from its ability to prevent MYC overexpression. Myc generates acute DNA damage by several mechanisms and accordingly, p53s anti-Myc function may be inextricably linked to its role in genome integrity surveillance.

Uracil-DNA glycosylase deficiency is associated with repressed tumor cell-intrinsic inflammatory signaling and altered sensitivity to exogenous interferons

Vendetti, F. P. — 2025-08-01

2-deoxyuridine (dU) is a common DNA lesion resulting from cytosine deamination and from dUMP incorporation by DNA polymerases, both of which are prevalent in cancer. The primary mechanism that repairs dU lesions in genomic DNA is base excision repair initiated by Uracil-DNA Glycosylase 1 (UNG1). We generated Ung knockout mouse B16 melanoma cells to investigate the consequences of UNG deficiency in a well-characterized, immunoproficient, syngeneic mouse cancer model. We show that UNG-deficient ({Delta}UNG) B16 tumors have altered growth kinetics in vivo and that their delayed growth is T-cell dependent. Immune profiling revealed reduced CD8+ T cell infiltration but augmented CD4+ Th1 responses in {Delta}UNG tumors. In vitro, {Delta}UNG tumor cells exhibit strongly suppressed cell-intrinsic type-I interferon, type-II interferon, and inflammatory signaling gene expression signatures as well as altered cytokine and chemokine secretion. In vivo, {Delta}UNG tumors exhibit a modified inflammatory cytokine and chemokine milieu. Furthermore, {Delta}UNG tumor cells have altered sensitivity to exogenous interferons in vitro, with increased sensitivity to IFN-{gamma} but decreased sensitivity to IFN-/{beta}. Collectively, our data show that tumor cell-specific UNG deficiency results in an altered tumor microenvironment in vivo and provide proof-of-concept data for the use of UNG inhibitors to modulate inflammatory pathways in tumors.

Spatiotemporal Dynamics of fMRI Signal Changes Induced by High Concentration Normobaric Oxygen Inhalation

Su, Y.-S. — 2025-08-01

While it is well known that oxygen supports the brains metabolic demands, it remains unclear how increased oxygen concentration influences intrinsic neural activity over time and across brain regions. Using resting-state functional magnetic resonance imaging (fMRI), we examined the dynamic responses to high-concentration normobaric oxygen across distinct phases of exposure and withdrawal. We revealed three patterns in the BOLD signals: increased activation during inhalation, an undershoot following immediate oxygen withdrawal, and reactivation even without continued oxygen. These responses were most pronounced in the default mode network (DMN), but also exhibited spatiotemporally heterogeneous patterns across the brain, a map we term Brain Oxygen Sensitivity Topography (BOST). Functional connectivity analyses further revealed increased between-network connectivity during inhalation and enhanced within-network connectivity in the DMN during the aftereffect. This spatiotemporal heterogeneity and transient network reorganization suggests that distinct physiological processes are engaged at each phase, enabling us to predict how different oxygen protocols will enhance specific cognitive functions.

MR-AIV reveals in-vivo brain-wide fluid flow with physics-informed AI

Toscano, J. D. — 2025-08-01

The circulation of cerebrospinal and interstitial fluid plays a vital role in clearing metabolic waste from the brain, and its disruption has been linked to neurological disorders. However, directly measuring brain-wide fluid transport--especially in the deep brain--has remained elusive. Here, we introduce magnetic resonance artificial intelligence velocimetry (MR-AIV), a framework featuring a specialized physics-informed architecture and optimization method that reconstructs three-dimensional fluid velocity fields from dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). MR-AIV unveils brain-wide velocity maps while providing estimates of tissue permeability and pressure fields--quantities inaccessible to other methods. Applied to the brain, MR-AIV reveals a functional landscape of interstitial and perivascular flow, quantitatively distinguishing slow diffusion-driven transport ([~] 0.1 {micro}m/s) from rapid advective flow ([~] 3 {micro}m/s). This approach enables new investigations into brain clearance mechanisms and fluid dynamics in health and disease, with broad potential applications to other porous media systems, from geophysics to tissue mechanics.

Enabling CAR-T Cell Immunotherapy in Glioblastoma by Modifying Tumor Microenvironment via Oncolytic Adenovirus Encoding Bispecific T Cell Engager

Choi, M. J. — 2025-08-02

Recent clinical trials show that CAR-T cell therapies can initially blunt tumor growth in glioblastoma (GBM) patients. However, the tumor microenvironment activates mechanisms that inhibit tumor-killing potential of the CAR-T cells and limit their therapeutic efficacy. To counteract this, we have utilized oncolytic adenovirus (OV) Ad5-{Delta}24-RGD as a platform to overexpress a bispecific T cell engager (BiTE) targeting both T cell marker CD3 and GBM specific tumor associated antigen IL-13R2. We first demonstrated that OV-BiTE could enhance recruitment of T cells to GBM in vitro and in vivo. We then showed that intratumoral injection of OV-BiTE followed by infusion of combined EGFR- and EGFRvIII-CAR-T cells was more effective than OV-BiTE supplemented with either CAR-T therapy alone, and led to significant tumor eradication in a GBM xenograft mouse model. In conclusion, our multimodal OV-BiTE & CAR-T cell immunotherapy is capable of overcoming immunosuppressive tumor microenvironment and GBM resistance to treatment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/667708v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@c55b15org.highwire.dtl.DTLVardef@deffb2org.highwire.dtl.DTLVardef@64ff90org.highwire.dtl.DTLVardef@c65fc2_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIOncolytic adenovirus encoding bispecific T cell engager (OV-BiTE) combines two immunotherapeutic agents into one. C_LIO_LIOV-BiTE strategy modifies tumor microenvironment and enhances recruitment of T cells to glioblastoma (GBM) in vitro and in vivo. C_LIO_LIMultimodal OV-BiTE & CAR-T cell immunotherapy effectively reduced tumor mass in a GBM xenograft mouse model and is superior to either immunotherapy alone. C_LI

A novel locus associated with decreased susceptibility of Plasmodium falciparum to lumefantrine and dihydroartemisinin has emerged and spread in Uganda

Niare, K. — 2025-08-02

Malaria control in Uganda is threatened by the emergence of artemisinin partial resistance (ART-R) and decreasing lumefantrine susceptibility. To identify loci contributing to decreased drug susceptibility, we assessed signatures of selection in Ugandan whole genome Plasmodium falciparum sequences. Extended shared haplotypes were seen for the ART-R associated Kelch13 (K13) C469Y and A675V mutations, but the strongest signal of recent selection was centered on a segment of chromosome 7 encoding the phosphoinositide-binding protein gene (px1, PF3D7_0720700). A haplotype, represented by three PX1 SNPs (L1222P, M1701I and D1705N) and two indels (designated PIN), was first seen in 2008 and rapidly increased, reaching prevalence >50% in northern Uganda by 2016 and eastern Uganda by 2023. PIN-carrying parasites showed significantly decreased ex vivo susceptibilities, measured as IC50s, to lumefantrine, mefloquine and dihydroartemisinin, either with or without coincident K13 mutations. Thus, PX1 polymorphisms may impact on the susceptibilities of African malaria parasites to key drugs.

Programmed meiotic errors facilitate dichotomous sperm production in the silkworm, Bombyx mori

Benner, L. — 2025-08-03

The goal of meiosis is typically to produce haploid gametes (eggs or sperm). Failure to do so is catastrophic for fertility and offspring health. However, Lepidopteran (moths and butterflies) males produce two morphs of sperm: nucleated (eupyrene) sperm which fertilize the egg, and anucleated (apyrene) sperm, both of which are essential for fertilization. The meiotic differences in the two types of spermatogenesis have not been well characterized, and our knowledge of the molecular differences between eupyrene and apyrene spermatogenesis are extremely limited in all systems. The only factor identified as being required for apyrene spermatogenesis is Sex-lethal (Sxl). Here, we show through cytological analysis of early meiotic events that there are several key differences in the genesis of apyrene sperm and eupyrene sperm. Specifically, apyrene spermatocytes fail to condense and pair their chromosomes during meiotic prophase I. In addition, telomeres do not attach to the nuclear envelope. Due to these differences, full-length synaptonemal complex does not form. RNA sequencing of both eupyrene- and apyrene-producing testes revealed distinct changes in transcriptional programs, including down-regulation of a myriad of meiotic genes and cell cycle checkpoint factors during apyrene meiosis. By comparing wild-type and Sxl-knockout apyrene testes, we found that Sxl is not required for the changes in the expression of the meiotic genes but instead plays a role in checkpoint inactivation to allow this error-prone meiosis to proceed. Together, our findings reveal significant insights into two converging molecular pathways that promote the formation of dimorphic sperm in Lepidoptera.

Decoding intended speech with an intracortical brain-computer interface in a person with longstanding anarthria and locked-in syndrome

Jude, J. J. — 2025-08-13

Intracortical brain-computer interfaces (iBCIs) for decoding intended speech have provided individuals with ALS and severe dysarthria an intuitive method for high-throughput communication. These advances have been demonstrated in individuals who are still able to vocalize and move speech articulators. Here, we decoded intended speech from an individual with longstanding anarthria, locked-in syndrome, and ventilator dependence due to advanced symptoms of ALS. We found that phonemes, words, and higher-order language units could be decoded well above chance. While sentence decoding accuracy was below that of demonstrations in participants with dysarthria, we are able to attain an extensive characterization of the neural signals underlying speech in a person with locked-in syndrome and through our results identify several directions for future improvement. These include closed-loop speech imagery training and decoding linguistic (rather than phonemic) units from neural signals in middle precentral gyrus. Overall, these results demonstrate that speech decoding from motor cortex may be feasible in people with anarthria and ventilator dependence. For individuals with longstanding anarthria, a purely phoneme-based decoding approach may lack the accuracy necessary to support independent use as a primary means of communication; however, additional linguistic information embedded within neural signals may provide a route to augment the performance of speech decoders.

Genomic resources for comparative analyses of obligate avian brood parasitism

Carroll, R. A. — 2025-08-15

Examples of convergent evolution, wherein distantly related organisms evolve similar traits, including behaviors, underscore the adaptive power of natural selection. In birds, obligate brood parasitism, and the associated loss of parental care behaviors, has evolved independently in seven different lineages, though little is known about the genetic basis of the complex suite of traits associated with this rare life history strategy. We generated genome assemblies for ten brood parasitic species plus eight species representatives of their parental/nesting outgroups. This includes nine long-read chromosome-level assemblies, with scaffold N50 sizes ranging from 38.1 to 72.6 MB, and gene representation completeness measures >97%. Leveraging this new catalog of avian genomes, we constructed clade-level alignments that reveal variation in chromosomal synteny, provide first-time or improved annotations of protein-coding and non-coding genes, and define cross-species ortholog reference sets. We also refine estimates for the timing of the seven independent origins of brood parasitism, ranging from recent events such as 1.6 to 4.5 million years ago in Molothrus cowbirds to much earlier origins over 30 million years ago in two of the three cuckoo lineages. These genomic resources lay the foundation for investigating the genetic and genomic underpinnings of brood parasitism, including the loss of parental care, shifts in mating systems, perhaps resulting in heightened sperm competition, elevated annual fecundity, improved spatial cognition related to nest-finding, and the diverse adaptations shaped by intense coevolution with host species.

VO: The Vaccine Ontology

Zheng, J. — 2025-08-15

With the widespread use of vaccines in research and clinical settings, there is an urgent need to standardize vaccine representation, integrate information across diverse vaccine types, and support computer-assisted reasoning. Accordingly, we have since 2007 developed the community-based Vaccine Ontology (VO), which aligns with the Basic Formal Ontology and adheres to OBO Foundry principles. VO models ontologically vaccines, vaccine components, vaccine immune responses, vaccine investigation studies and other vaccine-related topics. VO represents more than 10,000 vaccines targeting 289 infectious pathogens and cancers in humans and over 30 nonhuman animal species. VO provides mappings to external resources such as RxNorm, CVX, FDA, and USDA. Various VO use cases exist. VO facilitates vaccine standardization in resources such as the VIOLIN vaccine database, ImmPort, and the Vaccine Adjuvant Compendium (VAC). Semantic queries can be made to query VO. VO has been shown to enhance experimental and clinical vaccine data analysis and vaccine literature mining. Overall, VO standardizes vaccine modeling and representation and greatly supports vaccine AI research in the Semantic Web era.

A Cryptic Binding Pocket Regulates the Metal-Dependent Activity of Cas9

Ahsan, M. — 2025-08-26

Cas9 is a metal-dependent nuclease that has revolutionized gene editing across diverse cells and organisms exhibiting varying ion uptake, metabolism, and concentrations. However, how divalent metals impact its catalytic function, and consequently its editing efficiency in different cells, remains unclear. Here, extensive molecular simulations, Markov State Models, biochemical and NMR experiments, demonstrate that divalent metals - Mg2+, Ca2+, and Co2+ - promote activation of the catalytic HNH domain by binding within a dynamically forming divalent metal binding pocket (DBP) at the HNH-RuvC interface. Mutations in DBP residues disrupt HNH activation and impair the coupled catalytic activity of both nucleases, identifying this cryptic DBP as a key regulator of Cas9s metal-dependent activity. The ionic strength thereby promotes Cas9s conformational activation, while its catalytic activity is metal-specific. These findings are critical to improving the metal-dependent function of Cas9 and its use for genome editing in different cells and organisms.

Choice history biases evidence accumulation: a cross-species comparison from humans to mice

Fernandez, K. — 2025-08-27

Mice are increasingly used to study the neural circuit-level basis of behavior, often with the ultimate goal of extrapolating these insights to humans. To generalize insights about neural functioning across species, it is crucial to ensure correspondence in behavioral and cognitive strategy. We previously showed that human observers evidence accumulation is biased by previous choices (Urai et al., 2019). To replicate these findings across species, we fit Diffusion Decision Models (Fengler et al., 2025) to behavioral data from 62 mice performing a standardized perceptual decision-making task (The International Brain Laboratory et al., 2021). We identified the same cognitive strategy of history-dependent evidence accumulation: individual differences in choice repetition were explained by a history-dependent bias in the rate of evidence accumulation rather than its starting point. We argue that history-biased evidence integration reflects a fundamental aspect of perceptual decision-making, that may transcend the specific species.

The ecology and adaptive function of clownfish color patterns

Froehlich, C. Y. M. — 2025-09-01

Anti-predator coloration strategies vary widely from conspicuous to cryptic, and everything between. Coloration serves multiple functions, and determining the selective pressures that drive this diversity is a long-standing mystery. Clownfishes display enormous inter- and intra-specific variation in coloration without the influence of sexual selection, providing an excellent system in which to explore these drivers in closely related and sympatric species. Across clownfishes, three color-pattern phenotypes have evolved convergently with host anemone use, implying adaptive functions linked to the hosts physical and chemical properties. Here we conducted a comprehensive study to infer the ultimate functions of clownfish color patterns. By integrating multiple levels of biological organization, including behavior, visual ecology, dietary niche, and microbiome, our data overwhelmingly indicate that specialist and generalist clownfish color patterns have different underlying ecologies and adaptive functions. Specialist clownfishes background match their anemones, remain near hosts, and have specialized diets and microbiomes. Generalists are highly contrasting with their anemones, remain far from hosts, and have variable diets and microbiomes. Taken together, specialist color patterns align with functional expectations of camouflage while generalists align with aposematic or disruptive coloration. Our study provides novel insights into how ecological conditions shape the evolution of multiple anti-predator coloration strategies.

Neuroinflammatory crosstalk between microglia and astrocytes increases viral replication in an iPSC-derived model of CNS HIV infection

Gesualdi, J. — 2025-09-03

People living with HIV suffer multiple comorbid conditions related to chronic inflammation at increased rates compared to the general population, even when on effective antiretroviral therapy. In particular, current data indicate that the increased incidence and severity of neurocognitive impairment (NCI) are associated with unresolved neuroinflammation. Attempts to treat NCI in people living with HIV by reducing inflammation have thus far been unsuccessful, suggesting that a more mechanistic understanding of inflammatory processes in the CNS during HIV is necessary. Here, we use iPSC-derived microglia (iMg) and astrocytes (iAst) to model HIV infection in the CNS. We show that our iMg robustly express markers associated with microglial identity and are susceptible to HIV infection, but exhibit lower HIV replication rates and weaker immune response to HIV challenge compared to monocyte-derived macrophages. Coculture of iAst with iMg leads to a much stronger pro-inflammatory immune response, and, surprisingly, a robust increase in rates of HIV replication. Increased replication in iMg/iAst cocultures is associated with higher levels of multiple pro-inflammatory cytokines, including TNF, which is produced by iAst upon exposure to HIV-infected iMg. Addition of exogenous TNF to iMg during HIV infection is also sufficient to increase rates of replication, and neutralization of TNF via adalimumab/Humira treatment in iMg/iAst cocultures reduces replication. Blocking NF-kB signaling with iKK inhibitor Bay-11-7082 (Bay-11) demonstrates that increased HIV replication in iMg/iAst cocultures is due to increased NF-kB activity. Finally, we show that in HIV-infected iMg there is movement of lysosomes to the periphery of the cell membrane and release of lysosomal content into the extracellular space, suggesting that this dysregulated lysosomal flux could further contribute to the pro-inflammatory microenvironment. We propose that this altered lysosomal trafficking and increased cytokine production drives a pro-inflammatory phenotype in glia and represents a potential source of unresolved neuroinflammation in people living with HIV.

Fear, anxiety, and the extended amygdala- Absence of evidence for strict functional segregation

Didier, P. — 2025-09-04

Since the time of Freud, the distinction between fear and anxiety has been a hallmark of influential models of emotion and emotional illness, including the Diagnostic and Statistical Manual of Mental Disorders (DSM) and Research Domain Criteria (RDoC). Fear and anxiety disorders are a leading cause of human misery and morbidity. Existing treatments are inconsistently effective, underscoring the importance of developing accurate models of the underlying neurobiology. Although there is consensus that the extended amygdala (EA) plays a central role in orchestrating responses to threat, the respective contributions of its two major subdivisions--the central nucleus of the amygdala (Ce) and bed nucleus of the stria terminalis (BST)--remain contentious. To help adjudicate this debate, we performed a harmonized mega-analysis of fMRI data acquired from 295 adults as they completed a well-established threat-anticipation paradigm. Contrary to popular double-dissociation models, results demonstrated that the Ce responds to temporally uncertain threat and the BST responds to certain threat. In direct comparisons, the two regions showed statistically indistinguishable responses, with strong Bayesian evidence of regional equivalence. These observations underscore the need to reformulate conceptual models that posit a strict segregation of temporally certain and uncertain threat processing in the EA.

The Global Canopy Atlas: analysis-ready maps of 3D structure for the world's woody ecosystems

Fischer, F. J. — 2025-09-04

Woody canopies regulate exchanges of energy, water and carbon, and their three-dimensional (3D) structure supports much of terrestrial biodiversity. Remote sensing technologies such as airborne laser scanning (ALS) now enable the 3D mapping of entire landscapes. However, we lack the large, harmonized and geographically representative ALS collections needed to build a global picture of woody ecosystem structure. To address this challenge, we developed the Global Canopy Atlas (GCA): 3,458 ALS acquisitions transformed into standardized and analysis-ready maps of canopy height and elevation at 1 m2 resolution. The GCA covers 56,554 km2 across all major biomes. 19% of this area has been scanned multiple times, and 87% of all GCA products are openly available, covering 95% of the total area. To showcase its wide range of applications, we applied the GCA in three case studies. First, we validated three global satellite-derived canopy height maps, finding poor performance at native resolution (1-30 m, R2 < 0.38) and moderate performance at 250 m resolution (R2 < 0.65). Second, analyzing global patterns in canopy gap size frequency we discovered an unexpectedly large variation of power law exponents from branch to stand level ( = 1.52 to 2.38), pointing to a fundamental scale-dependence of forest structure. Third, we developed a framework to standardize forest turnover quantification from multi-source, multi-temporal ALS. In a temperate forest in North America it revealed that 21% of canopy gaps closed within 12 years of opening and would thus be missed by infrequent monitoring. As demonstrated by these case studies, the GCA provides a novel data source for ecologists, foresters, remote sensing scientists and the ecosystem modelling community that substantially advances our ability to understand the structure and dynamics of woody ecosystems at global scales.

Design Rules for Expanding PAM Compatibility in CRISPR-Cas9 from the VQR, VRER and EQR variants

Vieya, F. H. — 2025-09-04

Expanding the range of Protospacer Adjacent Motifs (PAMs) recognized by CRISPR-Cas9 is essential for broadening genome-editing applications. Here, we combine molecular dynamics simulations with graph-theory and centrality analyses to dissect the principles of PAM recognition in three Cas9 variants - VQR, VRER, and EQR - that target non-canonical PAMs. We show that efficient recognition is not dictated solely by direct contacts between PAM-interacting residues and DNA, but also by a distal network that stabilizes the PAM-binding domain and preserves long-range communication with REC3, a hub that relays signals to the HNH nuclease. A key role emerges for the D1135V/E substitution, which enables stable DNA binding by K1107 and preserves key DNA phosphate locking interactions via S1109, securing stable PAM engagement. In contrast, variants carrying only R-to-Q substitutions at PAM-contacting residues, though predicted to enhance adenine recognition, destabilize the PAM-binding cleft, perturb REC3 dynamics, and disrupt allosteric coupling to HNH. Together, these findings establish that PAM recognition requires local stabilization, distal coupling, and entropic tuning, rather than a simple consequence of base-specific contacts. This framework provides guiding principles for engineering Cas9 variants with expanded PAM compatibility and improved editing efficiency. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/673821v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1011b93org.highwire.dtl.DTLVardef@1b96a0eorg.highwire.dtl.DTLVardef@1e1c42corg.highwire.dtl.DTLVardef@11628be_HPS_FORMAT_FIGEXP M_FIG C_FIG

Structure, dynamics, and processing of 8oxoG:A in the nucleosome

Vito, A. F. — 2025-09-05

Eukaryotic genomic DNA is packaged into chromatin through a repeating unit known as the nucleosome. In this chromatin environment, DNA is constantly exposed to several sources of DNA damage, such as reactive oxygen species (ROS), which can lead to the formation of 8-oxo-7,8-dihydroguanine (8oxoG). 8oxoG can base pair with cytosine (8oxoG:C) or form a mutagenic base pair with adenine (8oxoG:A), which can lead to single base transversions if left unrepaired. To date, the structure and dynamics of these two possible 8oxoG base pairs in the nucleosome remain unclear. Furthermore, whether MutY homologue (MUTYH) excises 8oxoG:A base pairs in the nucleosome remains elusive. Here using a combination of cryogenic-electron microscopy, molecular dynamics simulations, and biochemistry we determined the structure and dynamics of 8oxoG:C and 8oxoG:A base pairs in the nucleosome and characterize MUTYH activity in nucleosomal DNA. We found that nucleosomal 8oxoG:C forms a stable base pair using its anti conformation, while nucleosomal 8oxoG:A forms a more dynamic base pair using its syn conformation that is unable to be processed by MUTYH. This work provides fundamental insight into the accommodation of oxidative damage in the nucleosome and how this damage contributes to increased mutagenic transversions in nucleosomal compared to linker DNA.

Exploratory Statistical Analysis of Rheumatoid Factor Based Subgroups in Sjögren's Syndrome.

Zhuang, N. Z. — 2025-09-12

This method describes a computational pipeline for stratifying autoimmune patient groups using exclusively binary autoantibody data. Our method addresses a methodological gap in computational immunology by providing a standardized framework for analyzing categorical serological data commonly found in electronic health records and resource-limited settings. The pipeline integrates three complementary analytical modules: O_LIModule 1: Exploratory screening using statistical association tests. C_LIO_LIModule 2: Quantification of overall immunological similarity and un-certainty. C_LIO_LIModule 3: Prediction modeling and validation against chance. C_LI We demonstrate the methods utility by applying it to two autoimmune disorders. We were successful in recapitulating established clinical relationships in these two closely linked diseases. The pipeline is implemented in Python and includes detailed configuration options for custom disease groups, autoanti-body panels and stratification variables. This method enables researchers to extract meaningful immunological patterns from underutilized binary clinical data, serving as a hypothesis-generation tool to help drive impactful exploration. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/670596v4_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@7c6df3org.highwire.dtl.DTLVardef@116ac5eorg.highwire.dtl.DTLVardef@18e76dcorg.highwire.dtl.DTLVardef@1d6b71_HPS_FORMAT_FIGEXP M_FIG C_FIG

RhoV Promotes Proliferation, Migration, and MAPK Activation in Pancreatic Ductal Adenocarcinoma

Wu, S. — 2025-09-16

Background and AimsPancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy characterized by diagnosis at advanced stages, limited therapeutic options, and frequent resistance to therapies. Although oncogenic KRAS mutations are central drivers of PDAC, alternative pathways indisputably contribute to its tumorigenesis and progression. RhoV, a member of the Rho family of small GTPases, has been implicated in tumor development in other cancer types, such as breast cancer and lung adenocarcinoma; however, its role in PDAC remains unclear. MethodsIn this study, we investigated the expression and functional impact of RhoV on PDAC. Analysis of publicly available datasets and immunohistochemical profiling of 114 PDAC patient specimens were used to evaluate the expression of RhoV in PDAC and its prognostic impact. Overexpression and CRISPR-Cas9-mediated knockout of RhoV were established in three pancreatic cancer cell lines. Functional analyses, such as cell proliferation, migration, invasion, colony formation, spheroid growth, and mouse xenograft, were used to evaluate the role of RhoV in PDAC cells. ResultsRhoV overexpression was associated with reduced overall and recurrence-free survival in public datasets and our own patient cohort. Functional assays demonstrated that RhoV overexpression promoted PDAC cell proliferation, colony formation, and spheroid growth, whereas knockout of RhoV suppressed these changes. Moreover, RhoV enhanced PDAC cell migration and invasion in vitro, accompanied by downregulation of E-cadherin and upregulation of N-cadherin and vimentin, indicating induction of epithelial-mesenchymal transition. Mechanistically, RhoV overexpression activated key MAPK pathway components, including phosphorylation of ERK, JNK, and p38. In vivo, xenograft models confirmed that RhoV drives tumor growth and increases tumor burden. ConclusionThese results establish RhoV as a novel oncogenic factor in PDAC progression and highlight its potential as a biomarker and therapeutic target, warranting further investigation into combinatorial targeting strategies to overcome KRAS inhibitor resistance.

Expression of vimentin intermediate filaments in epithelial cells promotes cell migration and cell matrix interaction in 3D

Rodriguez, C. — 2025-09-19

During a variety of physiological and pathological processes, such as development, wound healing, and tumor progression, epithelial cells collectively invade into their surroundings. Vimentin intermediate filaments (VIFs) are often observed to play a role in the epithelial cells located at the margins of 2D cultures. However, their role in 3D collective cell behavior remains underexplored. Here, we investigate how induced vimentin expression affects 3D multicellular architecture and mechanics in luminal breast cancer cells (MCF-7) that ordinarily express keratin intermediate filaments only. We find that vimentin expression significantly alters 3D cell cluster morphology, inducing protrusions and increasing boundary fluctuations. Furthermore, cells in vimentin-expressing clusters show enhanced, more stochastic migration. In addition, these clusters exert stronger and localized traction forces on the surrounding matrix, indicating increased cell-matrix interactions. Transcriptomic analysis corroborates these biophysical findings, revealing upregulated gene expression for cell migration and matrix adhesion, and downregulated cell-cell adhesion genes. Our results demonstrate that VIFs are critical in modulating 3D multicellular collective morphology and dynamics, promoting invasive-like behavior by enhancing cell migration and cell-matrix interactions. These results provide fundamental insights into understanding tissue morphogenesis and disease progression.

YeastSAM: A Deep Learning Model for Accurate Segmentation of Budding Yeast Cells

Zhao, Y. — 2025-09-20

An essential step for quantitative image analysis is cell segmentation, which is the process of defining the outline of individual cells in microscopy images. Segmentation of budding yeast is challenging due to their asymmetric cell division and mother-bud morphology. As a result, a dividing cell is frequently misidentified as two separate cells, causing errors in downstream analysis. Here, we overcame this challenge by developing YeastSAM, a deep learning-based segmentation framework derived from {micro}SAM and optimized for budding yeast. YeastSAM achieves more than threefold higher accuracy in segmenting dividing cells compared to existing methods. When combined with single-molecule RNA imaging and organelle imaging, YeastSAM facilitates quantitative analysis of the spatial regulation of gene expression. This study offers an accessible, high-accuracy model for yeast cell segmentation, empowering researchers with minimal programming experience to perform quantitative image analysis.

Bispecific Targeting of CHI3L1 and PD-1/PD-L1 Axis as a Novel Therapeutic Strategy for Idiopathic Pulmonary Fibrosis

Jeong, H.-S. — 2025-09-20

CHI3L1, a chitinase-like protein, plays a key role in the pathogenesis of pulmonary fibrosis, though the precise mechanisms remain unclear. This study explores how CHI3L1 regulates profibrotic macrophage activation and invasive myofibroblast differentiation and their interactions. In vitro, CHI3L1 induced profibrotic M2 macrophage activation and differentiation marked by increased expression of CD163, CD206, and PD-L1. CHI3L1 also enhanced TGF-{beta}1 effects on lung fibroblasts including myofibroblast transformation, migration and tissue invasion. Mechanistically, CHI3L1 increased TGF-{beta}1-stimulation of Smad, Akt and Erk signaling and PD-L1 played a significant role in TGF-{beta}1/CHI3L1-stimulated myofibroblast transformation. Coculture experiment further confirmed the ability of CHI3L1 to induce profibrotic macrophage activation that enhanced myofibroblast transformation mediated via a CD44-PD-L1 axis. Following in vivo bleomycin challenge, CHI3L1 transgenic mice exhibited significantly higher levels of PD-L1+ M2 macrophages, PD-L1+/PDGFR+ fibroblasts and increased numbers of PD-1+ and CD45+/PD-1+ cells compared to wild-type controls. Notably, combined treatment with anti-CHI3L1 and anti-PD-1 antibodies, or a bispecific anti-CHI3L1-anti-PD-1 antibody, resulted in greater inhibition of bleomycin-induced fibrosis than either antibody alone. These findings suggest that there is a stimulatory interaction between CHI3L1 and the PD-1/PD-L1 axis in promoting profibrotic macrophage activation and invasive fibroblast differentiation. The results also highlight the potential of bispecific targeting of CHI3L1 and the PD-1/PD-L1 pathway as an effective therapeutic approach for pulmonary fibrosis.

Role of nuclear ATPases in nuclear mechanics and cell migration through confined spaces: opposite effects of BRG1 and cohesin

Suprewicz, L. — 2025-09-21

Deformation of the nucleus often presents a barrier to cell migration through tight spaces, such as those encountered as cells move through tissues or across extracellular matrix barriers. Reorganization of the nucleus to allow its passage through spaces much smaller than its resting diameter requires forces generated by the cytoskeleton, as well as active reorganization within the nucleus driven by ATPases that crosslink or move chromatin. Here, we show that two different nuclear ATPases, the BRG1/SMARCA4 motor of the BAF or SWI/SNF complex and the bifunctional crosslinking and loop extruding complex, cohesin, have opposite effects on the stiffness of isolated nuclei. Inhibition of BRG1 stiffens the nucleus, and cohesin softens it in karyoplasts derived from multiple cell types, including four different cancer cells, fibroblasts, and mesenchymal stem cells. The effects on isolated nuclear stiffness coincide with the effects of these ATPases on the ability of cells to migrate through tight spaces. Stiffening the nucleus inhibits single cell migration through micron-sized pores and the outward migration of tumor cell spheroids into a surrounding collagen matrix. Softening the nucleus by inhibiting cohesin has the opposite effect: it enhances single-cell migration through pores, at least for some cell types, and facilitates the outgrowth of cells from a tumor cell spheroid into the surrounding matrix. These results emphasize the importance of active motions generated within the nucleus for the global mechanics of the nucleus and the way that it deforms in response to externally generated stresses. Statement of significanceCells often move through tight spaces within three-dimensional materials. This constricted motion requires deformation of the nucleus, which is often stiffer and less dynamic than the rest of the cell. Nuclear deformation is achieved in part by forces generated outside the nucleus, primarily by cytoskeletal motors, but nuclear deformation is also affected by intranuclear motor proteins that move and reorganize chromatin. Here we show that the activity of DNA binding chromatin remodeling ATPases affects not only the stiffness of the isolated nucleus but also the ability of cells to deform as they move through tight spaces. Inhibition of motors such as BRG1 that fluidize the nucleus prevents cell movement through tight spaces, whereas inactivation of the crosslinking ATPase cohesin tends to enhance it.

Collective Self-Assessment in Banded Mongoose Intergroup Contests

Rayner, C. W. — 2025-09-25

Contests over resources are widespread in nature. To optimize outcomes, animals assess fighting abilities, deciding to escalate conflicts based on their own strength (self-assessment) or comparing their own strength with that of their rival (mutual assessment). While most research focuses on one-on-one (dyadic) contests, the assessment strategies employed by groups remain poorly understood. Mutual assessment is frequently assumed, as more information is thought to improve decision-making; however, this assumption has rarely been tested. Here we used a dataset spanning 23 years and 641 intergroup contests in a banded mongoose (Mungos mungo) population in Queen Elizabeth National Park, Uganda. Our results support a model of self-assessment: groups with many males tend to escalate conflicts regardless of the rival groups strength, thus contrasting the commonly held assumption that decisions during intergroup contests are made by mutual assessment. We suggest that assessing rival group strength during conflict could be disproportionately costly, compared with assessing own group strength, which can be done over longer time periods and is easier to obtain. Greater understanding of these dynamics can shed light on the drivers and escalation patterns of intergroup conflict across social species, including humans. Lay SummaryWhen two rival groups come together, what determines whether or not they fight? We found support for the hypothesis that banded mongoose groups escalate into physical fights based upon an estimation of their own groups strength, which we term collective self-assessment. We did not find evidence for a commonly held notion that groups compare their own groups strength with their rivals. Distinguishing between assessment strategies is intensively researched in contests between individuals but has rarely been applied to intergroup conflict. Here we applied the successful "assessment strategy framework" to group conflict, analyzing over 20 years of data from over 600 intergroup interactions. We suggest that collective self-assessment is employed because it is faster than mutual assessment, which offers an advantage in conflicts where time is of the essence.

A landscape of intergroup conflict shapes den site choice in banded mongooses

Sankey, D. W. E. — 2025-09-25

BackgroundFor many social animals, intergroup conflict has major impacts on fitness and should therefore influence how groups navigate their environment. Yet most studies of group movement focus on the behaviour of single groups, usually foraging groups studied in isolation of all others. In many systems rival groups present both threats and opportunities, contributing to a landscape of intergroup conflict. This landscape could have a profound impact on group movement. MethodsHere we test how the potential for intergroup conflict influences movement decisions of wild cooperatively breeding banded mongooses, using 14 years of behavioural and GPS data. In this species, encounters between groups can present both risks (e.g. injury/mortality) and opportunities (e.g., extragroup mating). We expected that, to minimise risks and maximise rewards, the motivation to engage in conflict would depend on the groups state. Such that vulnerable groups (those that were babysitting young offspring) would select den sites closer to the core of their territory than groups without young offspring. ResultsWe found support for this pattern; non-babysitting groups were unusually risk prone choosing den locations in areas of frequent use by outgroups. During times of heightened recent conflict even choosing to den closer to the core of their rivals territory than their own. ConclusionsThese results suggest that groups of banded mongooses choose den sites that reflect not only the risks but also the potential rewards of defensive or competitive claims on space. Rival groups thus form an integral part of the social landscape shaping patterns of collective movement.

Preparatory encoding of diverse features of intended movement in the human motor cortex

Rigotti-Thompson, M. — 2025-09-25

Over the course of a voluntary movement, motor cortical activity exhibits a transition from preparation to execution, with markedly different activity across these phases. Preparatory activity in particular might be used to improve brain-computer interfaces (BCIs) that harness brain activity to control external assistive devices, for example by anticipating a users intended movement trajectory for quick and fluid performance. However, to leverage preparatory activity for clinical BCIs, we must first understand which features of upcoming movements are encoded by preparatory activity in humans. In this work, we collected intracortical recordings from 3 research participants in the BrainGate2 clinical trial to investigate whether diverse features of movement, such as direction, curvature, and distance, are encoded by preparatory activity in the human motor cortex. We first show that preparatory activity is tuned to the direction of upcoming movements, and this tuning is largely preserved across movements with different effectors. Further investigation demonstrated this preparatory activity is also informative of initial and endpoint directions of curved movement trajectories, and encodes movement distance and speed independently. Finally, we present an online control paradigm that leverages preparatory activity to predict movements towards intended directions in advance, yielding rapid, self-paced control of a computer cursor by human participants. Altogether, these results demonstrate that preparatory activity in the human motor cortex encodes rich features of upcoming movement, highlighting its potential use for high performance brain-computer interface applications.

Essential Role for Trf2 in Cardiac Development and Function

Sharifi-Sanjani, M. — 2025-09-30

Telomere Repeat-binding Factor 2 (Trf2) is essential for protecting our telomeres. While Trf2 global deletion is lethal, its role in organ-specific development, particularly in the heart, remains less understood. In this study, we investigated the role of Trf2 in cardiac development and function. Our studies reveal that cardiomyocyte (CM)-specific loss of Trf2 leads to profound defects in heart morphology, including impaired ventricular wall formation and compromised CM proliferation, concurrent with no CM telomere length attrition. Further, in vivo functional assessment and molecular analyses of CM-Trf2 deficient ventricles revealed severe cardiac dysfunction and, interestingly, altered nuclear envelope gene expression, respectively. Our work provides new insights into the essential role of Trf2 in heart development and function, and potential avenues for therapeutic intervention targeting telomere biology.

A proteogenomic approach to discover novel lncRNA-derived peptides and their potential clinical utility in hepatocellular carcinoma

Bingwu, L. — 2025-10-01

Peptides are increasingly recognized for their versatile functions in biological contexts but their clinical relevance and utility remain largely unexplored. Proteogenomic approaches can accelerate peptide discovery in clinical samples by integrating proteomic data with genomics and transcriptomics evidence. However, long noncoding RNA (lncRNA)-derived peptides (lncPeps) remain largely unidentified, resulting in unmatchable MS/MS spectra. To solve this problem, we have used high-quality Ribo-seq translatomic datasets to generate an extensive database of human liver lncPeps, which we subsequently applied to proteomics data of tumor-adjacent normal tissue pairs from hepatocellular carcinoma (HCC) patients. Using the new database, we discovered 105 novel lncPeps including lncPeps differentially expressed between tumor and non-tumor tissues, and lncPeps with significant correlation with prognosis. Remarkably, combining the expression of lncPeps with canonical proteins in a LASSO regression model improved predictive performance for recurrence, increasing the AUC by 0.005 to 0.085 across three recurrence time points. These findings suggest that lncPeps discovery contributes to our understanding of the molecular heterogeneity and progression of HCC, and broadens the range of potential biomarker candidates or treatment targets for the disease.

Myeloid Specific Ablation of SHIP1 Boosts ex vivo Expansion and Regulatory Function of Myeloid-Derived Suppressor Cells in Inflammatory Arthritis

So, E. Y. — 2025-10-02

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous cell population and the immunosuppressive function of MDSCs has been well established in tumor microenvironment. Recent studies show that adoptive transfer of MDSCs can ameliorate collagen-induced inflammatory arthritis (CIA), a mouse model of human rheumatoid arthritis (RA). Src homology 2 domain- containing inositol polyphosphate 5-phosphatase 1 (SHIP1) was previously shown to regulate MDSC differentiation. In this study, we aimed to generate immunosuppressive MDSCs from mouse bone marrow (BM) through genetic modification combined with cytokine treatments, and to investigate the ability of these ex vivo induced BM-MDSCs to suppress inflammatory responses in the CIA mouse model of RA. We found that myeloid specific ablation of SHIP1 increased the ratio of MDSCs and enhanced their regulatory functions in cytokine induced BM culture. MDSCs from LysMcre:SHIP1flox/flox mouse BM culture demonstrated stronger inhibitory effect on T cell proliferation than those from control mouse BM. Ex vivo induced MDSCs from either control mice or mice with myeloid specific ablation of SHIP1 were administered to the CIA mice as a cell-based therapy to treat inflammatory arthritis. Adoptive transfer of either BM-MDSCs significantly reduced disease incidence and severity, but SHIP1 deficient BM-MDSCs exhibited even higher efficacy compared to wild-type BM-MDSCs. Furthermore, pharmacological inhibition of SHIP1 enhanced the expression of immune regulatory genes in BM-derived MDSCs, and adoptive transfer of these cells protected against CIA development. In conclusion, myeloid specific ablation of SHIP1 boosts ex vivo expansion and immune regulatory function of MDSCs in experimental inflammatory arthritis. These ex vivo generated BM-MDSCs may provide novel therapeutic opportunities for the treatment of RA and other inflammatory diseases.

Coordination of Anle138b to Silver Results in Selective Reduction of a C-terminal truncated Alpha-synuclein Protein and Increased Aggregate Size

Rue, K. L. — 2025-10-03

Parkinsons disease (PD) is a prevalent age-related neurodegenerative syndrome, partially thought to be caused by a decrease in alpha-synuclein proteostasis. Anle138b = 5-(1,3-benzodioxol-5-yl)-3-(3-bromophenyl)-1H-pyrazole (HL), is undergoing clinical trials as a promising mitigator of alpha-synuclein aggregation. Because complexation to metals is known to modulate the activity of several drugs, we have prepared and characterized: H2L(ClO4), [CuI({micro}-L)]3, and [AgI({micro}-L)]3. To better understand the bioviability of these compounds, we monitored their effects in a cell culture model of alpha-synuclein protein aggregation using human alpha-synuclein pre-formed fibrils (PFFs). Using two different anti-alpha-synuclein antibodies, our data suggests that [AgI({micro}-L)]3 decreases a C-terminal truncated protein that is approximately 12.4 kDa, as well as increases the size and alters the shape of PFF-induced aggregates. This indicates that [AgI({micro}-L)]3 impacts aggregation in a manner different from HL and may serve as a novel tool for studying C-terminal truncation related aggregation chemistry. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/679869v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@4bbda5org.highwire.dtl.DTLVardef@8fc0b7org.highwire.dtl.DTLVardef@1b561f5org.highwire.dtl.DTLVardef@1322902_HPS_FORMAT_FIGEXP M_FIG C_FIG

Improved interpretability in LFADS models using a learned, context-dependent per-trial bias

Shah, N. P. — 2025-10-03

The computation-through-dynamics perspective argues that biological neural circuits process information via the continuous evolution of their internal states. Inspired by this perspective, Latent Factor Activity using Dynamical systems (LFADS, [1]) identifies a generative model consistent with the neural activity recordings. LFADS models neural dynamics with a recurrent neural network (RNN) generator, which results in excellent fit to the data. However, it has been difficult to understand the dynamics of the LFADS generator. In this work, we show that this poor interpretability arises in part because the generator implements complex, multi-stable dynamics. We introduce a simple modification to LFADS that ameliorates issues with interpretability by providing an inferred per-trial bias (modeled as a constant input) to the RNN generator, enabling it to contextually adapt a simpler dynamical system to individual trials. In both simulated neural recordings from pendulum oscillations and real recordings during arm movements in nonhuman primates, we observed that the standard LFADS learned complex, multi-stable dynamics, whereas the modified LFADS learned easier-to-understand contextual dynamics. This enabled direct analysis of the generator, which reproduced at a single-trial level previous results shown only through more complex analyses at the trial average. Finally, we applied the per-trial inferred bias LFADS model to human intracortical brain computer interface recordings during attempted finger movements and speech. We show that modifying neural dynamics using linear operations of the per-trial bias addresses non-stationarity and identifies the extent of behavioral variability, problems known to plague BCI. We call our modification to LFADS as "contextual LFADS".

Liver angiocrine myeloid-derived growth factor protects against endothelial dysfunction in pulmonary arterial hypertension

Singh, N. — 2025-10-04

Myeloid-derived growth factor (MYDGF) is a hepatic angiokine with protective effects in systemic vascular beds, but its role in pulmonary arterial hypertension (PAH) is unknown. We hypothesized that hepatic MYDGF deficiency contributes to pulmonary endothelial activation in PAH and that recombinant MYDGF could rescue endothelial injury. In the Sugen-hypoxia (SuHx) rat model, hepatic MYDGF expression was decreased, while pulmonary vascular cell adhesion molecule-1 (VCAM-1) expression was increased. Human hepatic sinusoidal endothelial cells exposed to pro-inflammatory macrophage conditioned media downregulated MYDGF, and recombinant MYDGF restored pulmonary artery endothelial cell resistance to inflammatory activation via MAP4K4-NF{kappa}B signaling. In the Brown University PHiNE PAH cohort (n=41 PAH, n=27 controls), plasma proteomics demonstrated increased MYDGF in PAH patients compared with controls, but MYDGF levels declined with worsening liver stiffness and correlated with higher pulmonary vascular resistance. In the independent Servetus PAH cohort (n=117), higher plasma MYDGF was associated with mortality and right ventricular dilation. Together, these findings demonstrate hepatic MYDGF deficiency in experimental PAH, tissue specificity of endothelial MYDGF to the liver, and MYDGFs potential to mitigate pulmonary endothelial inflammation. However, human data suggest a paradoxical association of elevated circulating MYDGF with adverse outcomes, underscoring the complex biology of angiogenic growth factors in PAH. MYDGF may represent a novel hepatic angiokine linking systemic inflammation, liver dysfunction, and pulmonary vascular disease.

The Liver is an Inflammatory Mediator of Pulmonary Arterial Hypertension

Singh, N. — 2025-10-06

The livers contribution to pulmonary arterial hypertension (PAH) pathogenesis remains unclear. We hypothesized that the liver promotes inflammatory injury to the pulmonary endothelium. PAH patients without liver disease with pulmonary artery endothelial cell (PAEC) biopsies were included. Liver serologies and imaging were analyzed by unsupervised classification and regression tree (CART) to identify subclinical liver dysfunction clusters. Two machine-learning models predicted cluster assignment and informed differential expression. PAEC transcriptomes were compared to liver and lung data from monocrotaline and Sugen-Hypoxia rats. Liver fibrosis was assessed in rat and human PAH livers. Among 25 PAH patients (76% female, median age 61 [30 - 84] years), CART identified clusters distinguished by Model for End-Stage Liver Disease Sodium (MELD-Na) [≥]12, predicting higher pulmonary vascular resistance ({beta}=0.5 Wood units per point increase in MELD-Na, 95% CI 0.2-0.8, p=0.005) after adjustment for right atrial pressure. Subjects with MELD-Na [≥]12 had decreased 6-minute walk distance (353 [120 - 576] m vs. 411[300 - 600] m, p=0.03), with upregulation of apelin, beta-catenin, and immune signaling. Rat lung ECs demonstrated survival and hepatic growth-factor signaling, while rat livers showed immune activation. Rat (20.8 vs 16.6 % area stained, p=0.09) and human PAH livers revealed fibrosis despite absent right ventricular failure, supporting a pathogenic lung-liver axis in PAH.

A parietal grid-like code rotates with cognitive maps but lags rapid behavioral transfer

Yu, L. Q. — 2025-10-07

The neural grid code has been proposed to provide a mechanism for generalization and transfer of relational knowledge between situations enabling rapid adaptation of behavior in novel circumstances. However, to date, very little is known about the dynamics with which grid representations change at context transitions, or how such dynamics relate to downstream behavioral adaptation. Here we tested whether grid representations measured with fMRI rotate to match behavioral goals at context transitions and whether such rotations underlie knowledge transfer. Human participants performed a task that included unsignaled state changes at which the position of multiple target locations abruptly and synchronously rotated by the same degree. After state changes, participants were able to leverage the relative positions of the targets to rapidly infer locations, even novel ones, constituting a form of zero-shot transfer. We observed a cognitive grid-like code in the right posterior parietal cortex with a consistent phase angle that rotated with the relative positions of the targets. However, this rotation was too slow to account for rapid improvements in performance after a state change, and instead these improvements were more closely related to representations of the identity and location of spatial targets in the frontoparietal and orbitofrontal cortex, respectively. Our results highlight the ability of humans to rapidly transfer knowledge and demonstrate that a parietal grid-like code rotates into behaviorally relevant reference frames, but raise questions about the function of such rotations, pointing instead to alternate neural mechanisms for rapid knowledge transfer.

Attention Improves Population Codes by Warping Neural Manifolds in Human Visual Cortex

You, Y.-Q. — 2025-10-10

Decades of research on visual attention have revealed its numerous effects on neural responses. Two competing models have been proposed for how these effects lead to improved population representations: one highlights changes in neural tuning, while the other points towards changes in trial-by-trial noise correlations. Here, we develop a neural population manifold framework that interprets changes in neural responses as geometric transformations in high-dimensional neural space, allowing us to disentangle and quantify the effects of tuning changes and correlation changes induced by attention. Applying this framework to extensive measurements of cortical responses during different attentional tasks, we find that tuning changes are the primary driver of improved population representations. In contrast, correlation changes, though present, have minimal--or even detrimental--effects to information content due to its strong interactions with other changes (e.g., tuning, variability). Our results support the "tuning change" model of visual attention and demonstrate a general framework for adjudicating how different aspects of neural coding affect information processing.

Ancient genomic insights into Salmonella enterica Paratyphi C in Central Mexico

Bravo-Lopez, M. J. — 2025-10-14

Salmonella enterica is a widespread pathogen of major global health relevance, with over 2,500 serovars classified into non-typhoidal and typhoidal groups. Within the typhoidal group, S. enterica Paratyphi C causes paratyphoid fever in humans. Ancient DNA from this bacterium has previously been recovered from epidemic-associated burials in Eurasia and Mexico, dating back 6,000 to 300 years. Here, we analyzed dental DNA from seven individuals (radiocarbon dated to 1800-1940 CE) buried at the Temple of the Immaculate Conception in Mexico City, Central Mexico, and identified ancient S. enterica Paratyphi C DNA in a young female with Native American ancestry. Using an in-house targeted enrichment strategy and deep shotgun sequencing, we reconstructed a S. enterica Paratyphi C genome (COYC5) with [~]11X coverage. Phylogenetic and comparative genomic analyses place COYC5 in close association with previously reported S. enterica Paratyphi C genomes associated with the 1545 cocoliztli epidemic in southern Mexico, as well as with European strains. Divergence estimates indicate that the Mexican and European lineages shared a common ancestor approximately 1,400 years before present (yBP), reflecting an ancient evolutionary split predating European colonization of Mexico. In contrast, the divergence between COYC5 and the southern Mexican genomes occurred around 516 yBP, coinciding with the onset of the colonial period. This pattern supports a European introduction of S. enterica Paratyphi C during colonization, followed by its local diversification within Mexico. Despite this regional differentiation, the conserved presence of key virulence loci--such as SPI-7 and an active shufflon system--across COYC5, southern Mexican, and European genomes underscores the enduring pathogenic potential of S. enterica Paratyphi C. The identification of this bacterium in 19th-century Mexico City provides the first genomic evidence of its persistence in urban contexts beyond major epidemic outbreaks, offering new insights into its evolutionary trajectory in Mexico. These findings raise new questions about how the pathogen spread and persisted across different ecological, social, and epidemiological contexts in ancient Mexico and the Americas.

The Impact of Malaria-Induced Neutrophil Subset Shift and a Link to Burkitt Lymphoma

Okoth, S. A. — 2025-10-14

Burkitt lymphoma (BL) is an aggressive B-cell lymphoma that remains a leading cause of childhood cancer mortality in sub-Saharan Africa. Although the epidemiological link between Plasmodium falciparum (Pf) malaria and BL has been established, our understanding of the underlying immunological mechanisms conducive to tumorigenesis is incomplete. To address a noted gap in our knowledge of the immune landscape, we profiled neutrophil subsets from children with different exposure histories to Pf-malaria and children diagnosed with BL from Western Kenya, along with healthy malaria-naive Kenyan adults. Using multiparameter flow cytometry, we characterized neutrophils by expression of CD15, CD16, CD10, CD11b, CD182, CD184, and CD62L and found that malaria-exposed children exhibited increased frequencies of aged neutrophil subsets, accompanied by a reduction in the mature subset frequencies compared to malaria-naive children. Malaria-exposed children also had neutrophil profiles that closely resembled those seen in the adults. Notably, a positive correlation (rs = 0.7; p < 0.0001) was observed in immature neutrophils between malaria-exposed healthy and BL children, indicating a similar expansion pattern of this subset in both groups. This finding suggests a malaria-driven expansion of the immature subset, potentially promoting a permissive environment for BL. Our data suggests that the observed shift in neutrophil profiles could contribute to the malaria-induced immunopathology associated with BL Visual abstractCreated in BioRender. Forconi, C. (2025) https://BioRender.com/oz60qvq O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/681992v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1640dc4org.highwire.dtl.DTLVardef@1c3e794org.highwire.dtl.DTLVardef@98bf12org.highwire.dtl.DTLVardef@2851b8_HPS_FORMAT_FIGEXP M_FIG C_FIG

Evaluation of a Polarization-Sensitive, Dual-Wavelength Wearable Photoplethysmography Sensor Across a Range of Skin Tones

Jakachira, R. — 2025-10-15

SignificanceHigh-quality photoplethysmography (PPG) signals are essential for accurate extraction of cardiovascular metrics such as heart rate, heart rate variability, and perfusion index. However, signal degradation for individuals with dark skin tones can compromise PPG quality and pose challenges for equitable sensing. AimWe develop a dual-wavelength, polarization-sensitive PPG device to assess perfusion index (PI) across a range of skin tones. ApproachTo evaluate the impact of polarization on PPG signal quality, we record PI for co-polarized (polarized illumination and parallel-aligned polarized detection), and cross-polarized conditions (polarized illumination and orthogonally aligned polarized detection) at 655 nm and 940 nm in participants representing light, medium, and brown skin tone categories. Skin tone classification are based on the individual typology angle (ITA) values derived from the CIE L*b* color space measurements. ResultsAt 940 nm, light from the cross-polarized light channel significantly increases PI (p < 0.05). At 655 nm, cross-polarization presents a statistically significantly enhanced PI (p < 0.05) relative to light from the co-polarized illumination condition, although the magnitude of the improvement decreases with lighter skin tone indication a possible interaction between skin tone and polarization. This improvement is consistent across all skin tones. ConclusionsOur results suggest that the cross-polarized condition improves PPG signal quality by reducing the influence of superficial scattering and enhancing deeper vascular signals. This approach may be especially beneficial for individuals with darker skin tones and offers a promising path towards more robust and inclusive physiological monitoring using PPG-based technologies.

Hunger Recruits a Parallel Circuit Encoding Alcohol Reward

Nunez, K. M. — 2025-10-15

Internal states like hunger, pain, thirst and arousal can bias behavior by affecting sensory and memory processing. Internal states are critical to understand in the context of alcohol addiction because they influence cravings, reinstatement, and relapse. Norepinephrine plays a key role in both hunger and alcohol-induced arousal and preference, but the circuit-level mechanisms through which it modulates the influence of hunger on alcohol preference are not well understood. We sought to address this using intersectional genetic tools for manipulating neurons expressing octopamine, the invertebrate analogue of vertebrate norepinephrine. We identified a single octopamine neuron required for ethanol seeking only when Drosophila are food-deprived. Hunger increased baseline activity in this neuron, making it more responsive to an odor cue previously paired with ethanol. A combination of genetic and connectome analyses revealed that synaptic partners of this octopaminergic neuron form a functional module that acts on Drosophila memory circuitry. Thus, we show that hunger recruits a parallel circuit that drives learned ethanol preference, providing a neuronal framework through which internal state influences the expression of memory for ethanol-associated cues.

AAV Kills Dividing Cells by Depleting PARP1 and Other DNA Damage Response Proteins

Friese, S. — 2025-10-15

Recombinant adeno-associated virus (rAAV) is a replication-defective viral vector used in hundreds of human gene therapy trials, resulting in five FDA-approved therapies. Despite this success, rAAV-based gene therapies suffer from dose-limiting toxicities, resulting in several severe adverse reactions, including death. Previously, we discovered that rAAV rapidly kills mouse NPCs in vitro and in vivo. This vector contains a minimal genome comprised of 145-base pair inverted terminal repeats (ITRs) with a T-shaped hairpin structure that appears to be necessary and sufficient for this toxicity. However, the mechanism for AAV ITR toxicity is not known, and there have been few attempts to engineer ITRs to attenuate rAAV toxicity. In the current study, we explore the molecular mechanisms that drive dose-dependent rAAV toxicity in dividing human NPCs (hNPCs) and test whether disrupting these mechanisms mitigates this toxicity. Recombinant AAV infection induces aberrant cell cycle progression with activation of the ATM /CHK1/CHK2 pathway and expression of the DNA damage markers {gamma}H2AX and 53BP1. Affinity-based proteomics indicate that AAV ITRs bind to Poly-(ADP-Ribose)polymerase 1 (PARP1) and other DNA damage response (DDR) proteins involved in single-strand break repair (SSBR). Recombinant AAV infection attenuates poly-(ADP-ribose) (PAR) formation and mimics the antiproliferative effects of pharmacological PARP inhibitors used in cancer therapy. Moreover, treatment of hNPCs with PARP inhibitors is sufficient to reproduce many features of rAAV-induced toxicity. Finally, we demonstrate that eliminating the T-shaped hairpin within the AAV ITR reduces binding to SSBR proteins and the resulting rAAV toxicity. These findings suggest that rAAV infection induces replication stress and cell death in dividing hNPCs by functionally depleting PARP1 and other DDR proteins that are essential for DNA replication. This work fills substantial gaps in the understanding of the mechanisms of rAAV toxicity and has important implications for the development of safer rAAV-based human gene therapies. One Sentence SummaryThe rAAV genome binds to and depletes PARP1 and other SSBR proteins that are essential for DNA replication, resulting in DNA double stranded breaks, checkpoint activation, and cell death in dividing cells.

Elevation of the mechanically-sensitive protein emerin links nuclear mechanotransduction to tau-induced cytoskeletal remodeling in neurons

Sohn, C. — 2025-10-15

Tauopathies are a group of neurodegenerative disorders, including Alzheimers disease, that are neuropathologically defined by deposition of pathological forms of tau in the brain. While tau is reported to drive neurotoxicity by negatively affecting cytoskeletal, nucleoskeletal, and genomic architecture, the mechanisms mediating tau-induced dysfunction of the cytoskeleton and nucleoskeleton are incompletely understood. Based on proteomic profiling, we identify a suite of cytoskeletal and nucleoskeletal proteins with differing abundance in a cellular model of tauopathy, iTau. Building upon previous findings that pathogenic forms of tau reduce nuclear tension, we find that protein levels of emerin, a central regulator of nuclear mechanotransduction, are significantly elevated in iTau cells and in induced pluripotent stem cell (iPSC)-derived neurons carrying a mutation in the microtubule-associated protein tau (MAPT) gene that causes autosomal dominant frontotemporal dementia. We find that neuronal emerin overexpression is sufficient to drive neurotoxicity, increase overall levels of filamentous actin (F-actin), and induce nuclear invagination, cellular phenotypes that also occur in settings of tauopathy. Mass spectrometry-based identification of emerin-interacting proteins in iTau-derived neurons reveals increased interactions with cytoskeletal proteins and reduced interactions with nuclear proteins. Indeed, we find that emerin relocalizes from the nucleus to the cytosol in the setting of tauopathy, suggesting that pathogenic tau impacts nuclear mechanotransduction pathways. Overall, we identify emerin as a mediator of cytoskeletal remodeling in tauopathy and provide a foundation for future studies into the mechanosensitive function of emerin in neurons. SIGNIFICANCE STATEMENTCells experience and respond to diverse mechanical forces that shape their morphology, function, and survival through a process termed "mechanotransduction." While well studied in non-neuronal cells, neuronal mechanotransduction remains poorly understood despite exposure of the brain to vascular flow, movement, injury, and disease. We identify the mechanosensitive protein emerin as a key regulator of nuclear mechanotransduction in neurons. Emerin overexpression is sufficient to increase filamentous actin, induce nuclear invagination, and drive neurotoxicity, revealing a novel function for emerin in neurons. In cellular models of tauopathy, emerin is elevated and relocalizes from the nucleus to the cytoplasm, where it alters cytoskeletal structure. These findings establish emerin as a mechanosensitive regulator in neurons and link disrupted nuclear neuronal mechanotransduction to neurodegeneration.

Decellularized Meniscus (MEND) as a biomaterial that supports stem cell invasion and chondrogenesis

Bonelli, H. M. — 2025-10-17

BACKGROUNDCartilage damage affects 25 million people globally each year. Tissue engineering strategies such as microfracture and matrix induced autologous chondrocyte implantation (MACI) are currently being used in the clinic; however, they are accompanied by their own limitations such as donor site morbidity, rapid clearance from the injury site, and extensive cost. To overcome these limitations, the tissue engineering field has shown increasing interest in the use of decellularized extracellular matrix (dECM) biomaterials due to their heightened integration with native tissue and regeneration rates. METHODSThe Gottardi Lab has developed a new dECM material sourced from porcine meniscus decellularization (MEND), in which elastin fibers are removed via enzymatic digestion, resulting in channels that can be easily recellularized. RESULTSIn this work we demonstrate that MEND can be seeded with bone-marrow derived mesenchymal stem cells (MSCs), achieving a uniform distribution of cell nuclei throughout the cross section of the scaffold. We also show that MEND retains its native structure in the presence of MSCs and can support chondrogenesis comparably to other commonly used tissue engineering materials such as methacrylated type I collagen and gelatin/hyaluronic acid hydrogels. CONCLUSIONOverall, MEND is a promising new dECM biomaterial for cartilage regeneration.

Ventral striatal astrocytes contribute to reinforcement learning

Pai, J. — 2025-10-20

Astrocytes influence synaptic plasticity and neuronal function through astrocytic calcium dynamics (ACD). However, astrocytes contribution to cognitive operations like reinforcement learning (RL) remains unclear. To examine this, we trained mice on a RL dependent probabilistic decision-making task. We attenuated ACD across distinct striatal regions, finding that ACD attenuation specifically in ventral striatum (VS) increased decision noisiness and impaired reward-guided choice. This effect was largely due to a reduction in "win-stay" behavior. Using in-vivo calcium imaging, we found that VS ACD correlated with reward prediction errors (RPEs). Furthermore, these trial-by-trial ACD fluctuations predicted trial-by-trial choice variability. Finally, ex-vivo slice electrophysiology and computational modeling revealed two mechanisms through which astrocytes could regulate behavioral variability: by regulating presynaptic excitatory-inhibitory balance, and by sharing RPE signals across populations of striatal neurons. Together, these results suggest that VS astrocytes contribute to cortico-striatal functions to mediate decision noisiness.

Tomato pollen tube integrity is critical for pollen function at high temperature and is controlled by RALF signaling.

ALTHIAB ALMASAUD, R. — 2025-10-26

Reproductive success at high temperature (HT) depends on the ability of pollen to germinate a pollen tube that delivers sperm for double fertilization. We recently found that Solanum lycopersicum (tomato) cultivars capable of setting fruit at HT produce pollen tubes that continue to grow under heat stress. Our present goal is to define the molecular basis of thermotolerant pollen tube growth using quantitative live imaging, transcriptomic and proteomic profiling, and genetic analysis. We found that pollen tube integrity - the ability to maintain an intact tip during germination and elongation - was the pollen performance parameter that distinguished thermotolerant cultivars. We identified the components of the Rapid Alkalinization Factor (RALF) pollen tube integrity signal transduction pathway in tomato and found that RALF peptides repress germination at control temperature and that signaling is antagonized by HT. Loss of a single RALF peptide changes pollen tube cell wall pectin distribution and enhances pollen tube integrity at HT in thermotolerant Heinz, revealing that RALF signaling regulates thermotolerant pollen tube growth and establishing a molecular framework for improving reproductive heat-resilience in crop plants.

In Silico Post-screening of Anti-polymerization Agents to Treat Sickle Cell Disease

Qian, Y. — 2025-10-26

Sickle cell disease (SCD) is a genetic disorder that affects approximately 100,000 individuals in the United States and millions globally. Although curative therapies, such as stem cell transplant and gene therapy, are available, their application is limited by high cost and donor availability. Conseuently, drug therapy remains the most feasible treatment option for the majority of patients with SCD. To date, only four drugs have been approved by the FDA, but none of these treatments comprehensively address all SCD-related symptoms or crises, suggesting the pressing need for developing new drugs. Several high-throughput screening campaigns have been performed for SCD drug discovery based on the in vitro sickling of red blood cells (RBCs) and they have identified several hits. However, it is challenging to replicate the organ-specific oxygen level and physiological deoxygenation time in these in vitro RBC sickling assays. Furthermore, these assays do not consider the pharmacokinetics (PK) and pharmacodynamics (PD) of the identified drugs, which are essential to determine whether the drugs can provide robust and sustained efficacy in treated patients with SCD. To address these technical gaps, we have developed a computational platform to perform post-screening analysis of potential anti-sickling agents. This platform sequentially combines PK/PD models with a kinetic model of RBC sickling, enabling efficient prediction of the dosage-dependent therapeutic efficacy of various anti-sickling agents based on patient-specific hematological factors and organ-specific oxygen levels. We first demonstrate the effectiveness of our integrated platform by showcasing the therapeutic efficacy of two FDA-approved drugs, Hydroxyurea (HU) and voxelotor. Next, we evaluate the therapeutic efficacy of two potential anti-sickling agents under clinical trial, namely Bitopertin and osivelotor. Our findings suggest that Bitopertin exhibits anti-sickling effects that are considerably less pronounced than those of HU and voxelotor. On the other hand, osivelotor can achieve similar anti-sickling effects as voxelotor with significantly lower doses due to its improved PK properties. Furthermore, we show the versatility of the proposed platform in predicting the anti-sickling effect of multi-agent therapies and evaluate the conse-quences of drug noncompliance. In particular, our analysis indicates that noncompliance with voxelotor may result in rapid increases in RBC sickling, whereas osivelotor is likely to mitigate noncompliance-induced adverse effects due to improved PK properties. We further quantify the relationship between drug dosage and the duration of noncompliance that leads to loss of thera-peutic efficacy for voxelotor and osivelotor, providing guidance for optimizing dosage strategies to reduce the risk associated with noncompliance. In summary, our in silico platform serves as a valuable tool for post-screening analysis of potential anti-sickling agents by considering their PK and anti-sickling efficacy under patient-specific hemoglobin level and organ-specific oxygen level, thereby gaining insights into their potential therapeutic efficacy alone or in combination before clinical trials.

Topological analysis of neuronal assemblies reveals low-rank structure modulated by cholinergic activity

Hansen, E. C. A. — 2025-10-27

1Neuronal assemblies are fundamental building blocks of brain function (e.g. memory, spatial navigation, eye fixation, etc.). However, the principles underlying the connectivity patterns that support their function have remained elusive. The optic tectum of the zebrafish larva is organized into distinct functional neuronal assemblies. These assemblies display all-or-none preferred activation states and inhibitory competition, mechanisms that improve the decoding of visual information. Here, we combined light-sheet microscopy to capture the dynamics of large neuronal networks ([~]2,000 neurons) in the optic tectum; genetic cell-type markers for studying the physiological and functional properties of tectal assemblies; and techniques from topological data analysis to study the dynamic connectivity patterns that enable the emergence and functional role of the neuronal assemblies. We found that during spontaneous activations, tectal assemblies maintain a tight and stable ratio of E-I activity despite the large increase in activity. Topological analysis of the spontaneous activations indicated a low-rank organization of the assemblies and a discrete number of temporal activation patterns. Finally, we observed that the cholinergic system can modulate the topological features of the assemblies to alter their functional role.

Temporal control of axonal floor plate crossing through a combination of incoherent feedforward and feedback loops of gene regulatory network regulating Robo3 expression

Sudakevitz-Merzbach, R. — 2025-10-27

Spinal commissural neurons play a fundamental role in motor control and sensory perception. Robo3, a receptor expressed on pre-crossing commissural axons, is required for midline crossing. Its downregulation in post-crossing axons is essential for forming synapses with contralateral CNS targets. We demonstrate that, at the transcriptional level, the dynamic expression of Robo3 in pre-crossing neurons is controlled by incoherent feed-forward loops (iFFLs) and negative feedback loops (NFLs) that precisely regulate its transience and thereby determine midline crossing. Lhx2 and Lhx9 activate Robo3 while also inducing its repressor, Barhl2. Additionally, negative feedback loops fine-tune the transience of Robo3 expression by adjusting the relative strength of the activation and repression modules. Our genomic analysis reveals that this regulatory circuitry converges on an enhancer element of the Robo3 gene. These findings imply that diverse iFFLs, together with NFLs, are essential for regulating the extent of midline crossing across different spinal commissural subtypes.

Control of Synaptic Communication through Molecularly Engineered Bioluminescence Light Emission and Sensing

Slaviero, A. N. — 2025-10-28

Synapses are sites of intercellular communication between neurons and from neurons to target organs, and of signal integration that underly physiological and behavioral responses. We have developed a modular platform, Interluminescence (Int), for experimental control of synaptic transmission: bioluminescent light, generated by a luciferase oxidizing a luciferin, from a presynaptic neuron is used to activate transsynaptic optogenetic ion channels in the postsynaptic neuron. Two strategies can activate or silence postsynaptic neurons in vivo in the presence of luciferin. In the Act-Int approach, a luciferase is genetically expressed in synaptic vesicles and released during depolarization-induced presynaptic vesicle fusion and exocytosis. In the Persist-Int approach, a luciferase is tethered to the presynaptic membrane where it can support sustained transsynaptic signaling. Both strategies can activate postsynaptic neurons with similar efficacy. By design, the modularity of the platform permits the use of luciferases and opsins ranging in brightness and light sensitivity, and the luciferase can be targeted to different subcellular regions of the presynaptic neuron. Our results demonstrate the utility and versatility of Interluminescence to mediate synapse-specific transmission that is either activity-dependent or activity-independent.

Repeated evolutionary turnover of vertebrate skeletal muscle myosins

Harvey, C. M. — 2025-10-28

Myosin heavy chain proteins are essential for muscle contraction and nearly every physiological function in animals, but their diversity and evolution outside mammals is largely unknown. We comprehensively model the evolutionary history of over 1100 heavy-chain myosins. We find that skeletal muscle myosins are located in a conserved tandem gene array in all vertebrate species, but repeated gene duplication-loss turnover has surprisingly led to an independently evolved set of core skeletal muscle myosins in each major vertebrate group. Despite these separate derivations of these myosin subfamilies, each major vertebrate group exhibits consistent tissue-specific patterns of subfamily expression and specialized myosin subfamily expression in extreme muscles. Our results show that muscle evolution across vertebrates is not based in conserved orthologous motor myosins, as might be expected for such a core structural protein family. Instead, we find that skeletal muscle myosins have evolved as a shifting cluster of genes that is constantly changing and diversifying to balance the need to maintain core physiology, while innovating new physiological possibilities.

Visualizing TERRA RNA G-quadruplex unfolding in FUS biomolecular condensates

Zheng, T. — 2025-10-29

RNA G-quadruplexes (rG4s) are remarkably stable secondary structures with critical regulatory roles in gene expression, RNA metabolism, and telomere maintenance. However, their behavior within cells remains controversial, partly due to challenges in detecting rG4s in complex environments. Here, we use solution NMR spectroscopy to investigate how condensates formed by the low-complexity and RGG domains of the RNA-binding protein FUS affect the structure of TERRA, a highly stable model rG4 RNA. We show that FUS LC-RGG1 interacts with TERRA in dilute solution and that binding perturbs, but does not disrupt, the G-quadruplex structure. When co-phase separated with FUS LC-RGG1, however, NMR signatures of TERRAs folded state disappear, and the remaining observable resonances indicate an unfolded conformation, even in buffer containing potassium where TERRA rG4 is exceptionally stable when outside a condensate. Quantitative comparisons with a mutant form of TERRA, used as a baseline for fully unfolded RNA, suggest that at minimum a third of TERRA RNA becomes unfolded in the condensed phase. Thus, our results demonstrate that condensates can shift the structural ensemble of rG4 towards unfolded species, offering a potential mechanistic explanation for their apparent lack of stability in vivo and revealing how phase-separated environments may actively modulate RNA structure and function. HighlightsO_LINMR spectroscopy directly probes RNA structure inside biomolecular condensate. C_LIO_LIFUS binding in solution perturbs TERRA RNA G-quadruplex. C_LIO_LIA sizable fraction of TERRA is unfolded in FUS condensates. C_LI

Plasticity promotes persistence across novel environments in experimental microcosms

Harmon, E. A. — 2025-11-03

Phenotypic plasticity--the ability of organisms to adjust their traits in response to changes in their environment--has long been thought to prevent extinction in novel or changing environments. However, there are few tests linking plasticity to population persistence. Here, we show that plasticity promotes population persistence in replicate populations of rotifers. We experimentally exposed 33 clonal lines that varied in plasticity to over 20 novel environments for up to 40 overlapping generations. We found that clonal populations expressing plasticity in morphology and life history traits persisted longer and were less likely to go extinct in novel environments than populations not expressing such plasticity. These results directly link plasticity to population persistence in novel environments and suggest that plasticity can buy time for organisms in a changing world. TeaserA multigenerational experiment finds that environmentally induced traits allow populations to persist in novel environments.

Model-based EEG phenotyping uncovers distinct neurocomputational mechanisms underlying learning impairments across psychopathologies

Ging-Jehli, N. R. — 2025-11-04

BackgroundMajor depressive disorder (MDD), bipolar disorder (BP), and schizophrenia (SCZ) involve learning impairments with poorly understood mechanisms. Understanding both the similarities and differences in these mechanisms is important to guide the development of new, targeted interventions. Methods255 participants diagnosed with MDD (n=54), BP (n=47), SCZ (n=67) or without any diagnoses (CTRL; n=87) performed an associative learning task. Computational modeling quantified the mechanistic interplay between working memory (WM) and reinforcement learning (RL). The latent RL and WM signatures in the EEG dynamics showed shared and distinct neurocognitive mechanisms underlying learning. ResultsAll clinical groups showed learning impairments at the behavioral level. Model-based EEG analyses linked these impairments to distinct patterns in the dynamic interplay between latent RL and WM mechanisms, contrasting with the typical patterns observed in CTRL. SCZ was characterized by reduced neural markers of WM, weakening the cooperative influence of WM onto RL (reduced WM recruitment), and reduced integration of negative feedback. Conversely, MDD was characterized by reduced reciprocal influence of RL onto WM, reducing the tendency to upregulate WM contribution with reward history (impaired WM management). Finally, BP was characterized by deficits in both WM and RL recruitment, along with higher WM decay. ConclusionsBehavioral learning impairments that appear similar across clinical groups can be linked to distinct neurocognitive mechanisms via integrative neurocomputational modeling. Our approach provides insights into the interplay of underlying learning mechanisms and how they manifest differently across psychopathologies. CitationThis manuscript is a preprint version of the later manuscript accepted for publication in Biological Psychiatry: Global Open Science. The content may differ from the final published version following peer review and editorial revisions. Ging-Jehli, N.R., Rac-Lubashevsky, R., Bera, K., Roberts, A., Loder, A., Boudewyn, M.A., Carter, C.S., Erickson, M., Gold, J., Luck, S.J., Ragland, J.D., Yonelinas, A.P., MacDonald III, A.W., Barch, D.M., & Frank, M.J. (2025). Model-based EEG phenotyping uncovers distinct neurocomputational mechanisms underlying learning impairments across psychopathologies. Preprint at bioRxiv.

Phospotyrosine proteomics reveals novel Zap70 and Itk pathway targets downstream of TCR and CAR in Jurkat T cells

Callahan, A. — 2025-11-05

{zeta}-associated protein of 70 kDa (Zap70) and interleukin-2-inducible T cell kinase (Itk) propagate the primary and CD28-integrated phosphotyrosine (pY) signalling, respectively, to achieve full T cell activation. Despite their canonical roles in T cell activation, our understanding of how each kinase controls canonical and noncanonical pY signalling is incomplete. Here, using three T cell activation methods (soluble antibodies, APC- pMHC/TCR, and CD19-CAR/Raji), we evaluated the effects of two novel inhibitors, RDN2150 (RDN, Zap70) and Soquelitinib (Soq, Itk), on T cell activation. We validated the published working concentrations of RDN and Soq on phosphorylation of key T cell signalling proteins and on T cell activation markers, finding that RDN provides more complete inhibition of T cell signalling and activation. We used LC-MS/MS to evaluate how RDN and Soq treatment affected the phosphotyrosine (pY) signalling and proteome of T cells, finding that RDN, as opposed to Soq, completely downregulated the TCR signalling pathway. Finally, we identified new, noncanonical pY sites responsive to RDN and Soq, providing new insights into the pathways regulated by Zap70 and Itk. Together, our work provides a basis for further study on RDN and Soq, as well as a molecular roadmap for the effects of these inhibitors.

Spatial Conformation of Pancreatic Cancer Is Associated with Disease Recurrence after Curative-Intent Total Neoadjuvant Therapy

Cisneros, L. H. — 2025-11-06

Pancreatic ductal adenocarcinoma (PDAC) frequently recurs after total neoadjuvant therapy (TNT) and curative-intent resection. Traditional histopathologic response assessments demonstrate that major pathologic response (<5-10% viable residual cancer) is associated with favorable outcomes. However, most TNT cases achieve only a minor response. We investigated whether the spatial organization of residual PDAC encodes clinically meaningful biology beyond residual tumor burden. In a retrospective cohort of 203 resected PDAC patients, all with minor pathologic response, H&E whole-slide images were segmented into cancer and stroma to subsequently quantify spatial composition (e.g., patch size/density, edge density) and configuration (e.g., patch complexity/compactness, spatial intermixing) to model treatment-resistant tumor architecture. Non-response was associated with a more fragmented interface-rich ecology, higher edge density and diversity, and reduced homotypic aggregation, independent of conventional clinicopathologic features. This demonstrates a link between emergent tissue architecture of treated PDAC and therapeutic resistance. Further, two multivariable, spatial risk models were independently associated with disease-free survival (DFS): (1) cancer mean shape index and stromal shape-index variability (high-risk median DFS 7.23 vs 11.57 months; adjusted HR 1.75, p=0.007) and (2) mean stromal area with edge density (high- vs low-risk adjusted HR 1.94, p=0.006), outperforming traditional treatment response assessments. Quantifying residual cancer-stroma topology thus yields independent, prognostic signals in post-TNT PDAC and motivates prospective, spatially informed adjuvant strategies and mechanistic studies of edge habitats and mixing as therapeutic vulnerabilities.

Unconscious switching of dorsal and medial pathways for plasticity and stability during NREM and REM sleep

Yamada, T. — 2025-11-06

While visual perceptual learning improves during non-REM sleep and stabilizes during REM sleep via excitatory-inhibitory neurotransmitter (E/I) balance in early visual areas (EVA), the role of prefrontal regions remains unclear. Here, we show that contributions of the dorsolateral prefrontal cortex (DLPFC) and medial prefrontal cortex (mPFC) differ by sleep stage in human adults. During non-REM sleep, plasticity increased in DLPFC--indexed by elevated E/I balance measured with magnetic resonance spectroscopy and polysomnography--in correlation with performance gains. During REM sleep, stability increased in mPFC--indexed by reduced E/I balance--in correlation with resilience to retrograde interference from new learning. E/I balance changes and their effects on learning paralleled those in EVA. Connectivity weights between EVA and DLPFC, and between EVA and mPFC, switched with sleep stage. These findings suggest the presence of dorsal and medial pathways that unconsciously alternate between non-REM sleep and REM sleep to improve and stabilize learning.

Rhythmic light stimulation elicits multiple concurrent neural responses that separably shape human perception

Griffiths, B. J. — 2025-11-06

Rhythmic light stimulation offers solutions to innumerable cognitive and neurological disorders. However, like any neuromodulatory technique, responses to rhythmic light stimulation are highly variable, producing challenges in replicating lab-based studies and translating findings to the clinic. Across three MEG/EEG experiments, we show that this variability can, in part, be attributed to rhythmic light stimulation eliciting multiple, coexisting neural responses which have separable impacts on cognition. Specifically, we find that rhythmic light stimulation produces distinct neural responses at the fundamental (f) and second harmonic (2f) frequencies, and that these responses are differentially shaped by endogenous oscillatory dynamics that vary across participants. Importantly, these responses separably contribute to perception, with harmonic gamma-band responses supporting the representation of stimulus-specific information, and the phase of harmonic alpha-band responses causally contributing to near-threshold visual perception. We reproduce these effects across datasets, paradigms, and oscillatory bands, suggesting that the multiplex oscillatory responses elicited by rhythmic light stimulation are a robust and pervasive phenomenon. We propose that the complexity of neural responses to rhythmic stimulation can explain why there is substantial variability between studies using these techniques, and that understanding these complex responses may help advance neuromodulatory technologies for both fundamental and clinical neuroscience. Supplementary MaterialAll supplementary material can be found at the end of this document.

Inferring the regulation dynamics of oscillatorynetworks from scRNA-seq data

Zhao, W. — 2025-11-10

Oscillatory processes such as the cell cycle play critical roles in cell fate determination and disease development. Yet, most current gene regulatory network (GRN) inference methods are based on gene-gene correlations or temporal progression, not adequately accounting for the recurrence in cyclic processes. We hypothesize that constraining the continuous ordering of relative positions along the cell cycle can enhance GRN inference accuracy of cell cycle regulation. To test performance, we evaluated eight representative methods and applied three of them to a mouse retinal progenitor single-cell gene expression dataset [1]. Incorporating cell cycle positions inferred by Tricycle [2] led to significant improvements compared against using experimental times, particularly for early progenitor cells that been hypothesized to be more intrinsically driven by cell cycle regulation. These findings highlight the promise of integrating oscillatory processes into causal inference frameworks to advance our understanding of gene regulation.

Ribosomal Architecture and rRNA Modification Landscape in the Tick-Borne Parasite Babesia divergens

Gutierrez-Vargas, C. — 2025-11-11

Babesia is a tick-borne intracellular apicomplexan parasite responsible for diseases ranging from mild to fatal, with a broadening geographic distribution. Due to the complex life cycle of Babesia species, their survival depends on the precise control of gene expression, which is primarily regulated by epigenetic, transcriptional, and post-transcriptional mechanisms. High-resolution structural information on key components of the translation machinery, such as ribosomes, could aid in the development of antiparasitic drugs. Here, we report cryo-EM ribosome structures (2.6 [A]) from the tick-borne apicomplexan pathogen Babesia divergens, showing associated tRNAs, an mRNA fragment, and RACK1, a signaling scaffold crucial to translation regulation. Density map analysis displays ribosome regions at atomic resolution (1.7 [A]), which, when combined with nanopore sequencing, enabled the comprehensive identification of rRNA modifications, including modifications unreported in other organisms. The new rRNA modifications localize not only to the reduced Babesia rRNA expansion segments but also to functionally essential ribosomal sites, uncovering new avenues for therapeutic intervention against babesiosis.

Selective distractor representations resolve multidimensional interference

Gheza, D. — 2025-11-17

How can humans manage multiple sources of information competing for attention? To approach this question, we adopted a multi-dimensional task-set interference paradigm that requires individuals to handle distractions from three independent dimensions. Behavioral results suggest that people track prior interference from each dimension to selectively modulate their attentional gain. Testing the mechanism of this adaptation at the neural level requires measuring multi-dimensional task representations. To achieve this, we applied representational similarity analyses and encoding models to human EEG and investigated how the history of interference simultaneously affected the time-resolved representations of target and distractor dimensions. EEG analyses revealed that target and distractor features are initially encoded in parallel, but distractors are rapidly suppressed around 250 ms, with suppression scaling with prior interference. Next, we introduced a task-specific proportion-congruency manipulation to study how learning the control demands of each task dimension shapes the proactive handling of multidimensional distractors. Proactive task-level control enhanced the efficiency of the same reactive suppression mechanism observed for trial-to-trial adaptation, without producing sustained preparatory changes. Finally, behavioral and neural effects converged to show that the strength of dimension-specific interference and adaptation scales with the speed of visual integration for each dimension, as captured by a connectionist model with temporal integration. Altogether, these results suggest that proactive control mechanisms operate on the speed and efficiency of a reactive suppression mechanism that constrains selective distractor representations from biasing the response process. More broadly, they show that multidimensional attentional control relies on selectively suppressing distractor representations after they are encoded, revealing a dynamic, dimension-specific mechanism that extends biased-competition and conflict-monitoring theories.

Insular error network enables self-correcting intracranial brain-computer interface

Weger, P. — 2025-11-17

Error recognition is fundamental to adaptive behavior, enabling rapid compensatory action when outcomes deviate from expectations. Central to this function are neural circuits for performance monitoring, encoding cognitive signals that could support more reliable neural interfaces. Here, we recorded intracranial electroencephalography (iEEG) in epilepsy patients to enable a motor brain-computer interface (BCI) while sampling error-related activity across a distributed network. Our work reveals high-frequency population dynamics emerging in the anterior insula and propagating to the prefrontal cortex as the interface fails to follow the users intention. We identify spatially organized insular responses to error processing and movement feedback, highlighting it as a heterogeneous hub linking action and outcome. Real-time integration of error responses enables a self-correcting neural interface that enhances usability by reducing the need for manual user intervention. Together, our work demonstrates a human intracranial BCI harnessing insular brain activity, integrating cognitive processes directly into device control.

Methamphetamine Conditioned Place Preference in Adolescent Mice: Interaction Between Sex and Strain

Severino, L. N. — 2025-11-18

BackgroundMethamphetamine (METH) use is higher in adolescent women than men. While rodent studies support a sex difference in the reinforcing effects of METH, few have investigated sex differences in the underlying neural circuits, none of which tested rodents during adolescence. AimsInvestigate whether there are sex differences in the rewarding effects of METH in two strains of adolescent mice that are commonly used to generate transgenic lines. Identify changes in associated neural circuits. MethodsWe tested METH-induced conditioned place preference (CPP) in male and female 129Sv/Ev and C57Bl/6 mice during middle adolescence using 1mg/kg of METH. Behaviorally-induced upregulation of c-Fos protein expression was quantified in the nucleus accumbens (NAc) and area CA1 of the hippocampus following the post-conditioning test. ResultsIn C57Bl/6 mice, METH induced CPP in females, but not males. Conversely, METH-induced CPP in 129Sv/Ev males, but not females. In both strains, groups that exhibited CPP had more c-Fos+ cells in the NAc and CA1 when compared to saline-treated control groups. The number of c-Fos+ cells in these two brain regions correlated in groups exhibiting CPP, indicating increased NAc-CA1 communication during retrieval of the conditioned memory. Finally, we found evidence of behavioral sensitization in 129Sv/Ev males only. ConclusionsOur study reveals that the rewarding effects of the METH in adolescent mice are both sex- and strain-dependent, indicating that METH response may result from an interaction between sex-specific and genetic mechanisms. Our findings will be informative when selecting an appropriate background strain in future studies using genetically modified mice.

CDH3 as a Novel Therapeutic Target in Basal-like Double-Negative Prostate Cancer

Liu, G. — 2025-11-19

PurposeBasal-like (also known as double-negative) prostate cancers are aggressive tumors that lack effective targeted therapies. We aimed to delineate the role of CDH3 (P-cadherin) in basal-like prostate cancer and evaluate CDH3-directed therapeutic strategies. MethodsWe integrated genetically engineered mouse models (GEMMs) of prostate cancer, bulk and single-cell transcriptomic analyses, and a suite of in vitro and in vivo experiments. CDH3 expression and associated signaling pathways were examined in Pten/Apc double-knockout mouse prostates and human datasets. Functional studies included antibody-drug conjugate (ADC) cytotoxicity assays and the development of chimeric antigen receptor (CAR) T cells targeting CDH3, tested in prostate cancer cell lines and xenograft models. ResultsPten/Apc double deletion in prostate GEMMs led to highly aggressive tumors with markedly elevated CDH3 expression and enrichment of non-canonical WNT signaling components. Transcriptomic analyses of patient-derived prostate tumors confirmed that CDH3 is significantly upregulated in basal-like prostate cancer subtypes relative to luminal subtypes. Single-cell RNA sequencing revealed CDH3 expression predominantly in basal epithelial cells. Mechanistically, we found that active YAP1 signaling and a WNT5A-ROR2 non-canonical WNT axis drive CDH3 expression. Targeting CDH3 with a CDH3-specific ADC induced potent, antigen-dependent killing of CDH3 prostate cancer cells in vitro and significantly suppressed tumor growth in in vivo metastatic prostate cancer models. Likewise, CDH3-targeted CAR T cells specifically recognized and lysed CDH3-expressing prostate tumor cells while sparing CDH3-negative cells, leading to tumor regression and improved survival in mouse models, especially when combined with PD-1 checkpoint blockade. ConclusionsCDH3 is a key marker and functional driver of basal-like prostate cancer. Therapeutic strategies leveraging CDH3, including ADCs and CAR T cells, demonstrate strong preclinical efficacy, supporting the development of CDH3-targeted treatments to overcome resistance in aggressive prostate cancer.

A New Method for Optimal Placement of Tumor Treating Fields Electrodes

Weise, K. — 2025-11-20

OverviewTumor Treating Fields (TTFields) provide a non-invasive treatment option for newly diagnosed glioblastoma. While optimization of electrode placement is important to increase treatment efficacy, clinical therapy planning is done using an undisclosed and proprietary software (NovoTAL(R)), which is clinically unvalidated. This study investigates a new computational approach for optimizing TTFields electrode placement and is compared to the current clinical standard. MethodsWe developed a new computational pipeline integrating patient-specific anatomical data to optimize electrode configurations in five representative glioblastoma cases with diverse tumor locations and sizes. Two optimization strategies were employed: one maximizing electric field intensity at the tumor, and another enhancing coverage of the adjacent brain while maintaining sufficient tumor intensity. Results were compared to electrode placements generated by NovoTAL(R). Additional simulations with artificial tumors assessed the effects of tumor size and location. ResultsOptimized electrode placements improved electric field intensity in tumors by 18%-34% compared to the clinical standard. Coverage-weighted optimizations provided broader field coverage without significantly compromising tumor intensity. Smaller or surface-adjacent tumors benefited most from optimization, achieving precise targeting and enhanced coverage. Extensive randomized placement analyses highlighted the superior performance of the optimized configurations. Analysis of artificial models showed consistent improvements across varying tumor locations and sizes. ConclusionPersonalized optimization of TTFields electrode placement significantly improves electric field targeting of tumors and adjacent brain regions. This approach outperforms standardized planning software and clinical practices and supports future development of adaptive, automated strategies for individualized TTFields therapy in glioblastoma. Keypoints- Optimized TTFields array placement enhanced field intensity by 18-34% vs. clinical standard. - Optimized TTFields planning improved field coverage in tumor-adjacent regions. - Optimized TTFields planning outperformed standard methods and random placement. Importance of the studyTTFields are increasingly used as adjuvant therapy for glioblastoma. However, current individualized treatment planning relies on proprietary, undisclosed, and clinically unvalidated software, limiting transparency, optimization, and innovation in the field. This study introduces an individualized, semi-automated, and open-source method for optimal electrode placement based on standard MRI data, addressing a critical need for validated and adaptable planning tools. Our approach increased field intensity in tumors by 18-34% and achieved broader coverage compared to the standard clinical tool (NovoTAL(R)), without compromising therapeutic strength. Notably, the method also consistently outperformed extensive random electrode placements across diverse tumor types and sizes, highlighting its robustness and ability to achieve true optimal configurations. Open-source availability enhances reproducibility and clinical translation, representing a significant step toward more effective, individualized TTFields therapy. This advancement has the potential to improve outcomes for glioblastoma patients and underscores the importance of technology validation in neuro-oncology.

Diffusion-based Representation Integration for Foundation Models Improves Spatial Transcriptomics Analysis

Jain, A. — 2025-11-21

MotivationWe propose DRIFT, a framework that integrates spatial context into the input representations for foundation models by leveraging diffusion on spatial graphs derived from spatial transcriptomics (ST) data. ST captures gene expression profiles while preserving spatial context, enabling downstream analysis tasks such as cell-type annotation, clustering, and cross-sample alignment. However, due to its emerging nature, there are very few foundation models that can utilize ST data to generate embeddings generalizable across multiple tasks. Meanwhile, well-documented foundational models trained on large-scale single-cell gene expression (scRNA-seq) data have demonstrated generalizable performance across scRNA-seq assays, tissues, and tasks; however, they do not leverage the spatial information in ST data. We use heat kernel diffusion to propagate embeddings across spatial neighborhoods, incorporating the local neighborhood context of the ST data while preserving the transcriptomic representations learned by state-of-the-art single-cell foundation models. ResultsWe systematically benchmark five foundational models (both scRNA-seq and ST-based) across key ST tasks such as annotation, alignment, and clustering, ensuring a comprehensive evaluation of our proposed framework. Our results show that DRIFT significantly improves the performance of existing foundational models on ST data over specialized state-of-the-art methods. Overall, DRIFT is an effective, accessible, and generalizable framework that bridges the gap toward universal models for modeling spatial transcriptomics. Availability and ImplementationCode and data available at https://github.com/rsinghlab/DRIFT. Contactritambhara@brown.edu Supplementary informationSupplementary notes are provided with the manuscript.

JADE: Joint Alignment and Deep Embedding for Multi-Slice Spatial Transcriptomics

Guo, Y. — 2025-11-25

As spatially resolved transcriptomics (SRT) datasets increasingly span multiple adjacent or replicated slices, effective joint analysis across slices is needed to reconstruct tissue structures and identify consistent spatial gene expression patterns. This requires resolving spatial correspondences between slices while capturing shared transcriptomic features, two tasks that are typically addressed in isolation. Multi-slice analysis remains challenging due to physical distortions, technical variability, and batch effects. To address these challenges, we introduce Joint Alignment and Deep Embedding for multi-slice SRT (JADE), a unified computational framework that simultaneously learns spatial location-wise alignments and shared low-dimensional embeddings across tissue slices. Unlike existing methods, JADE adopts a roundtrip framework in which each iteration alternates between alignment and embedding refinement. To infer alignment, we employ attention mechanisms that dynamically assess and weight the importance of different embedding dimensions, allowing the model to focus on the most alignment-relevant features while suppressing noise. To the best of our knowledge, JADE is the first method that jointly optimizes alignment and representation learning in a shared latent space, enabling robust multi-slice integration. We demonstrate that JADE outperforms existing alignment and embedding methods across multiple evaluation metrics in the 10x Visium human dorsolateral prefrontal cortex (DLPFC) and Stereo-seq axolotl brain datasets. By bridging spatial alignment and feature integration, JADE provides a scalable and accurate solution for cross-slice analysis of SRT data.

From Circles to Signals: Representation Learning on Ultra-Long Extrachromosomal Circular DNA

Li, J. — 2025-11-25

Extrachromosomal circular DNA (eccDNA) is a covalently closed circular DNA molecule that plays an important role in cancer biology. Genomic foundation models have recently emerged as a powerful direction for DNA sequence modeling, enabling the direct prediction of biologically relevant properties from DNA sequences. Although recent genomic foundation models have shown strong performance on general DNA sequence modeling, their application to eccDNA remains limited: existing approaches either rely on computationally expensive attention mechanisms or truncate ultra-long sequences into kilobase fragments, thereby disrupting long-range continuity and ignoring the molecules circular topology. To overcome these problems, we introduce eccDNAMamba, a bidirectional state space model (SSM) built upon the Mamba-2 framework, which scales linearly with input sequence length and enables scalable modeling of ultra-long eccDNA sequences. eccDNAMamba further incorporates a circular augmentation strategy to preserve the intrinsic circular topology of eccDNA. Comprehensive evaluations against state-of-the-art genomic foundation models demonstrate that eccDNAMamba achieves superior performance on ultra-long sequences across multiple task settings, such as cancer versus healthy eccDNA discrimination and eccDNA copy-number level prediction. Moreover, the Integrated Gradient (IG) based model explanation indicates that eccDNAMamba focuses on biologically meaningful regulatory elements and can uncover key sequence patterns in cancer-derived eccDNAs. Overall, these results demonstrate that eccDNAMamba effectively models ultra-long eccDNA sequences by leveraging their unique circular topology and regulatory architecture, bridging a critical gap in sequence analysis. Our codes and datasets are available at https://github.com/zzq1zh/eccDNAMamba.

AlphaFlex: Ensembles of the human proteome representing disordered regions

Liu, Z. H. — 2025-11-25

Over a third of residues in the canonical human proteome are predicted to fall within intrinsically disordered protein regions (IDRs), which do not adopt stable folded structures. These IDRs play critical roles in biological regulation and organization, including as targets for post-translational modifications, scaffolds and mediators of biomolecular condensates. To address the pressing need for valid structural models providing biological relevance and enabling functional insight, we developed the AlphaFlex workflow, using IDPConformerGenerator or IDPForge to calculate fully atomistic conformer ensembles for proteins predicted to have disordered regions, modeled in the context of highly confident folded domains from AlphaFold2. We illustrate our approach by generating conformational ensembles of the human proteins in the AlphaFold2 database, with completed AlphaFlex models deposited in the Protein Ensemble Database that is mirrored in UniProt. This transformative resource of AlphaFlex ensembles provides more realistic and biologically relevant full-length protein models for proteins with IDRs, which we illustrate for scaffold proteins with folded domains connected by IDRs, those with IDRs that interact with folded domains, regulatory and condensate proteins requiring exposed binding elements, and a conditionally folding IDR.

COATswga: A Coverage Optimizing and Accurate Toolkit for fast primer design in selective whole genome amplification

Zuckerman, K. — 2025-11-27

BackgroundDespite the transformative nature of next-generation sequencing in genomics, efficiently capturing underrepresented microbial DNA from complex biological mixtures, such as pathogens from host samples, remains a persistent challenge. Unwanted genomes make whole-genome sequencing (WGS) inefficient and expensive without enrichment of the targeted genome. Selective whole genome amplification (sWGA) uses primers that selectively bind to the target genome to preferentially amplify and enrich an entire microbial genome. However, existing sWGA primer design is often complicated and time-consuming, with primers usually having biased amplification producing uneven genome coverage. Developing faster primer design methods that ensure uniform, reliable microbial genome recovery are critical to improving sWGA. MethodsWe developed COATswga, a Coverage Optimizing and Accurate Toolkit for designing sWGA primer sets. The pipeline consists of three key steps: (1) primer discovery, using k-mer counting to identify all candidate primers in the target genomes; (2) filtering, where primers are screened for amplification potential, thermodynamic stability, and specificity; and (3) set formation, which uses a novel interval-based tiling algorithm. COATswga is parallelized for efficiency and supports pre-existing primer set refinement and gap filling. The pipeline was evaluated by designing primer sets for Plasmodium falciparum (strain 3D7) and benchmarking their performance against sets generated using similar pipelines. ResultsCOATswga-designed primers demonstrated superior enrichment based on both qPCR and long-read sequencing. In P. falciparum-spiked human DNA samples at parasitemia levels of 400, 100, and 10 parasites/L, COATswga primers achieved higher amplification efficiency and specificity compared to alternatives. At 100 parasitemia, COATswga yielded [~]99% of reads mapping to the P. falciparum genome with 82.5% of the genome having 5x coverage. Even at 10 parasites/uL, COATswga primers enabled effective amplification of key drug resistance genes (pfcrt and pfmdr1). Additionally, integration with molecular inversion probe (MIP) genotyping showed that COATswga greatly improved sequencing depth and target recovery in low-density infections compared to established sWGA primers. ConclusionCOATswga provides a robust and flexible solution for designing sWGA primers to reliably recover targeted genomes from complex samples. The new P. falciparum primer set performs well in low-parasitemia samples allowing more extensive application of sWGA in falciparum malaria genomics.

PD-1 regulates CD4+ T cell-mediated CD8+ T cell responses in the brain to balance viral control and neuroinflammation

Butic, A. B. — 2025-12-01

Programmed cell death protein 1 (PD-1) is expressed by T cells during progressive multifocal leukoencephalopathy (PML), a life-threatening brain disease caused by the human-only JC polyomavirus. Why PD-1 blockade finds variable success in PML patients is unclear. Brain CD4+ and CD8+ T cells are PD-1high during mouse polyomavirus (MuPyV) encephalitis. Here, we show that PD-1 loss during MuPyV infection acts in a brain-autonomous manner to increase the magnitude of brain-infiltrating CD4+ and CD8+ T cells and the function of virus-specific CD8+ T cells; in concert, brain virus levels decline and neuroinflammation increases. Deletion of PD-1 in CD4+ T cells, but not CD8+ T cells, recapitulates effects of global PD-1 loss. Single-cell RNA sequencing shows that PD-1-deficient CD8+ T cells cluster as effectors while transcripts associated with proliferation and function are upregulated with loss of PD-1. Thus, CD4+ T cell-intrinsic PD-1 signaling balances antiviral defense against neural injury during polyomavirus CNS infection.

Widespread sex-biased gene expression reflects female-biased longevity in a species with environmental sex determination

Bock, S. — 2025-12-01

Sexes frequently differ in life history traits including body size, lifespan, and age at sexual maturity. Aging, the progressive decline in physiological function and cellular resilience over time, is a central process contributing to sex-specific life histories, yet the mechanisms driving sex differences in aging remain largely unresolved. Long-term mark-recapture efforts revealed a striking pattern of female-biased longevity in the painted turtle (Chrysemys picta), a species with temperature-dependent sex determination. As a result, this species provides a compelling system to examine the mechanisms of sex-specific aging in the absence of sex chromosomes. Here, we characterize sex- and age-associated patterns in the blood transcriptomes of wild painted turtles (n = 93). We identified widespread gene expression differences between females and males (2,347 genes; 13.4% of all filtered genes). In contrast, only six genes showed significant linear relationships with continuous age in both sexes. We also employed a machine learning approach which identified distinct sets of genes for which expression was predictive of age in each sex. Age-related gene expression patterns highlight both conserved molecular pathways with known roles in aging as well as novel gene targets. These findings suggest sex-specific molecular processes underlie sex-biased demographic aging and raise questions regarding the environmental and developmental drivers of sex-biased gene expression.

{Phi}X174 bacteriophage viability predicted by protein biophysical modeling

Van Leuven, J. T. — 2025-12-02

The relationship between genotype and phenotype underlies our ability to understand and predict evolution. Efforts to build genotype-phenotype (GP) maps have revealed several unifying rules: epistasis is pervasive, fitness effects are not normally distributed, and the GP map is non-linear and complicated for high-level phenotypes. A critical step in developing GP maps is evaluating how well predictive models do in explaining observed phenotypes. We utilize the simplicity of a bacteriophage ({Phi}X174) study system to test if an intermediate phenotype (predicted stability of the G capsid protein) explains more complex phenotypes. In doing so, we compare the predictive performance of free energies of folding and binding obtained using numerous molecular modeling methods as well as phylogenetic and basic biochemical/biophysical properties of amino acid substitutions. By creating a large mutational library, we find that {Phi}X174 tolerates about 50% of the amino acid substitutions we inserted into the G protein and that molecular modeling compliments other substitution models for predicting viability. Mutations predicted to have large destabilizing effects are especially informative and are almost universally detrimental. These large-effect substitutions often coincide with the most conserved residues in the G protein. Apart from large-effect mutations, our ability to predict {Phi}X174 phenotypes is fairly poor and we explore various potential confounding factors (e.g., codon bias) that could be considered to improve viability predictions.

Behavioral Opportunism and Altered Dopamine Dynamics in Mice Exposed to Early Life Adversity

Gallo, M. E. — 2025-12-08

Early life adversity (ELA) confers risk for reward-related psychopathologies. These risks may stem from adaptations optimizing reward pursuit in anticipation of unreliable, resource poor environments. One rational adaptation to poor, unreliable environments is Behavioral Opportunism: updating expectations more slowly and acting vigorously only when reward is immediately available. To systematically test the impact of ELA on behavioral strategies and underlying reward processing mechanisms, we exposed mice to resource restriction (limited bedding and nesting materials for 7 days) to manipulate the reliability and quality of early life care. Subsequently, we tested adults reward learning and decision making in a two-arm bandit task and recorded dopamine signaling using dLight1.2 fiber photometry in the nucleus accumbens core. Exposure to ELA led to poorer choice discrimination, impaired learning, and decreased adaptation to changes in reward availability. Furthermore, ELA mice were slower to choose between levers but were faster to retrieve immediately available rewards when delivered, consistent with a strategy of behavioral opportunism. Dopamine signaling predicted behavior in both rearing conditions, and its fluctuations were strongly predictive of faster retrieval in ELA mice and an increased likelihood of choice repetition, implying that aberrant dopamine signals underlie slowed learning and vigorous action for immediately available rewards. To understand key features of maternal interactions driving these effects, we used home cage video monitoring to quantify maternal behaviors, continuously, across early life. We found that specific experiential outcomes, such as maternal kicking, intensified behavioral opportunism in adults, predicting poorer bandit task performance beyond the group effect of ELA. Behavioral opportunism provides an explanatory framework for interpreting altered reward processing and reward pursuit in adulthood for individuals exposed to ELA.

Phasic dopamine encodes persistent attraction to reward cues

Townsend, E. S. — 2025-12-12

Reward-predictive cues can become attractive themselves due to incentive salience attribution. This manifests as persistent engagement with cues, known as sign-tracking. Cue-evoked phasic dopamine in the nucleus accumbens core (NAc) is critical for sign-tracking and cue-evoked reward craving, yet is also known to follow learning rules requiring outcome value updating. Would cue-evoked dopamine track incentive salience persistence, or change in accord with reward value alteration while failing to encode persistent behaviors? We assessed NAc dopamine activity using fiber photometry in sign-tracking rats during an omission task, in which physical cue interaction cancels reward. Animals adapted to reward loss while maintaining non-physical cue interaction reflective of persistent incentive salience. Cue-evoked dopamine remained unchanged during omission, mirroring robust incentive salience but not behavioral adaptations occurring with learning. In contrast, outcome-related dopamine reflected reward gain or loss. Therefore, cue-related dopamine encodes persistence of cue value, independent of outcome-related dopamine signals tracking reward occurrence.

RNA modulates FUS condensate assembly, dynamics, and aggregation through diverse molecular contacts

Zheng, T. — 2025-12-13

Fused in sarcoma (FUS) is an RNA-binding protein that undergoes phase separation with RNA and other cellular components, forming ribonucleoprotein (RNP) granules. While recent advances delineating the molecular forces that underlie phase separation have largely focused on protein-protein interactions (1-6), the molecular details of protein-RNA interactions within condensates remain limited. In this study, we demonstrate that RNA modulates the phase separation of the low-complexity (LC) and arginine-glycine-glycine motif (RGG1) domains of FUS: low RNA concentrations enhance protein phase separation and excess RNA disrupts it. By integrating biochemical assays, NMR spectroscopy, and molecular dynamics simulations, we show that RNA incorporates into FUS condensates, reducing condensate density while enhancing local relaxation and diffusional motion of FUS. Surprisingly, whereas RNA binding in the dispersed phase primarily involves the RGG1 domain, within the condensed phase, both LC and RGG1 domains contribute to interactions with RNA. NMR and simulation data show diverse interactions between amino acids and RNA moieties, including prominent glutamine-RNA contacts, that stabilize FUS-RNA co-condensates. Furthermore, we found that RNA accelerates the liquid-to-solid transition of FUS LC-RGG1 condensates, promoting fibrillar aggregate formation. Together, these results provide mechanistic insight into how RNA regulates the assembly, dynamics, and maturation of protein condensates. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/694118v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@19168a9org.highwire.dtl.DTLVardef@16e6878org.highwire.dtl.DTLVardef@18cb8a9org.highwire.dtl.DTLVardef@1af6a4c_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO Using NMR and molecular simulations, we map how RNA engages FUS LC-RGG1 within condensates through electrostatic, {pi}-stacking, and hydrogen-bond contacts. We find that RNA incorporation dilutes condensate density, tunes protein mobility, remodels interaction networks, and accelerates the formation of fibrillar aggregates. C_FIG

Retrotransposon Activation in the Aged and Alzheimer's Disease Brain Examined by Nanopore Long-read DNA Sequencing

Kelsey, M. M. G. — 2025-12-15

BackgroundCellular defenses against retrotransposable elements (RTEs) weaken with age and RTEs have been reported to contribute to Alzheimers disease (AD) pathogenesis by promoting neuroinflammation. The mechanisms implicated include DNA damage promoted by retrotransposition and interferon system activation by RTE-derived cDNA intermediates. LINE-1 (L1) retrotransposons are of particular interest because they are the only autonomously active RTEs in the human genome. ResultsTo investigate L1 activation and retrotransposition in AD, we performed Nanopore long-read DNA sequencing on six late-onset AD (LOAD) and six age-matched control human prefrontal cortex (PFC) samples. We developed and validated a stringent RTE insertion calling pipeline and identified two high-confidence somatic insertions, one AluY and one L1HS. We estimate that [~]1% of cells in the aged PFC have a somatic RTE insertion. AD samples were hypomethylated, and genome-wide analysis of differentially methylated regions (DMRs) supports a process of epigenetic drift in AD. DMR-associated gene sets primarily related to brain function and inflammation. To investigate L1 activation we used CpG methylation as a proxy for L1 expression. We observed decreased methylation at young L1 elements. While most reads overlapping the L1HS promoter were highly methylated (>80% methylated), 7% were <50% methylated, 1% were <25%, and the highly demethylated read fraction increased in AD. L1HS 5 UTR methylation was strongly correlated with RNA expression. ConclusionsCpG methylation-mediated repression of young RTEs is compromised in old age - our findings indicate that this is further exacerbated in AD. Amid these failing defenses, we report somatic retrotransposition events in the aging and demented brain.

Echolocation calls of some bat species in western Uganda

Kloepper, L. N. — 2025-12-15

With the rise of accessible recording technology, passive acoustic monitoring can be an affordable and rapid way to assess species richness, even when individual animals cannot be captured due to regulatory or practical obstacles. Motivated by the relative lack of data and in partnership with the local populace, we recorded echolocation calls of freely-flying bats across six locations in rural western Uganda using opportunistic passive acoustic recordings. Frequency-modulated echolocation calls were recorded at all six locations, while constant-frequency calls were recorded only at sites near entrances to caves. Preliminary species identifications were made using Kaleidoscope Pro, habitat distribution maps for Uganda, and by reference to published work. We make our acoustic recordings publicly available to serve as a resource for further explorations of the richness of bat species in Uganda.

Projection-specific Routing of Odor Information in the Olfactory Cortex

Daste, S. — 2025-12-16

Sensory processing in the mammalian cortex relies on extensive feedforward and feedback connections, yet how information is routed along these pathways remains poorly understood. Here, we examined the functional properties of feedback and feedforward neurons in the mouse olfactory (piriform) cortex. We selectively labeled neurons projecting to the olfactory bulb (OB, feedback) or medial prefrontal cortex (mPFC, feedforward) and recorded their activity during passive odor exposure and learning of an odor discrimination task. We found that odor identity and reward associations were encoded by OB-projecting ensembles early during odor exposure, whereas mPFC-projecting neurons encoded this information later, aligned with behavioral responses. Moreover, mPFC-projecting neurons maintained a stable representation of valence across days, while OB-projecting neurons exhibited pronounced plasticity. Together, these findings reveal that odor information is selectively routed through feedforward and feedback pathways and suggest that the functional properties of piriform neurons mirror the computational demands of their downstream targets.

Site-specific methionine oxidation alters structure and phase separation of TDP-43 C-terminal domain

Ozguney, B. — 2025-12-16

TAR DNA binding protein 43 (TDP-43), a key protein linked to ALS pathology, undergoes phase separation and forms functional assemblies via condensation within cells. The conserved region (CR) within its C-terminal domain (CTD) mediates self-assembly through helix-helix interactions, while the flanking intrinsically disordered regions (IDRs) contribute to phase separation through transient interactions involving aromatic and hydrophobic residues. The CTD contains ten methionine residues distributed equally between these regions, making it particularly susceptible to oxidative modifications. While methionine oxidation is known to impair phase separation, neither the precise mechanism nor the specific contribution of methionines in the CR compared to the IDRs has been determined. Here, we combine NMR spectroscopy and all-atom molecular dynamics (MD) simulations to reveal if and how methionine oxidation in each region differentially affects CTD structure and phase separation. We demonstrate that all methionine residues are vulnerable to oxidation, leading to distinct regional effects: oxidation of CR methionines disrupts helical structure and directly impairs intermolecular helical association, while oxidation of IDR methionines disrupts long-range contacts. Hence, oxidation of methionines in both regions contributes to impaired phase separation, albeit through different mechanisms. These findings establish methionines as critical redox-sensitive modulators of TDP-43 phase behavior and provide molecular insights into how oxidative stress may contribute to TDP-43 dysregulation in neurodegenerative diseases.

Absence of Mothers Curse for performance traits among divergent mtDNAs in heterozygous nuclear backgrounds in Drosophila

Rand, D. M. — 2025-12-18

Maternal inheritance allows selection to act on mtDNA-encoded effects in females but prevents direct selection on mtDNA in males. Mutations that are deleterious in males but neutral or beneficial in females can persist in populations. This predicts that mtDNA-based phenotypic variation should be more common among males than among females, a pattern referred to as Mothers Curse (MC). Most studies of MC place alternative mtDNAs on common homozygous nuclear chromosomal backgrounds, a condition not common in nature. Moreover, it is not known whether MC effects accumulate as mtDNAs acquire nucleotide substitutions between populations or species. We tested the MC hypothesis using mtDNAs from Drosophila melanogaster (OreR, Zimbabwe or w1118), D. simulans (siI and siII) and D. yakuba each placed on several D. melanogaster nuclear backgrounds heterozygous for different chromosomal deficiencies paired with a common w1118 chromosome set. Females and males were tested for starvation resistance, climbing speed, and flight performance. In the majority of chromosomal backgrounds the variance among mtDNA genotypes was greater in females than in males, opposite from the central prediction of Mothers Curse. This suggests that additive and dominance variation across the nuclear genome may provide nuclear blessings that can counter the curse of maternally inherited mtDNA. Teaser textMothers Curse (MC) posits that selection on mtDNA should be stronger in females than in males due to maternal inheritance of mtDNA. This predicts that phenotypic variation among mtDNA genotypes should be lower for females and higher for males. There is conflicting experimental evidence for MC. Most studies of MC have used a common, homozygous nuclear background and have not explored the influence of divergent mtDNAs as strong predictors of MC effects. We address both issues by assaying performance traits among mtDNAs of varying levels of divergence on heterozygous backgrounds. The data fail to support the MC hypothesis and even reveal the opposite effect that females have greater phenotypic variation across mtDNAs. MC may operate in some contexts, but it is not a consistent force in evolutionary genetics.

Revealing the genetic mechanisms underpinning the parasite-induced water-seeking behaviour of insects through RNA-seq

Bhattarai, U. R. — 2025-12-22

The water-seeking behaviour in arthropods infected by mermithid nematodes or nematomorphs is a classic example of host manipulation, yet how these parasites induce the behavioural change is poorly known. We investigated the molecular mechanisms at the basis of this aberrant behaviour in European earwigs (Forficula auricularia) infected with the nematode Mermis nigrescens. We performed comparative RNA-seq analysis at different stages of infection and behaviour manipulation in both the host and the parasite. We detected a total of 12,876 and 9,722 expressed genes in the earwig and nematode, respectively. Differential gene expression (DGE) analysis showed 673 genes upregulated and 593 downregulated in earwigs, and 2,672 genes upregulated and 2,293 downregulated in nematodes across all comparisons between stages of infection and parasite sizes. Clustering analysis produced six and four clusters of differentially expressed genes in earwigs and nematodes, respectively, based on temporal patterns of expression across stages of infection. With overrepresented analysis using gene ontology terms, we found that the earwig shows increased signalling and sensory pathways, while the nematode ramps up transport and secretory genes during manipulation. We found shared GATA transcription factor motif enriched in the differentially over expressed genes during manipulation in both host and the parasite. Our study provides valuable insights into candidate pathways and genes in the host and the parasite, improving our understanding of the molecular regulation of water-seeking behaviour.

Pooled CRISPRi screening reveals fungal-specific vulnerabilities across environments and genetic backgrounds

Wensing, L. — 2025-12-22

The rising rate of drug-resistant fungal infections and the emergence of fungal pathogens with intrinsic resistance phenotypes are a growing concern. The close evolutionary distance between mammals and fungi complicates the design of new antifungals and increases the chances of toxic off-target effects. As such, antifungal drug development usually focuses on fungal-specific proteins when considering potential new targets. Ideal drug targets should mediate essential cell processes and be highly sensitive to inhibition. Targeted gene repression can serve as a model for drug-mediated inhibition and for determining the dosage-sensitivity profile of genes of interest. In the fungal pathogen Candida albicans, classical approaches for gene repression can be labour-intensive and limited to one genetic background due to low throughput. Here, we adapt pooled CRISPRi screening in C. albicans for the first time and exploit this technique for large-scale functional genomic analysis. Through pooled CRISPRi screening, we test the repression sensitivity of over a hundred essential genes conserved in fungi but absent in humans, and successfully identify highly dosage-sensitive genes across multiple cell components and pathways. By extending our analysis to ten diverse environmental conditions, we show how the environment influences dosage-sensitivity profiles. Finally, we extend our experiments to two clinical drug-resistant C. albicans strain backgrounds and demonstrate that many of the fitness defects we observed are conserved in resistant clinical isolates. Together, our results highlight a set of genes that are highly dosage-sensitive across different genetic and environmental contexts, making them attractive targets for further investigation. By facilitating rapid, efficient large-scale functional genomics assays across diverse genetic backgrounds, CRISPRi pooled screening will open new frontiers in C. albicans biology.

Distinct neural temporal architectures encode rapid social expressions and sustained internal mood states

Kakusa, B. — 2025-12-22

Affective processing operates across multiple temporal scales, from rapid social signaling through facial expressions to sustained internal mood states, yet the neural computational principles governing these different timescales remain unclear. Understanding how the brain implements distinct temporal architectures for momentary versus persistent affective phenomena is important to comprehending emotional processing and developing objective biomarkers for psychiatric conditions. Here, we introduced a multimodal approach combining automated facial expression monitoring and continuous intracranial electroencephalography in 2,037 electrode contacts across 16 epilepsy patients, over multiple days. Of these, 15 and 12 patients met criteria for facial expression and for mood analysis, respectively. Among patients meeting criteria, we captured 1,396 naturalistic smiles, and 3,746 neutral expressions - separated by at least 10 seconds, alongside 336 periodic mood assessments. This paradigm revealed distinct behavioral and neural computational architectures. Aperiodic neural activity in the lateral temporal cortex (79.5% accuracy) encoded facial expressions with high cross-participant generalizability. Mood states, however, showed different encoding patterns. Facial expressions provided no consistent mood indicators across participants. Critically, low-gamma power dynamics in limbic regions encoded mood states in only a subset of individuals (5 of 12 participants) with expression-mood behavioral correlations, suggesting a distinct encoding phenotype. Cross-domain analysis confirmed computational independence: neural features optimized for facial expression decoding failed to predict sustained mood states, and vice versa. These findings suggest that multiple neural mechanisms may influence underlying affective processing, with variations in their contributions between individuals. The results provide a framework for understanding individual differences in neural mood representation and establish methodological approaches for objective measurement of naturalistic affective behaviors.

Towards an evolutionary baseline model of Plasmodium falciparum for population-genomic inference

Henry, C. M. — 2025-12-22

Malaria has caused over 15.7 million deaths in the 21st century and was responsible for [~]600 thousand deaths globally in 2023 alone. Although many effective antimalarial drugs have been developed and widely adopted to reduce the occurrence and severity of the disease, recurrent resistance to the frontline treatment has been of major concern. Multiple drug resistance alleles at intermediate and high allele frequency have been identified in specific Asian and African populations of P. falciparum, the deadliest malaria parasite. With the improvement in throughput of sequencing technologies and global efforts such as the MalariaGEN project to build genomic surveillance, we now have access to tens of thousands of genomes of P. falciparum from across the world. With this data, it is becoming increasingly possible to employ powerful population genetics approaches to understand the selective pressures and demographic history of the parasite. While several empirically motivated outlier-based approaches have been employed to identify targets of drug resistance, there is a lack of a framework that jointly accounts for the multiple concurrent processes occurring in natural populations of P. falciparum. We argue that a baseline evolutionary model that accounts for simultaneously acting evolutionary processes is needed to understand patterns of genomic variation in P. falciparum populations. Here, we identify key components essential for building such a baseline model for the malaria-causing pathogen. Such effort will be important to develop an appropriate null model to test evolutionary hypotheses using genomic datasets, will provide a path forward to improve the accuracy of inference of evolutionary parameters, and help identify new gene candidates involved in drug resistance.

The Structural Basis for RNA Binding and Recognition of the Disordered Prion-Like Domain of TDP-43

Puterbaugh, R. Z. — 2025-12-23

Though the structural details of how RNA interacts with folded RNA-binding domains are well established, how intrinsically disordered regions (IDRs) found in a large fraction of RNA-binding proteins mediate contacts with RNA and if they contribute to binding specificity has not been extensively characterized. The human RNA-binding protein TDP-43 is associated with many RNA processing functions that require its predominantly disordered C-terminal domain (CTD) that forms disease-associated inclusions in ALS, and other neurodegenerative conditions. Here, we demonstrate that TDP-43 CTD directly interacts with RNA primarily via a region of the IDR composed of clustered positively charged residues. Large RNAs act as a multivalent scaffold for CTD monomers, inducing the -helical segment of TDP-43 CTD to form multimeric protein-protein structures. Additionally, we probe the nucleotide base and amino acid specificity of CTD-RNA interactions, showing that arginine, aromatic and polar residues display a preference for U and G nucleic acid bases over C and A. Finally, we probe the molecular basis for the strong binding interaction between TDP-43 and G4 quadruplex structures and discover similarly avid interactions with cytosine-rich DNA I-motifs. This work deepens our understanding of how disordered regions of proteins contribute to RNA recognition, drive function, and contribute to disease. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=107 SRC="FIGDIR/small/695782v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@f1d256org.highwire.dtl.DTLVardef@284c2forg.highwire.dtl.DTLVardef@1a1d715org.highwire.dtl.DTLVardef@2fbb1a_HPS_FORMAT_FIGEXP M_FIG C_FIG

Improved short nascent strand sequencing (iSNS-seq) enhances DNA replication origin detection and reduces non-origin biases

Sokka, M. — 2025-12-24

Identifying DNA replication origins in human and other metazoan genomes has been challenging, as highlighted by the fact that various methods for mapping them have produced conflicting results. A popular method, short nascent strand sequencing (SNS-seq or NS-seq), enriches newly replicated short single-stranded DNA by size selection and{lambda} exonuclease ({lambda}-exo) digestion of parental DNA. Surprisingly, SNS-seq has never been validated in Saccharomyces cerevisiae where origins have been well characterized genome-wide. We improved the SNS-seq protocol through biochemical optimization and benchmarked its origin-mapping sensitivity and precision with traditional SNS-seq in asynchronous populations of S. cerevisiae. The improved SNS-seq protocol significantly enhanced the enrichment of origin-derived DNA. S trikingly, the traditional SNS-seq failed to detect known origins and instead enriched non-origin DNA, likely arising from RNA:DNA hybrids. These findings have important implications for the interpretation of previously p ublished datasets that rely on{lambda} -exo for origin mapping. Overall, our biochemical and genomic analyses help unravel the mystery of the inconsistencies between SNS-seq and other techniques used to map DNA replication origins genome-wide.

Hierarchical processing and polarization encoding in the cephalopod visual system

Mano, T. — 2026-01-08

Coleoid cephalopods (octopus, cuttlefish and squid, hereafter cephalopods) have evolved a range of complex visually-guided behaviours, from dexterous hunting to skin-pattern based camouflage and communication1. They have also evolved sensitivity to the polarization of light2, an adaptation thought to help detect camouflaged or semitransparent predators and prey in low visibility underwater environments3-10. How visual information is processed by the cephalopod brain to support their behaviours remains unclear. Here, studying the bigfin reef squid Sepioteuthis lessoniana, we performed calcium imaging and electrophysiological recordings from populations of neurons in the large visual center of the cephalopod brain, the optic lobe (OL). We revealed that the retina-recipient superficial OL contains a diversity of functionally distinct cell types, spatially organized into sub-layers, processing spatio-temporal features of light intensity and possessing polarization angle specificity. More complex features, e.g. direction selectivity, are seen in deeper regions of the OL cortex, which also exhibits spontaneous waves of neural activity in the absence of visual input. Neurons in the downstream OL medulla exhibit visual receptive field sizes and spontaneous activity levels which increase with brain depth, consistent with the hierarchical processing of visual information through the medullas tree-like anatomical organization. Medulla neurons exhibit sensitivity to local decreases in the degree of linear polarization (DoLP), which they integrate additively with light intensity information. Underwater imaging in the squids habitat off the coast of Okinawa, Japan, demonstrate that polarization sensitivity confers a robust short-range boost in object-background contrast over a range of objects and environmental conditions. These findings reveal convergent principles of hierarchical visual processing shared between cephalopods and vertebrates, and highlight how cephalopods utilise their distinct adaptation of polarization sensitivity to solve universal visual challenges underwater.

Navigators learn a local graph, not a global map

Strickrodt, M. — 2026-01-12

It is still an unresolved issue how humans represent navigable space and use this information to relate distant locations, as when shortcuting or pointing. In the present study we compare two competing theoretical approaches to the structure of this spatial knowledge. We contrast the metric embedding of spatial information in a global reference frame (i.e., a Euclidean mental map) with models that assume only local place-to-place information (i.e., a labeled graph), which can be used to estimate shortcuts and pointing direction when needed. Two groups of participants learned a multi-corridor virtual maze by walking around a zig-zag loop that connected seven objects placed on a circle. One group experienced a possible Euclidean maze, and the other group an impossible, non-Euclidean, broken version of the maze. In the impossible environment, after walking one lap the participant was covertly teleported to the starting place again, despite having walked to a different Euclidean location. Thus, the local place-to-place metrics were globally inconsistent. During the test phase, participants pointed to targets in a clockwise or counterclockwise sequence around the circle from their current location. Whereas the possible maze group was fairly accurate, the estimates of the impossible maze group were systematically biased by the test sequence, as predicted by local place-to-place metrics. Despite being queried about the same target, participants pointed in significantly different directions, violating the metric postulates. The results suggest that human knowledge of navigable space is not a globally consistent Euclidean map, but can be characterized as a labeled graph.

Evolutionary Divergence of mTOR-mediated Transcriptional Regulation Between Drosophila melanogaster and Drosophila simulans is Modulated by Sex and Tissue

Raynes, Y. — 2026-01-13

Evolutionary divergence in gene regulation is a major source of phenotypic novelty, yet understanding how deeply conserved, pleiotropic signaling pathways evolve under selective constraint while maintaining essential cellular functions remains an important challenge. Here, we investigate the evolutionary divergence in transcriptional regulation mediated by the mechanistic target of rapamycin (mTOR) pathway in two related species, Drosophila melanogaster and Drosophila simulans. mTOR is a highly conserved central regulator of cellular growth and metabolism, making it well-suited for the study of regulatory evolution under functional constraint. Using a fully factorial RNA-seq design, we quantified transcriptional responses to mTOR inhibition by rapamycin across three tissues (head, thorax, and abdomen) and both sexes. Despite highly conserved tissue-specific expression - reflecting the close phylogenetic relationship between species - mTOR-mediated transcriptional responses showed clear evidence of evolutionary divergence. Divergence varied across tissues and sexes: heads of both sexes and female thoraces and abdomens showed the most rapid gene-level divergence, suggesting stronger directional selection, whereas male thorax and abdomen exhibited comparatively conserved responses, consistent with stabilizing selection. Gene-level divergence patterns were consistent across multiple metrics and generally mirrored pathway-level divergence, except in the female abdomen, which showed relative pathway-level conservation despite extensive gene-level divergence. Species-by-treatment interaction analyses further revealed divergence in core mTOR-regulated biological processes. Together, our results suggest that regulatory modularity may allow even highly conserved signaling pathways to evolve under context-specific selective pressures while maintaining critical functionality. Significance statementThe evolution of gene regulation is increasingly recognized as a major contributor to biological diversification. However, how highly pleiotropic and deeply conserved pathways evolve new regulatory effects without disrupting essential cellular functions remains an open question in evolutionary biology. Using the nutrient-sensing mTOR pathway as a model, we show that gene regulation mediated by this core, selectively-constrained signaling network has diverged between two closely related fruit fly species in a tissue- and sex-specific manner. Our findings suggest that even pleiotropic pathways like mTOR can diverge over short evolutionary timescales, and that regulatory modularity may facilitate the evolution of novel transcriptional effects in specific contexts while preserving essential functions.

Nutrient response strategies drive coastal range shifts of phytoplankton taxa

Krinos, A. I. — 2026-01-14

Coastal phytoplankton blooms are important drivers of regional and global marine primary production. Recently documented increases in coastal phytoplankton blooms in the first quarter of the 21st century may be a consequence in part of changing environmental conditions and have important ecological implications. Here, we explore coastal phytoplankton dynamics in Cape Cod Bay, MA, USA. We use a 20-year phytoplankton ecology dataset to identify potential drivers of the increasing prevalence of two regionally-rare phytoplankton taxa: a coccolithophore thought to thrive in the global open ocean, and a dinoflagellate genus with potentially toxic members. Using metatranscriptomics, we show that these minor phytoplankton taxa leverage unique strategies to gain a competitive advantage under nutrient limitation compared to traditionally dominant taxa and compared to a diatom taxon that became modestly more abundant over the study period. Our results highlight the ecological dynamics arising from long-term shifts in temperature and nutrient status in coastal ecosystems.

MAPKAP Kinase 2 Orchestrates Memory T Cell Inflation in Cytomegalovirus Infection

Panagioti, E. — 2026-01-20

Memory T cell inflation is a distinctive immunological phenomenon observed during persistent viral infections, such as Cytomegalovirus (CMV). Unlike conventional memory T cell responses, which contract after infection resolution, a subset of CMV-specific T cells undergoes a progressive and sustained expansion, termed "inflation", which is thought to be critical for long-term immune surveillance. The molecular mechanisms that govern memory T cell inflation remain incompletely understood, yet they are pivotal for understanding immune persistence and designing strategies against chronic viral infections. In this study, we investigate the role of MAP kinase-activated protein kinase 2 (MK2), a key downstream effector of p38 MAPK signaling, in regulating T cell responses during murine CMV (MCMV) infection. Using MK2 knockout (MK2-KO) mice, we demonstrate that MK2 deficiency alters the dynamics of MCMV-specific CD8 T cell responses without impairing viral control or tissue replication. MK2 deficiency led to a reduction in non-inflationary MCMV-specific CD8 T cells during the acute phase, followed by enhanced expansion of inflationary CD8 T cell subsets during persistence. Furthermore, MK2-KO mice exhibited impaired effector differentiation, as evidenced by decreased expression of the terminal differentiation marker KLRG1 on MCMV-specific CD8 T cells. Collectively, these findings identify MK2 as a pivotal regulator of CD8 T cell magnitude, kinetics, and phenotype during both acute and chronic MCMV infection. By elucidating the role of MK2 in the regulation of memory T cell inflation, this study provides new mechanistic insight into immune regulation with implications for vaccination, chronic infection, and immune aging. Graphical abstractThe graphical abstract was created using BioRender (https://biorender.com/). O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/699974v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@140d602org.highwire.dtl.DTLVardef@1a23691org.highwire.dtl.DTLVardef@ac75b6org.highwire.dtl.DTLVardef@640d68_HPS_FORMAT_FIGEXP M_FIG C_FIG

Exploring the GM-CSF Histidine Triad as a Modulator of Structure, Molecular Motion, and Ligand Binding

Cui, J. Y. — 2026-01-21

Granulocyte macrophage-colony stimulating factor (GM-CSF) is a cytokine that plays a role in immune modulation. Its expression is associated with a multitude of different effects ranging from harmful, as in diseases such as rheumatoid arthritis and multiple sclerosis, to beneficial, as in the case of mitigation of diabetes type I and neutropenia. However, there is a large gap in knowledge explaining how GM-CSF toggles its structure for such physiological and pathological interactions. Our work describes an ongoing attempt to address this gap by focusing on a clustered histidine triad within -helices near the N-terminus, which prior studies have suggested play a role in binding ligands at an acidic pH. While GM-CSF is known to be highly flexible at a more acidic pH, several properties of its histidine triad remain unclear at the physiological pH at which GM-CSF would encounter its binding partners. We describe an effort to characterize the role of the GM-CSF histidines under physiological pH, specifically to determine if these histidines are key to GM-CSF structural integrity, and whether individual histidine residues modulate binding as they do at a lower pH. Our findings reveal that, while the histidine residues have an impact on GM-CSF structure, flexibility, and stability, they alone do not modulate the affinity for ligands at neutral pH. These data provide an initial explanation for the pleiotropic functions and interactions of GM-CSF within a biophysical context. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/700583v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@a6fffcorg.highwire.dtl.DTLVardef@1f00c30org.highwire.dtl.DTLVardef@b04c50org.highwire.dtl.DTLVardef@6224d9_HPS_FORMAT_FIGEXP M_FIG C_FIG

The glycine-arginine-rich motif of 53BP1 modulates RNA interactions necessary for its liquid-liquid phase separation during DNA Damage Response

Terraneo, F. — 2026-01-30

The DNA damage response relies on the rapid assembly of repair factors into foci with properties of liquid-liquid phase separation, driven by de novo transcription of damage-induced RNAs. 53BP1 is a key component of these condensates, yet the molecular determinants driving this process remain unknown. Here, through computational, structural and in vitro approaches, we identify the oligomerization domain of 53BP1 and its glycine-arginine-rich (GAR) motif as crucial for RNA interactions and phase separation. Biophysical characterization reveals that 53BP1-RNA condensates can progressively mature into a more stable state, and that GAR mutants display aberrant material properties. Using a cellular model of telomere fusion events, we demonstrate that the GAR motif is essential for 53BP1-mediated DNA repair, which depends on the combined contributions of RNA binding and appropriate condensate biophysical properties. Therefore, RNA-driven 53BP1 condensation is functionally required to maintain genome integrity.

immgenT: A Comprehensive Reference of Convergent T-cell States in the Mouse

Magill, I. — 2026-02-02

The immgenT collaborative project generated a comprehensive molecular atlas of T cells spanning virtually all mouse organs and disease states, profiling [~]800,000 cells from 750 samples with RNA, 128-plex surface protein, and {beta}TCR sequence. Applying a deep generative model to joint RNA and protein data defined a finite landscape of T-cell states organized into eight lineages and 110 robust clusters, integrating identical cells from different contexts, and resolving prior nomenclatures. Analysis of effector molecules, transcription factors and modules showed that both immunological functions and regulatory programs are shared across cell states. This framework provides a stable, reusable reference, demonstrated by computationally integrating 16 external datasets from diverse biological contexts. A set of public web tools supports browsing of these data, allows mapping of any dataset onto the immgenT framework. These results propose a molecular classification of T cells organized around a set of shared states reused across immunological contexts.

The CD8 immgenT framework as a universal reference of mouse CD8 Tαβ cell differentiation states

Galletti, G. — 2026-02-04

Mouse CD8 T cell differentiation has been studied extensively in models of infections and cancer, yet no unified framework spans the full spectrum of immunological contexts. We present the CD8 immgenT framework, integrating >200,000 single-cell transcriptomes and 128-plex surface proteomes from 734 samples spanning multiple perturbations, tissues, and timepoints. Unbiased analysis identifies 21 states encompassing naive, effector, circulating memory, tissue-resident memory, progenitor-exhausted, and terminally-exhausted compartments, among others. These states re-emerge with striking molecular convergence across acute/chronic infections, cancer, autoimmunity, aging, and homeostasis, showing that near-identical transcriptional programs support protective or dysfunctional outcomes depending on developmental history and microenvironment. Classic archetypes map to discrete clusters but exhibit unappreciated heterogeneity and overlap, cautioning against rigid nomenclature. We provide validated combinatorial markers, flow cytometry gating strategies, and immgenT reference-based integration for reproducible annotation of new datasets. This universal coordinate system harmonizes fragmented CD8 T cell literature and clarifies relationships across diverse immune challenges.

Emergence of microglia structural and functional heterogeneity between hippocampal subregions during development into early adulthood

Salter, E. W. — 2026-02-05

Phagocytosis is a key function performed by microglia to maintain tissue homeostasis. The degree of microglial phagocytic activity differs across ages and between gross anatomical brain regions, dictated by the local environment. Here, we asked whether microglial phagocytic phenotype exhibits subregion-scale tuning by circuit-specific features within a brain region. To address this, we took advantage of the highly stereotyped architecture of the hippocampus. We examined three adjacent synaptic subregions, the CA1 stratum radiatum (SR), stratum lacunosum moleculare (SLM) and dentate gyrus molecular layer (DGML). These three subregions provide an ideal system for examining local microglia heterogeneity, as each subregion contains distinct neuropil features, creating three adjacent unique micro-environments to which the microglia are exposed. We measured the phagocytic activity and morphological properties of over 1,000 individual microglia at two developmental points, mid-postnatal (P16) and early adulthood (P60) in the CA1 SR, SLM and DGML. We found that microglial phenotype diversified with development into early adulthood. At the mid-postnatal age, phagocytic activity and morphology were homogeneous across subregions. Conversely, in young adulthood, microglia in the CA1 SR and DGML exhibited a reduction in phagocytic activity, while microglia in the CA1 SLM maintained a highly phagocytic phenotype reminiscent of an immature-like state. These findings uncover a fine-scale tuning of microglia activity that emerges during maturation and is dictated at the sub-region level of the hippocampus, uncovering a distinct population of microglia in the CA1 SLM that exhibit a persistent immature phenotype under physiological conditions. Understanding the target(s) of this phagocytosis and consequences for CA1 SLM function will provide new insight into the role of local tuning of microglia properties for circuit-specific needs in both health and disease.

A transcriptional code controlling fluid shear stress-induced gene expression

Fleisinger, L. — 2026-02-06

The formation and health of the vascular system is dependent on fluid shear stress (FSS), a hemodynamic force exerted onto endothelium by flowing blood. FSS strongly induces the endothelial expression of Kruppel-like factor 2 (KLF2), an atheroprotective TF essential for vascular development and homeostasis. Despite its early and crucial role in the cascade of cardiovascular events triggered by FSS, the transcriptional mechanisms by which FSS regulates KLF2 expression remain unclear, although they are known to involve the widely expressed MEF2 proteins. Here, we identified and characterized two FSS-dependent enhancers for KLF2 which collectively recapitulate endogenous endothelial KLF2 expression, and determined the TFs contributing to their regulation. This analysis identified an essential and precisely spaced MEF2-TBP double motif also shared by the FSS-sensitive KLF2 promoter. MEF-TBP double motifs are extremely rare across the genome but were also found within regulatory elements of three other FSS-induced KLF genes, including KLF4. Although normally part of the basal transcriptional machinery, TBP specifically bound all KLF elements at the MEF-TBP double motifs in a FSS-dependent manner. Collectively, this work demonstrates a specific and targetable requirement for combined MEF2-TBP binding during FSS-induced gene activation. Significance statementBlood flow induces a force known as fluid shear stress (FSS) which is required for vascular development and for the health of the mature arterial system. One of the first endothelial responses to FSS is the induction of Kruppel-like transcription factors (KLFs). However, the mechanisms by which FSS activates KLF gene expression are incompletely understood. In this paper, we characterized all regulatory elements involved in driving FSS-induced expression of KLF2. This identified an essential MEF2-TBP double motif that was extremely rare across the genome, yet found within regulatory elements for multiple FSS-responsive KLF genes including KLF2 and KLF4. This MEF2- and TBP-bound motif therefore enables blood flow to specifically activate the cascade of cardiovascular responses necessary for atheroprotective gene expression.

EBV Type 1 versus Type 2: A determinant of NK cell anti-tumor activity in Burkitt lymphoma

Forconi, C. S. — 2026-02-10

Terminally differentiated CD56negCD16pos NK cells have been described after chronic viral and malaria infections, and in children diagnosed with Burkitt lymphoma (BL). Despite CD56neg NK cells appearing to be poor at direct cytotoxicity, they express high levels of cytotoxic granules (i.e. granzymes, perforin), activation markers, and Fc-{gamma} receptors (CD32 and CD16) that are typically engaged in antibody-dependent cell cytotoxicity (ADCC). In addition, the abundance of CD56neg NK cells strongly correlates with IgG1 and IgG3 plasma levels, which are essential subclasses for ADCC. To determine whether CD56neg NK cells have superior ADCC capacity relative to CD56dim NK cells, we performed ADCC assays using effector cells from pediatric cancer patients and healthy children from malaria endemic regions of Kenya, targeting in vitro rituximab-treated commercial and newly established BL cell lines. We found that CD56neg NK cells were indeed capable of in vitro ADCC, showing a significant increase of CD107a-mediated degranulation in the presence of rituximab; however, they were not as efficient as CD56dim NK cells. Moreover, we found that the ADCC magnitude was significantly lower against EBV-Type 2 (EBV-T2) BL lines compared to EBV-Type 1 (EBV-T1). EBV-T2 tumor cell lines expressed significantly more lytic viral proteins than EBV-T1, making them more sensitive to direct cytotoxicity. Results from this study highlight the importance of assessing inter-patient variation in NK cell profiles in conjunction with ADCC sensitivity and EBV type within tumor cells when evaluating clinical outcomes for NK-mediated immunotherapies. SignificanceEBV type dictates NK cytotoxicity: EBV-T1 BL cells require rituximab for NK killing, while EBV-T2 BL cells are eliminated without antibody assistance, highlighting target-specific immune response to EBV-associated cancers.

BRIDGE: Biological Antimicrobial Resistance Inference viaDomain-Knowledge Graph Embeddings

Iyer, A. — 2026-02-11

Antimicrobial resistance (AMR) is a growing global health crisis, responsible for an estimated 1.27 million deaths in 2019 alone. Traditional approaches to identifying antibiotic resistance genes (ARGs) are often labour-intensive and limited in their ability to detect novel resistance mechanisms. In this study, we propose BRIDGE, a knowledge graph-based framework, to improve AMR gene prediction by integrating gene neighbourhood information and protein-protein interaction networks. Focusing on Klebsiella pneumoniae and Escherichia coli, we construct a comprehensive and biologically grounded knowledge graph using curated data from CARD, STRING, and DrugBank. We apply knowledge graph embedding models which are fed into deep neural networks to infer novel AMR links, achieving classification accuracy of up to 97%. Our results demonstrate that incorporating biologically meaningful relationships, such as gene neighbourhood information and protein interactions, enhances the predictive accuracy and interpretability of AMR link predictions. This work contributes to the development of scalable and data-integrated approaches for advancing antimicrobial resistance surveillance and drug discovery. BRIDGE implementation and data are available at https://github.com/GraphML-lab/BRIDGE.

Breaking β-sheets in FUS prion-like domain preserves phase separation and function but prevents aggregation and toxicity

Wake, N. — 2026-02-18

The RNA-binding protein Fused in Sarcoma (FUS) undergoes phase separation associated with RNA processing. However, the prion-like low complexity (LC) domain of FUS forms solid-like aggregates in neurodegenerative diseases. Whether the formation of {beta}-sheet structure associated with pathology is also physiologically/functionally relevant is debated. Similarly, if mislocalization alone or concomitant aggregation is responsible for FUS gain-of-function toxicity remains to be probed. Here, we introduce {beta}-sheet breaking proline residues into FUS LC with the goal of preventing cross-{beta}-driven aggregation without disrupting essential functions and phase separation. {beta}-sheet-deficient FUS variants maintain native-like global motions, disorder, and phase separation, but no longer show a liquid-to-solid transition (LST). Biochemical partitioning, cellular localization, and auto- and cross-regulatory functions of FUS all remain essentially unchanged. Conversely, FUS-induced neurodegeneration in several Drosophila models is drastically reduced. These findings suggest a strategy for mitigating disease-related toxicity through backbone structure modulation to prevent prion-like domain protein aggregation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/706410v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@d15f63org.highwire.dtl.DTLVardef@1cd6221org.highwire.dtl.DTLVardef@e58126org.highwire.dtl.DTLVardef@181ec67_HPS_FORMAT_FIGEXP M_FIG C_FIG SUMMARYThe RNA-binding protein Fused in Sarcoma (FUS) undergoes phase separation as part of its physiological function but can aberrantly aggregate into solid-like assemblies in amyotrophic lateral sclerosis and frontotemporal dementia. To dissect the role of {beta}-sheets in both function and pathological transition, we engineered {beta}-sheet-preventing FUS variants via targeted proline residue insertions in the prion-like disordered region. These variants retained native structure, motions, and phase behavior yet showed dramatically reduced aggregation, both as an isolated prion-like domain and in full-length FUS. Crucially, these variants maintained a panel of FUS cellular functions that depend on FUS condensation but prevented FUS toxicity in fly models of neurodegeneration. Our findings implicate {beta}-sheets as key drivers of FUS condensate maturation and neuronal toxicity, highlighting {beta}-sheet modulation as a therapeutic strategy against FUS-related neurodegeneration. HIGHLIGHTSO_LITargeted proline additions disrupt {beta}-sheet formation in FUS without altering native conformations, dynamics, or phase separation behavior C_LIO_LI{beta}-sheet-deficient FUS variants prevent aggregation and liquid-to-solid transitions while retaining key biological functions C_LIO_LIIn vivo models reveal attenuated toxicity of {beta}-sheet-deficient FUS in Drosophila C_LIO_LI{beta}-sheets are identified as central drivers of condensate maturation and neuronal death, offering a therapeutic entry point for modulating prion-like domain pathology C_LI

Cacna1b alternative splicing is linked to associative learning

Dhillon, S. K. — 2026-02-18

Voltage-gated CaV2.2 channels are essential for neurotransmitter release throughout the nervous system including areas related to learning and memory like the hippocampus. Previous results have shown that CaV2.2 channels are involved in cognitive processes. However, a link between alternative splicing of the Cacna1b (gene that encodes for CaV2.2) pre-mRNA and cognitive processes has not been described. The Cacna1b pre-mRNA undergoes extensive cell-specific alternative splicing. In this body of work, we focus on the cassette exon 18a. Alternative splicing of exon 18a generates two splice variants, +18a-Cacna1b and {Delta}18a-Cacna1b. Exon 18a encodes a 21-amino acid sequence within the SYNaptic PRotein INTeraction (synprint) site. Splice variants containing exon 18a (+18a-CaV2.2) show reduced cumulative inactivation and increased Ca{superscript 2} current density compared to splice variants lacking exon 18a ({Delta}18a-CaV2.2), suggesting functional specialization. We previously showed that +18a-Cacna1b splice variants are enriched in cholecystokinin-expressing interneurons (CCK+INs). This neuronal type is strongly implicated in associative learning. Therefore, we tested whether alternative splicing of exon 18a contributes to associative learning. To test this hypothesis, we used genetically engineered mice that constitutively express either +18a-Cacna1b (+18a) or {Delta}18a-Cacna1b ({Delta}18a). We first validated that restricted splicing of exon 18a did not alter downstream alternative or constitutive spliced exons in the Cacna1b pre-mRNA, nor total CaV2.2 protein levels. We then performed a comprehensive behavioral analysis that included assessment of associate learning. We found that in the trace fear conditioning task, +18a mice exhibited less freezing during the trace interval in both the acquisition and memory phases compared to WT mice. Whereas {Delta}18a mice showed enhanced freezing during the same intervals relative to WT mice. These bidirectional phenotypes reveal that exon 18a shapes aversive associative learning. Furthermore, exon 18a splicing did not influence spatial working memory, spatial navigation under stress, nociceptive responses in basal and inflammatory conditions, overall locomotion or exploratory behavior. These results suggest that the behavioral impact of exon 18a splicing is highly selective. Together, our findings identify alternative splicing of exon 18a as a molecular contributor to associative learning.

A modular, high-bandwidth, bidirectional implantable device for neural interrogation

Darie, R. — 2026-02-24

Modern neuroelectronic interfaces have shown great potential to diagnose conditions, address neurological dysfunction, and advance neuroscientific knowledge. However, neural interface systems today require tethered connections that restrict mobility, prevent testing across ecological contexts, and inhibit clinical translation to at-home use. Fully implantable commercial systems have previously been developed, but exhibit significant constraints, including a bulky design, limited modularity, low bandwidth, or unidirectional communication (e.g. deep brain stimulation systems, DBS; spinal cord stimulation systems, SCS). Here, we have developed the Modular Bionic Interface (MBI), a system composed of a fully implantable device and a worn unit for high-bandwidth, bidirectional interfacing with the nervous system. The MBI can record high fidelity electrophysiological signals and deliver spatiotemporally modulated electrical stimulation for clinical and research purposes through flexible interaction with third party implantable devices. We performed benchtop evaluation to validate the recording and stimulation capabilities of the MBI across a diverse range of inputs and outputs. We then evaluated the MBI system in vivo through chronic implantation within a sheep, where results were stable for the length of evaluation, over three months. While connected to an actively powered, third-party high-resolution spinal cord stimulation electrode array, the MBI system was able to deliver stimulation to evoke lower extremity motor responses and record spinal compound action potentials evoked by peripheral nerve and spinal stimulation. Through rigorous evaluation, we demonstrate a fully implantable system with a small footprint capable of high-resolution, bi-directional communication with the nervous system via modular connections to third-party devices. We expect that modular devices will further our ability to treat complex neurological disease and injury.

Rapid sequence-based screening of structure-disrupting protein mutations

Oh, J. — 2026-02-25

Recent advances in AI-based protein structure prediction have dramatically reduced the cost of obtaining three-dimensional protein models and have become integral to modern protein engineering workflows. However, full structure prediction remains computationally prohibitive in high-throughput settings for mutation-based protein engineering, where thousands of candidate variants may need to be evaluated. In many such scenarios, the primary objective is not to resolve the complete structure of a candidate mutant, but rather to identify whether the introduced mutations are likely to induce substantial structural changes for rapid down-selection of candidates that conserve the wildtype structure. Protein language models (PLMs), trained solely on unlabeled natural protein sequence data, are known to encode rich structural information within their hidden representations. Motivated by this observation, we investigate a range of sequence-based ranking metrics derived from PLMs as efficient surrogates for structural deformation prediction. Through systematic evaluation across multiple proteins, mutation regimes, and structure-prediction backbones, we show that embedding distance--particularly the L1 distance between representations--provides a robust and computationally efficient signal for identifying structure-disrupting mutations. Our results demonstrate that sequence-level screening can substantially reduce the need for expensive structure prediction while preserving sensitivity to large structural perturbations, thereby providing the means to significantly speed up mutation-based protein design.

The Translatome of Senescent Cells Revealed by Ribosome Profiling

Kalekar, R. L. — 2026-03-01

Cellular senescence drives aging-related tissue dysfunction in part through the senescence-associated secretory phenotype (SASP), an inflammatory secretome linked to retrotransposable element (RTE) derepression. Transcriptomic and proteomic approaches have characterized the senescent program extensively, but mRNA abundance does not predict protein output well, and limited proteomic depth constrains the detection of low-abundance SASP factors and RTE-derived proteins. To bridge this gap, we used AHARibo, a metabolic labeling-based method that selectively enriches mRNAs associated with actively elongating ribosomes, to generate translatome profiles in human fibroblasts across proliferating, early senescent, and late senescent states. Comparison of total and ribosome-associated mRNA pools reveals marked translational uncoupling in early senescence: transcriptomic changes explain only 34% of translatomic variance, compared to 70% in late senescence, indicating that early senescence is substantially shaped by post-transcriptional regulation. Key senescence programs are actively regulated at the translational level: cell cycle and extracellular matrix remodeling genes are translationally suppressed and enhanced, respectively, while inflammatory SASP components are translationally depleted in early senescence - a depletion relieved in late senescence. Translationally depleted SASP genes are enriched for binding motifs of the ZFP36 family (ZFP36, ZFP36L1, ZFP36L2), implicating these RNA-binding proteins in the post-transcriptional gating of inflammatory signaling. More broadly, translational efficiency is associated with 3UTR GC content and codon optimality, and translationally depleted mRNAs are enriched for numerous RBP and microRNA target motifs. Finally, we detect robust, locus-resolved translation of evolutionarily young LINE-1 retrotransposons, identifying full-length elements with stage-specific translational activity. Together, these findings establish translational control as a pervasive regulatory layer shaping the senescent phenotype.

Single-molecule spatial genomics reveals the multi-scale organization and plasticity of extrachromosomal DNA in glioblastoma

Taylor, B. — 2026-03-07

Extrachromosomal DNA (ecDNA) is a major driver of intratumoral heterogeneity and is associated with poor clinical outcomes across cancers, yet how individual ecDNA molecules are organized and regulated within intact tumors remains unknown. Here, we leveraged single-molecule, multi-modal spatial genomics to resolve the three-dimensional chromatin organization and transcriptional activity of individual EGFR-containing ecDNA molecules in glioblastoma (GBM) cells in vitro, in orthotopic xenografts, and in patient-derived GBM tissue. At the larger scale, we find that distinct GBM molecular and functional states emerge depending on the local cellular environment. EGFR expression was markedly different between GBM subpopulations, and perturbations of EGFR dosage shifted GBM cellular states. ecDNA expression was modulated by multiple mechanisms, including variation in copy number, chromatin organization, DNA sequence, and chromosomal reintegration, which were simultaneously measured within the same cells. At the single-molecule scale, ecDNA adopts a physically expanded chromatin configuration with larger ecDNA molecules having higher transcriptional activity and interaction with active transcriptional machinery. ecDNA regulation was coordinated within cells and across GBM states, and ecDNA copy number, structure, and transcription were spatially organized across the tumor architecture. Co-culturing GBM cells with neurons recapitulated key features of infiltrative regions, including lower EGFR expression, reduced ecDNA copy number, and increased chromosomal reintegration, suggesting a causal role for the microenvironment in shaping ecDNA regulation. Collectively, these findings support a model in which GBM states and ecDNA are linked, plastic, and influenced by microenvironmental contexts, revealing a previously inaccessible layer of genome organization underlying tumor heterogeneity and malignant cell behavior.

A minimal transcriptomic signature predicts intravascular tumor extension in renal cell carcinoma

Mao, C. A. — 2026-03-09

Renal cell carcinoma (RCC) with venous tumor thrombus, termed renal intravascular tumor extension (RITE), is associated with aggressive behavior and poor clinical outcomes. Yet, its underlying molecular determinants remain incompletely defined. We analyzed RNA sequencing data from three independent RCC cohorts comprising 721 samples. Two cohorts included matched samples of index tumor, tumor thrombus, and normal adjacent kidney tissue. Analyses integrated dimensionality reduction, differential gene expression, interpretable machine learning, and gene ontology approaches. Principal component analysis revealed that only these two cohorts exhibited a coherent RITE-associated transcriptional structure. Their sequencing depth was sufficient to delineate 6,317 differentially expressed genes that distinguish RITE from non-RITE tumors. SHAP-based feature attribution across logistic regression, random forest, and XGBoost yielded a robust 29-gene consensus signature, which was further distilled into a compact 13-gene panel that preserved maximal classification performance. These genes converged on biological themes, including loss of distal epithelial identity, dysregulation of ion transport pathways, and consistent enrichment of mitochondrial processes such as oxidative phosphorylation. Together, these findings define a newly discovered and uniquely refined molecular signature of venous tumor extension in RCC and highlight mechanistically relevant pathways that may inform biomarker development and future translational strategies for predicting or mitigating RITE progression.

Systematic clustering alignment and feature characterization for single-cell omics using ACE-OF-Clust

Liu, X. — 2026-03-12

Clustering is widely used to identify cell types in cellular-resolution transcriptomic data, including single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics (ST). Mixed-membership clustering assigns fractional memberships across clusters and captures continuous variation beyond hard clustering, but integrating and interpreting results from either approach is complicated by the "clustering alignment problem," which arises from label switching, multi-modality, and differences in model settings (including differing numbers of clusters). We introduce ACE-OF-Clust, enabling a four-step workflow for single-cell clustering: multiple clustering, clustering alignment, model comparison, and identification of informative features. ACE-OF-Clust introduces direct comparison of clustering solutions, assesses consistency against annotations, and leverages feature-level clustering profiles to prioritize genes discriminating among cell types. We demonstrate its utility on PBMC scRNA-seq and breast cancer ST data, and on multi-omic single-cell data. ACE-OF-Clust quantifies cross-omic clustering variability and suggests putative cross-omic regulatory links. Overall, ACE-OF-Clust increases the interpretability, flexibility, and robustness of single-cell clustering, providing a scalable tool for studying cellular heterogeneity and gene expression dynamics.

Improving Local Ancestry Inference through Neural Networks

Medina Tretmanis, J. — 2026-03-13

MotivationLocal Ancestry Inference (LAI) allows us to study evolutionary processes in admixed populations[1], uncover ancestry-specific disease risk factors[2], and to better understand the demographic history of these populations[3]. Many methods for LAI exist, however, these methods usually focus on cases of intercontinental admixture. In this work, we evaluate both existing and novel methods in challenging scenarios, such as downsampled reference panels, intracontinental admixture, and distant admixture events. ResultsWe present four novel LAI implementations based on neural network architectures, including Bidirectional Long Short-Term Memory and Transformer networks which have not previously been used for LAI. We compare these novel implementations to existing methods for LAI across a variety of scenarios using the 1 Thousand Genomes dataset and other synthetic datasets. We find that while all networks achieve high performance for intercontinental admixture scenarios, inference power is comparatively low for scenarios of intracontinental or distant admixture. We further show how our implementations achieve the best performance of all methods through specialized preprocessing and inference smoothing steps. AvailabilityAll implementations and benchmarking code available at https://github.com/Jazpy/LAINNs.

Antarctic marine microplastics reveals environmental persistence and rapid evolution of Candida auris

van Rhijn, N. — 2026-03-13

Candida (Candidozyma) auris is a critical priority fungal pathogen that emerged two decades ago near simultaneously on multiple continents. Since emergence, C. auris resistance to all four classes of antifungal drugs has been described, including pan-drug resistant isolates, sometimes evolving within patients. Here, we confirm the first isolation of C. auris from Antarctica and show cold-adapted phenotypes and an affinity for binding to nylon. We also provide evidence to suggest mutator phenotypes contribute to the rapid evolution of C. auris and are responsible for the emergence of multiple, distinct genetic clades worldwide. Isolates in clades I, III and IV with a mutator phenotype displayed elevated mutation rates compared to non-auris Candida species. This phenotype had a complex genetic basis and was associated with drug resistance mutations. We postulate that the mutator phenotype has a significant effect on evolutionary potential and is responsible for the emergence and rapid spread of drug-resistance C. auris and novel genetic clades.

Cell type-specific gene regulatory network inference from single cell transcriptomics with ctOTVelo

Chang, S. — 2026-03-14

Inferring gene regulatory networks (GRNs) from gene expression is a crucial task for understanding functional relationships. Gene expression data (transcriptomics) provide a snapshot of gene activity, encoding information about gene regulatory relationships. However, gene regulation is a dynamic process, modulating across time and with different cell types. Temporal GRN inference methods aim to capture these dynamics by utilizing time-stamped transcriptomics, gene expression data of similar samples captured across discrete timepoints, or pseudotime transcriptomics, computationally ordering cells based on an inferred trajectory. These methods can estimate constant or temporal gene regulatory relationships, but may not capture finer, cell type specific relationships. We propose ctOTVelo, an extension to our previous work to account for cell type specificity during GRN inference. ctOTVelo incorporates cell type labels or proportions when inferring the GRN from single cell transcriptomics data. Our methods achieve state-of-the-art performance in GRN prediction in time-stamped and pseudotime-stamped transcriptomics. Furthermore, ctOTVelo is able to generate cell type specific GRNs, allowing cell type resolution analysis of gene regulatory relationships.

Differential chromatin looping regulated by two GA-binding transcription factors creates an X-specific chromatin environment for dosage compensation

Aguilera, J. L. — 2026-03-14

The mechanisms by which differential occupancy of transcription factors (TFs) at similar binding sites leads to context-specific targeting of large transcription complexes remain poorly understood. X chromosome upregulation (XCU), the most highly conserved step in dosage compensation and best studied in Drosophila, serves as a model for understanding how differential occupancy of similar TFs functions context-specifically. Sequence variation within GA-repeat motifs that accumulated on the X chromosome over evolutionary time promotes the binding of a specific GA-binding TF (CLAMP) that recruits the dosage compensation complex (DCC) while outcompeting another similar TF (GAF). However, the mechanism by which CLAMP-GAF competition drives specific targeting of the DCC to the X chromosome remains unknown. Because DCC binding sites cluster in 3D space, we combined Micro-C and Hi-ChIP to determine that CLAMP and GAF directly mediate largely mutually exclusive 3D genomic contacts. Specifically, we show that CLAMP but not GAF drives local short-range interactions that directly link high affinity DCC binding sites with active, dosage-compensated housekeeping genes. In contrast, GAF mediates interactions between transcriptionally silent insulator regions on the X chromosome spanning a wider range of genomic distances. Together, these findings demonstrate that CLAMP outcompetes GAF at active regions on the X chromosome, but not autosomes, to create an X-chromosome specific chromatin environment for dosage compensation. Overall, we provide new insight into how differential TF binding at similar binding sites drives context-specific targeting of transcription complexes.

Selective regulation of transsynaptic alignment and postsynaptic assembly by a novel NCAM family synaptic adhesion molecule

Van der Linden Costello, P. — 2026-03-16

Synapse formation underlies the organization of neurons into functional circuits during brain development and requires precise alignment and maturation of pre-and postsynaptic compartments. Many synaptogenic adhesion molecules have been identified that drive target recognition and sustained adhesion between appropriate synaptic partners. Yet the degree to which individual molecules serve selective functions in distinct aspects of synapse formation and maturation remains poorly understood. In particular, how exquisite nanoalignment of pre-and postsynaptic specializations flows from micron-scale adhesive interactions between synaptic partners remains a key unanswered question. Here we shed new light on this question by establishing a specialized set of synaptic functions for Epithelial limiter of Fasciclin II function (Elff), an NCAM family member with previously unknown roles in the nervous system. Our structural and functional studies at the glutamatergic Drosophila NMJ indicate that Elff is required for postsynaptic assembly, maturation, and transsynaptic alignment; however, it is not required for presynaptic function, bouton formation, or developmental expansion of the NMJ. Notably, NMJs in elff null mutants display reduced glutamate receptor clustering beginning at the embryonic stage when NMJ synapses first form. These poorly defined postsynaptic specializations are frequently out of register with presynaptic release sites, disrupting neurotransmission. Unexpectedly, the striking defects in elff nulls occur in the context of both normal active zone number and developmental expansion of the NMJ. These findings suggest a surprising degree of specialization among transsynaptic adhesion complexes and demonstrate that Elff-mediated signaling is critical for the transsynaptic nanoarchitecture of glutamatergic synapses.

A novel framework for expanding RNNs with biophysical detail to solve cognitive tasks

Tolley, N. — 2026-03-17

Recurrent neural networks (RNNs) have proven to be highly successful in emulating human-like cognitive functions such as working memory. In recent years, RNNs are evolving to incorporate more biophysical realism to produce more plausible predictions on how cognitive tasks are solved in real neural circuits. However, there are major challenges in constructing and training networks with the complex and nonlinear properties of real neurons. A major component of the success of RNNs is that they share the same mathematical base as deep neural networks, permitting highly efficient optimization of model parameters using standard deep learning techniques. To do so, they use abstract representations of neurons which fail to capture the impact of cell-level biophysical and morphologic properties that may benefit network-level function. Expanding task-trained RNNs with biophysical properties such as dendrites and active ionic currents poses substantial challenges, as it moves these models away from the validated training regimes known to be highly effective for RNNs. To address this gap, we developed a biophysically detailed reservoir computing (BRC) framework with the goal of extracting mechanistic insights from biophysical neural models, and propose that these insights can be used to guide model choices that will work for specific categories of cognitive tasks. The BRC network was constructed with synaptically coupled excitatory and inhibitory cells, in which the excitatory cells include multicompartment biophysically active dendrites; motivated by empirical studies suggesting dendrites have desirable computational benefits (e.g. pattern classification and coincidence detection). We trained the BRC network to do a simplified working memory task where it had to maintain the representation of an extrinsic "cue" input. We studied the impact of extrinsic input time constants (fast AMPA vs slow NMDA) and location (dendrite vs soma) on the ability of a network to solve the task. Our results revealed that cue inputs through NMDA receptors are particularly efficient for solving the working memory task. Further, the properties of NMDA receptors are uniquely suited for cue inputs delivered at the dendrite, as networks trained with dendritic AMPA cue inputs failed to solve the task. Detailed examination of the cell and network dynamics that solve the task reveals distinct local network configurations and computing principles for the different types of extrinsic input. Overall, much like the body of mechanistic insights that have underpinned the success of training RNNs, this study lays the groundwork for applying the BRC framework to train biophysically detailed neural models to solve complex human-like cognitive tasks.

POLQ-driven repair scars shape the immunogenic landscape of homologous recombination-deficient pancreatic cancer

Park, W. — 2026-03-17

Pancreatic cancer (PC) is broadly resistant to immune checkpoint blockade, although a subset of homologous recombination-deficient (HRD) tumors exhibits durable immune engagement. The genomic features that distinguish these immune-responsive tumors from immune-inert HRD tumors remain poorly understood. Here we identify a microhomology-mediated end joining (MMEJ) repair scar, the MMEJ Deletion Footprint (MDF), as a genomic readout of POLQ-associated error-prone repair that enriches for frameshift indels. Across the multi-omic discovery cohort integrating tumor genomics, single-nucleus transcriptomics and spatial immune profiling, MDF-high HRD PC exhibited increased frameshift-indel-derived neoantigens and interferon programs. MDF was further associated with remodeling of the myeloid compartment toward MHC II-high dendritic cell-like antigen-presenting macrophage states and the immune synapse architecture marked by increased spatial interaction between APC-like macrophages and cytotoxic CD8+ T cells. These tissue-level features aligned with a functional trajectory shift of CD8+ T cells, consistent with effective anti-tumor immunity and was associated with favorable clinical outcomes of patients. Together, our findings position MMEJ-linked repair scarring as actionable biology that connects an HRD genotype to immune organization and suggests rational immunotherapy combinations that may enhance antigen presentation and myeloid activation to extend durable benefit in HRD-lineage cancers.

WINDEX: A hierarchical integration of site- and window-based statistics for characterizing the footprint of positive selection in genome-wide population genetic data

Snell, H. — 2026-03-26

Adaptive mutations, or mutations that confer a fitness benefit, can leave behind distinct signals in genetic data. Computational methods have improved the localization of adaptive mutations in genetic samples using a range of statistical and machine learning classification techniques. However, these methods miss the opportunity to jointly integrate statistics at both the site and window-based level, thus failing to harness all available statistical evidence to detect selection. Our method, WINDEX, combines these different resolutions of statistics to improve the detection of adaptive mutations among hitchhiking signals. Our model simultaneously integrates emissions at different resolutions by defining site-based and window-based latent states corresponding to neutral, linked, and sweep regions, with the site-based states and transition models nested within the window-based states. Using evolutionary simulations with varying selection parameters, we validate the ability of WINDEX to classify positive selective sweeps. Using data from the 1000 Genomes Project, we show that WINDEX is able to identify regions harboring signals of selective sweeps, and provides improved localization within those regions over existing methods. In addition, using WINDEX genome-wide allows for estimation of the proportion of whole genomes that are under positive selective pressures; our estimates of between 9.7-10.5% across different populations provide support for other preliminary estimates of these quantities. Author summaryPopulation geneticists often seek evidence for positive selective sweeps, or an evolutionary event in which a beneficial allele increases in frequency over time in a population, resulting in increased fitness of the individuals that have said allele. Positive selective sweeps, however, are difficult to detect due to varying patterns of linkage disequilibrium (LD), or the nonrandom association of alleles, and detecting these signals reliably among differing LD structures remains a challenge in the field. In this work, we present WINDEX, a probabilistic framework designed to leverage signals of positive selective sweeps at both the site- and window-levels in the form of a hierarchical hidden Markov model (HHMM), to localize regions of positive selective sweeps in aligned haplotype data. We validate WINDEX in evolutionary simulations over varying positive selective sweep scenarios, showcasing the improved resolution that the HHMM structure provides. We apply WINDEX in comparative genomic scans of canonical sites of positive selection as well as whole-genome scans to demonstrate the tools power in localizing functionally-validated signals of selection and to offer insights into the proportion of the human genome currently under positive selective pressures. WINDEX is publicly available and easy to apply to many cases of human genetic data.

A network for self-transcendence derived from patients with brain lesions

Healey, M. R. — 2026-03-27

Self-transcendence, the reorientation of experience away from the self toward others, nature, or broader meaning, is a fundamental dimension of human psychology, yet its causal neural architecture remains poorly understood. Here we applied lesion network mapping to 88 neurosurgical patients with pre- and post-operative assessments of trait self-transcendence to identify the distributed brain network whose disruption alters this capacity. The resulting network showed significant spatial correspondence with the default mode network and, at a finer parcellation level, with frontoparietal control subnetworks. Leave-one-out analyses identified posterior midline regions as the most stable correlates of increased self-transcendence following brain lesions. Independent validation against fMRI meta-analyses of self-referential processing, compassion, and ketamine administration, alongside a neuromodulation target previously shown to modulate the sense of self, converged on a consistent model. These findings provide causal evidence for a network architecture in which posterior midline hubs constrain, and brainstem and anterior midline regions facilitate, self-transcendent experience.

Sulfide:quinone oxidoreductase drives mitochondrial supersulfide metabolism to regulate bioenergetics and longevity in eukaryotes

Yao, J. — 2026-04-07

Sulfide:quinone oxidoreductase (SQR) is a critical enzyme that maintains sulfur metabolism by oxidizing sulfide to supersulfides, currently defined as sulfur metabolites with six valence electrons and no charge that are covalently catenated with other sulfur atoms and excludes disulfides. While SQR is known to contribute to mitochondrial electron transport, its physiological impact on systemic energy metabolism and longevity remains largely undefined. In this study, we investigated the role of SQR in mitochondrial bioenergetics and aging using SQR-deficient Schizosaccharomyces pombe ({Delta}hmt2) and a mitochondria-selective SQR-deficient (Sqrdl{Delta}N/{Delta}N) mice model. Functional analysis demonstrated that{Delta} hmt2 grew normally in glucose but not in glycerol, indicating impaired mitochondrial respiration. It showed reduced membrane potential, ATP, and lifespan. Consistent with the yeast findings, Sqrdl{Delta}N/{Delta}N mice exhibited accumulated levels of hydrogen sulfide and persulfides, and demonstrated impaired mitochondrial energy metabolism. Furthermore, supersulfide donor supplementation selectively conferred lifespan extension in wild-type yeast, but not in SQR-deficient strain, and similarly improved mitochondrial function exclusively in wild-type mouse embryonic fibroblasts, with no benefit observed in SQR-mutant counterparts. Together, our findings demonstrate that mitochondrial SQR plays an essential role in sulfur respiration, critically supporting mitochondrial function and organismal longevity across eukaryotes. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=175 SRC="FIGDIR/small/716515v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@cfada1org.highwire.dtl.DTLVardef@904e79org.highwire.dtl.DTLVardef@1024b1org.highwire.dtl.DTLVardef@1d911bc_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIDeveloped an SQR-deficient S. pombe ({Delta}hmt2) model that exhibits sulfur metabolism, mitochondrial dysfunction, and shortened chronological lifespan C_LIO_LISulfide and supersulfide donors prolong yeast lifespan in a SQR-dependent manner C_LIO_LIMitochondrial SQR is essential for membrane potential formation and ATP production in yeast and mammals C_LI