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<title>bioRxiv Subject Collection: Cell Biology</title>
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This feed contains articles for bioRxiv Subject Collection "Cell Biology"
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<title>bioRxiv</title>
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<link>https://www.biorxiv.org</link>
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<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.09.723948v1?rss=1">
<title>
<![CDATA[
Sex-differentiated hormonal microenvironments recapitulate in vivo liver metabolism in human iPSC-derived organoids 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.09.723948v1?rss=1
</link>
<description><![CDATA[
Bioengineers strive to recreate in vivo microenvironments in vitro to reduce our use of animal models and provide insights into human biology. While liver models show promise, sex differences in liver biology remain largely neglected in preclinical studies. Despite the 2014 EU mandate for the inclusion of women in clinical trials, decoupling of research data by sex is historically rare, with only 11% of papers disaggregating data by sex. This gap contributes to women being more susceptible to drug-induced liver injury (DILI) and being underserved in drug development, as well as to costly drug attrition levels. Here we present a novel approach to modelling sex differences in vitro. Human induced pluripotent stem cells (iPSCs) from both male (XY) and female (XX) donors, were differentiated into hepatocyte liver spheroids and exposed to in vivo-mimicking levels of testosterone, progesterone, and oestrogen in high-throughput microwell format. We successfully recapitulated sex-specific metabolic profiles and demonstrated significant differences in CYP1A2 and CYP3A4 drug metabolism and gene expression patterns consistent with reported in vivo observations, without compromising cell viability. These findings validate the utility of sex-differentiated microenvironments in early-stage research, offering a pathway to refine animal and clinical trials and improve therapeutic outcomes for all sexes.
]]></description>
<dc:creator><![CDATA[ Grant, R., Giselbrecht, S. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.09.723948</dc:identifier>
<dc:title><![CDATA[Sex-differentiated hormonal microenvironments recapitulate in vivo liver metabolism in human iPSC-derived organoids]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723688v1?rss=1">
<title>
<![CDATA[
Targeting cancer-associated cell surface RNAs with oligonucleotide-drug conjugates enables broad antitumor activity 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723688v1?rss=1
</link>
<description><![CDATA[
Recent studies have revealed the presence of RNAs on mammalian cell surfaces, yet the linkage of cell surface RNAs (csRNAs) to cellular states and their potential as extracellular accessible drug targets remains unexplored. Here we develop Cell-surface and Intracellular RNAs Co-mapping (CIRCmap), a highly multiplexed in situ profiling approach that simultaneously detects thousands of cell-surface and intracellular RNAs at single-cell resolution. Using CIRCmap, we uncover that csRNA distributions are correlated with cellular states and identified cancer-associated csRNAs. Integrative analysis of cell-surface and intracellular RNAs within the same cells implies that the csRNAs undergo endocytosis and endolysosomal trafficking, which is further supported by perturbation experiments and co-localization visualization. We design oligonucleotide-drug conjugates (ODCs) that target cancer-associated csRNAs for endocytic delivery of cytotoxic payloads selectively to cancer cells. ODCs exhibit broad antitumor activity in cell lines and an in vivo mouse tumor model, opening a promising new avenue for targeted cancer therapy.
]]></description>
<dc:creator><![CDATA[ Luo, C., Cheng, C., Zhao, X., Zhou, J., Jing, Z., Ma, Q., Ye, L., Zhu, K., Zhao, L., Xu, H., Zeng, H. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723688</dc:identifier>
<dc:title><![CDATA[Targeting cancer-associated cell surface RNAs with oligonucleotide-drug conjugates enables broad antitumor activity]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723889v1?rss=1">
<title>
<![CDATA[
Size-dependent nucleus-vacuole interactions in budding yeast demonstrate a role for steric packing in organelle shape and positioning 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723889v1?rss=1
</link>
<description><![CDATA[
Although organelles are often studied one at a time, whole-cell imaging studies show that organelles take up a large part of the cell volume such that they are crowded together. Here we use whole cell soft X-ray tomography imaging to investigate how such crowding affects organelle size scaling, position, and shape, focusing on the nucleus and vacuole of budding yeast. We find that as the vacuole becomes larger, the nucleus loses its normal scaling relation with respect to cell volume, becomes displaced from its normal position near the cell center, and becomes progressively deformed from a sphere into a pancake shape. Using a whole-cell integrated modeling framework, we find that these changes are statistically correlated and give rise to distinct modes in cell organization space. Using a simplified mechanical model for two initially spherical compartments contained inside a confined intracellular space, we are able to recapitulate the effects seen in the experimental data, indicating that these observations are consistent with a purely mechanical interaction. Taken together, our work indicates that, in addition to the well-known protein-based organelle-organelle interactions, physical steric packing of organelles inside a limited cellular volume also plays a large role in the inter-organelle relationships and the overall geometry of the cell.
]]></description>
<dc:creator><![CDATA[ Mirvis, M., Akenuwa, O. H., Lee, C. T., Marshall, W. F. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723889</dc:identifier>
<dc:title><![CDATA[Size-dependent nucleus-vacuole interactions in budding yeast demonstrate a role for steric packing in organelle shape and positioning]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723767v1?rss=1">
<title>
<![CDATA[
A genome-wide RNAi screen identifies host cell cycle regulation as a determinant of Orientia tsutsugamushi infection 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723767v1?rss=1
</link>
<description><![CDATA[
Orientia tsutsugamushi (Ot) is an obligate intracellular bacterium that causes scrub typhus, a potentially life-threatening disease. To systematically identify host factors regulating early stages of infection, we performed a microscopy-based genome-wide siRNA screen in HeLa cells. This approach identified 2,989 genes grouped into 55 functional networks that modulate bacterial entry and intracellular translocation. In addition to confirming previously well described pathways, including endocytosis and microtubule-dependent trafficking, the screen revealed an association between Ot infection and host cell cycle regulation. We found that Ot preferentially infects host cells in the S and G2 phases, where intracellular bacterial accumulation is increased relative to G1. Early infection was associated with a shift in host cell cycle distribution, consistent with a delay in progression through S and G2 phases. Longitudinal analysis further showed that these cell cycle states support enhanced bacterial expansion. In parallel, infected cells exhibited reduced proliferation compared to uninfected cells, suggesting that Ot infection alters host cell division dynamics. Together, these findings support a model in which host cell cycle state influences susceptibility to Ot infection and intracellular growth. This work provides a systems-level map of host pathways involved in early infection and identifies cell cycle regulation as an important component of host-pathogen interactions in scrub typhus.
]]></description>
<dc:creator><![CDATA[ Chusorn, P., Pittayasathornthun, Y., Kanchanapiboon, P., Saharat, K., Phongkitkarun, K., Sampattavanich, S., Salje, J. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723767</dc:identifier>
<dc:title><![CDATA[A genome-wide RNAi screen identifies host cell cycle regulation as a determinant of Orientia tsutsugamushi infection]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723799v1?rss=1">
<title>
<![CDATA[
The proximal lipid phase of PI3K signaling is confined to the plasma membrane 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723799v1?rss=1
</link>
<description><![CDATA[
Class I phosphoinositide 3-kinases (PI3Ks) generate the lipid second messengers PIP3 and PI(3,4)P2 to control diverse cellular processes including growth, metabolism, and survival. Although these signals are classically thought to arise at the plasma membrane, several recent studies have proposed that PI3K signaling is propagated from intracellular membranes along the endocytic pathway. Here, we combined genomic tagging of endogenous PI3K pathway enzymes with single-molecule imaging and sensitive lipid biosensors to define the spatial organization of PI3K signaling in living cells. We find that PI3K catalytic subunits are recruited to the plasma membrane but do not undergo detectable endosomal translocation during receptor activation. Consistently, PIP3 and PI(3,4)P2 accumulation is restricted to the plasma membrane, despite enrichment of lipid phosphatases along the endocytic pathway. Functional perturbation experiments further show that degradation of PI(3,4)P2 occurs predominantly at the plasma membrane, indicating that both synthesis and termination of proximal lipid signals are spatially confined to this compartment. Together, these results resolve the subcellular localization of proximal PI3K signaling and support a model in which lipid second messenger production is restricted to the plasma membrane, with diversification of downstream pathway outputs occurring through redistribution of activated effector proteins rather than intracellular propagation of lipid signals.
]]></description>
<dc:creator><![CDATA[ Ricci, M. M. C., Patel, H., Montoya, M. J., Jayasuriya, K. L., Rectenwald, A., Motter, M. A. K., Wills, R. C., Hammond, G. R. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723799</dc:identifier>
<dc:title><![CDATA[The proximal lipid phase of PI3K signaling is confined to the plasma membrane]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723543v1?rss=1">
<title>
<![CDATA[
HS3ST1 regulates pulmonary inflammation and is a determinant of clinical outcomes after trauma and hemorrhagic shock 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723543v1?rss=1
</link>
<description><![CDATA[
Mechanisms that promote organ injury after trauma and hemorrhagic shock (T/HS) remain poorly defined. Endothelial heparan sulfates with a 3-O-sulfate (3-OS) modification controlled by the HS3ST1 gene have anticoagulant and anti-inflammatory properties through their interaction with antithrombin. Our objective was to determine whether HS3ST1 deficiency drives organ injury and poor outcomes after T/HS. Hs3st1-/- and wild-type (WT) mice were subjected to T/HS followed by resuscitation with lactated ringers (LR) or fresh frozen plasma (FFP). While no differences were observed between WT and Hs3st1-/- LR resuscitated mice, lung injury and leukocyte infiltrates were significantly increased in FFP resuscitated Hs3st1-/- compared to WT mice. In vitro, leukocyte slow rolling and adherence was increased in HS3ST1 KO compared to WT cells. Among 472 T/HS patients, of which 31 (7%) were homozygous for the rs16881446 variant allele (GG), the number of ventilator free days was lower, and mortality was significantly higher in AG and GG patients. The rs16881446 genotype was independently associated with mortality. In conclusion, HS3ST1 deficiency mitigates organ protection from FFP resuscitation, partly through mediating EC:leukocyte engagement, and predicts mortality after T/HS. These findings identify a novel therapeutic target and prognostic tool that can be leveraged towards improved risk stratification after trauma.
]]></description>
<dc:creator><![CDATA[ Mokhtari, A. K., Cotton, M. E., Thomas, K. A., Chitrakar, A., Krocker, J. D., Pokharel, M., Osborn, B. K., Huby Viduarre, M. d. P., Mankame, A. R., Wade, C. E., Wang, Y.-W., Orlicky, D. J., Cohen, M. J., Richter, J. R., Shworak, N. W., Cardenas, J. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723543</dc:identifier>
<dc:title><![CDATA[HS3ST1 regulates pulmonary inflammation and is a determinant of clinical outcomes after trauma and hemorrhagic shock]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723186v1?rss=1">
<title>
<![CDATA[
Decoding murine corneal epithelial specification and homeostasis by single-cell spatial transcriptomics with scRNA-seq enrichment 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723186v1?rss=1
</link>
<description><![CDATA[
Investigation of gene regulatory programs underlying corneal epithelial cell specification and homeostasis is essential for understanding how the cornea maintains vision. Here, we describe the use of true single-cell resolution spatial transcriptomics (ST), enriched with full-tissue single-cell RNAseq (SC), to improve spatial resolution and enhance cell cluster size up to 65-fold and per-cell transcriptomic depth up to 17-fold. This enabled cell type specification across the full differentiation trajectory from limbal stem cells (LSC) to superficial corneal epithelium and identification of an activated signature (Atf3, Zfp36, Gsta4 and Dapl1) marking differentiation-primed states across multiple cell types, including a major activated intermediate epithelium (AIE) population. Validation using ST data from murine corneas at different postnatal ages and multiple human SC datasets confirms a large AIE population, which spatial localization and transcriptomic profiling suggest is an active intermediate state distinct from quiescent wing cells. Sub-clustering further revealed early (Sox9, Hes1), proliferative (Mki67, Top2a) and mature (Ccdn1, Dapl1) transient amplifying cell subpopulations and four LSC subpopulations, including putative active (Atf3, Socs3, Zfp36), quiescent (Gpha2, Ifitm3, Cd63) and Apoe-specific. Direct ST-to-SC comparison revealed enhanced axonal processes and genes (Sema3f, Sema 4d, Pax6) and cell-cell adhesion and cell-matrix markers (Itgb4, Tns4, Tjp3) in ST data, suggesting cell dissociation from tissue in SC masks epithelial innervation, adhesion and barrier functions. Our findings identify and localize key transcriptional programs in situ, prompting a re-evaluation of epithelial states in scRNA-seq data.
]]></description>
<dc:creator><![CDATA[ Javidjam, D., Vattulainen, M., Lagali, N., Moustardas, P. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723186</dc:identifier>
<dc:title><![CDATA[Decoding murine corneal epithelial specification and homeostasis by single-cell spatial transcriptomics with scRNA-seq enrichment]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723609v1?rss=1">
<title>
<![CDATA[
A CAMKK2-UBR4-19S Proteasome Axis Regulates Chondrocyte Proteostasis and SOX9 Stability 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723609v1?rss=1
</link>
<description><![CDATA[
In early osteoarthritis (OA), chondrocytes undergo a shift from anabolic to catabolic states driven by a coordinated interplay between inflammation and the ubiquitin-proteasome system (UPS) that results in a rapid turnover of anabolic proteins, including the master chondrogenic transcription factor Sox9. However, the molecular mechanisms linking these processes remain unclear. Ca2+/calmodulin dependent protein kinase kinase 2 (CaMKK2) is elevated in OA cartilage, where it coordinates chondrocyte inflammatory responses. Here, we identify CaMKK2 as a novel regulator of chondrocyte UPS and Sox9 degradation. Pharmacological inhibition of CaMKK2 increased SOX9 in human OA cartilage, whereas its loss or inhibition preserved Sox9 following destabilization of the medial meniscus (DMM) surgery in mice. Expression of kinase-active, not kinase-inactive, CaMKK2 reduced Sox9 in human and mouse chondrocytes, indicating a role in Sox9 turnover. Proteomic analysis of CaMKK2 immunoprecipitates revealed the presence of ubiquitin E3 ligase Ubr4 and the 19S proteasome regulatory particle (RP). CaMKK2 kinase activity was dispensable for its interactions with Ubr4 and 19S RP subunits, and kinase-inactive CaMKK2 enriched higher molecular weight Sox9-ubiquitin conjugates and attenuated Sox9 degradation in chondrocytes. Mechanistically, CaMKK2 directly phosphorylated the 19S RP ATPase Psmc5 (Rpt6) on Ser136, and an intact kinase increased proteasome activity, which was diminished in Camkk2-/- chondrocytes. Our findings define CaMKK2 as a dual-function regulator of chondrocyte UPS with a scaffolding role to assemble Ubr4-19S RP complexes around polyubiquitinated proteins such as Sox9, and a catalytic role to enhance proteasome function, potentially through Psmc5 phosphorylation, thereby linking chondrocyte inflammatory signaling to Sox9 degradation and cartilage degeneration.
]]></description>
<dc:creator><![CDATA[ Ding, X., Li, Y., Hansen, K., Mosley, A. L., Yeh, E. S., Doud, E. H., SANKAR, U. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723609</dc:identifier>
<dc:title><![CDATA[A CAMKK2-UBR4-19S Proteasome Axis Regulates Chondrocyte Proteostasis and SOX9 Stability]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723713v1?rss=1">
<title>
<![CDATA[
Benzopyrene induces keratinocyte senescence and p21-dependent differentiation 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723713v1?rss=1
</link>
<description><![CDATA[
In this study, we demonstrate that Benzo[a]pyrene (B[a]P) induces keratinocyte senescence and p21Cip1-dependent keratinocyte differentiation. Atmospheric and environmental pollution are known to induce senescence and promote terminal differentiation in human primary keratinocytes, thus driving skin aging. However, much is still unknown about the underlying molecular mechanisms. We observed that B[a]P, a common atmospheric pollutant, induced senescence in primary keratinocytes in both two-dimensional and three-dimensional (reconstructed human epidermis) culture. This was accompanied by signs of DNA damage in B[a]P-treated cells. B[a]P-treated cells also underwent accelerated late-stage terminal differentiation, indicated by increased IVL and FLG expression from 48 to 96 hours post-exposure. While pharmacological and genetic attenuation of p21Cip1 did not rescue cellular senescence, it prevented the expression of IVL and FLG, suggesting that the late-stage terminal differentiation induced by B[a]P exposure was p21-dependent. Our data thus suggest a key role for the p21Cip1 in the keratinocyte response to pollution-induced damage, where p21Cip1 induces terminal differentiation to maintain skin barrier homeostasis.
]]></description>
<dc:creator><![CDATA[ Law, D. C. L., Tang, M. L. F., Van Steensel, M. A. M. ]]></dc:creator>
<dc:date>2026-05-12</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723713</dc:identifier>
<dc:title><![CDATA[Benzopyrene induces keratinocyte senescence and p21-dependent differentiation]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723579v1?rss=1">
<title>
<![CDATA[
Alternative organelle targeting of OPA1 mediates fatty acid release from lipid droplets 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723579v1?rss=1
</link>
<description><![CDATA[
Mitochondria and lipid droplets (LDs) are functionally coupled to coordinate fatty acid utilization and storage. However, a comprehensive understanding of mitochondria-LD alliances remains elusive. We have identified a previously unrecognized role for optical atrophy 1 (OPA1), a mitochondrial fusion factor, in the regulation of fatty acid release from LDs. We demonstrated that OPA1's exon 4 adapts an amphipathic helix to target OPA1 to LDs. OPA1 localized to LDs promote fatty acid release by facilitating the recruitment of lipases to LDs. In addition, OPA1's residence on LDs competes with its mitochondrial entry, influencing mitochondria fusion and connectivity. Furthermore, the S158N polymorphism within OPA1's exon 4 exhibiting attenuated fatty acid release from LDs is associated with changes in metabolic traits in pediatric cancer survivors. Altogether, our findings reveal that OPA1 actively mediates fatty acid release from LDs and provide a mechanistic link between OPA1 and human metabolism.
]]></description>
<dc:creator><![CDATA[ Li, X., Voronin, D., Bhattacharyya, R., Klein, J., Haas, M., Cho, W. J., Robinson, C. G., Throm, R. E., Wu, G., Li, C., Sapkota, Y., Niemi, N., Pruett-Miller, S. M., Opferman, J. T., Chang, C.-L. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723579</dc:identifier>
<dc:title><![CDATA[Alternative organelle targeting of OPA1 mediates fatty acid release from lipid droplets]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723498v1?rss=1">
<title>
<![CDATA[
Liver sinusoidal endothelial cells integrate metabolic and immune signals for MAPK-dependent BMP6 regulation and hepcidin induction 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723498v1?rss=1
</link>
<description><![CDATA[
Liver sinusoidal endothelial cells (LSECs) separate the blood from the hepatic parenchyma and thus are at the frontline as scavengers of blood-borne waste, pathogens and metabolic stimuli. LSECs are also critical for sensing systemic iron availability by controlling the synthesis of bone morphogenetic protein (BMP) 6, which is essential for hepcidin expression in hepatocytes. Hepcidin maintains systemic iron homeostasis by inhibiting dietary iron uptake and iron release from iron recycling macrophages. Hepcidin is also an acute-phase protein and its activation by inflammation requires active BMP signaling. It is incompletely understood how signals derived from inflammation, cellular damage and iron are integrated by the liver to assure adequate hepcidin expression. Here, we show that Bmp6 expression is activated in primary LSEC cultures upon their exposure to danger-associated molecular patterns (DAMPs), such as heme and myoglobin, pathogen-associated molecular pattern (PAMPs), such as lipopolysaccharide (LPS) and Fibroblast-Stimulating Lipopeptide-1 (FSL1), or oxidative stress inducers (H2O2). Interestingly, all regulatory cues converge at the MAPK signaling pathway, although the specific signaling branches involved are stimulus-specific. Of note, Bmp6 upregulation in LSECs in response to all signals tested is strongly enhanced by the hepatocyte secretome. As hepatocytes critically depend on active BMP/SMAD signaling to control hepcidin activation, our results reveal that multiple sources of signaling input activating Bmp6 in LSECs and hepcidin in hepatocytes serve to determine BMP/SMAD signaling strength. Furthermore, our findings identify hypoferremia (low plasma iron levels), the result of high hepcidin levels due to elevated Bmp6, as a convergent response in conditions of inflammation, oxidative stress and cellular damage.
]]></description>
<dc:creator><![CDATA[ Qiu, R., Cucinelli, S., Mertens, C., Colucci, S., Altamura, S., Hentze, M. W., Muckenthaler, M. U. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723498</dc:identifier>
<dc:title><![CDATA[Liver sinusoidal endothelial cells integrate metabolic and immune signals for MAPK-dependent BMP6 regulation and hepcidin induction]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723512v1?rss=1">
<title>
<![CDATA[
Cannabidiol confers neuroprotection against 6-OHDA toxicity by rescuing Nrf2 proteostasis and preserving mitochondrial integrity 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723512v1?rss=1
</link>
<description><![CDATA[
Oxidative stress and the progressive degeneration of dopaminergic neurons are key features of Parkinsons disease (PD). The intrinsically disordered structure of the transcription factor Nuclear factor erythroid 2-related factor 2 (Nrf2), which coordinates the main cellular antioxidant response of the body, makes it highly susceptible to misfolding and aggregation under severe oxidative stress, compromising cellular survival. Cannabidiol (CBD) has potent neuroprotective properties, but its exact molecular mechanism within the dopaminergic redox environment remains unclear. In this study, we investigated the protective effects of CBD against 6-hydroxydopamine (6-OHDA)-induced toxicity in both undifferentiated and mature, post-mitotic differentiated SH-SY5Y cells. We found that CBD confers robust Nrf2-dependent neuroprotection against 6-OHDA. Importantly, we uncover a previously unexplored mechanism of neuroprotection by which CBD actively prevents the stress-induced sequestration of Nrf2 into insoluble cytoplasmic inclusions under oxidative stress. We find that CBD keeps Nrf2 in a soluble, functional state, increases Ser40 phosphorylation, restores nuclear localization, and drives the robust transcriptional upregulation of antioxidant enzymes. This targeted activation of Nrf2 effectively reduces intracellular ROS, significantly attenuates mitochondrial fragmentation, and decreases aberrant mitophagic activity. Overall, our results show that rather than merely scavenging reactive oxygen species, CBD directly increases Nrf2 activity during oxidative stress, enabling a sustained cytoprotective response. We thus identify CBD as a highly specific, targeted molecule with a high potential for neuroprotective therapy in PD.
]]></description>
<dc:creator><![CDATA[ Jurado Coronel, J. C., Duennwald, M. L. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723512</dc:identifier>
<dc:title><![CDATA[Cannabidiol confers neuroprotection against 6-OHDA toxicity by rescuing Nrf2 proteostasis and preserving mitochondrial integrity]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723032v1?rss=1">
<title>
<![CDATA[
Angiotensin II and cAMP signaling pathways regulate mitochondrial biogenesis and activity in human adrenocortical cells. 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723032v1?rss=1
</link>
<description><![CDATA[
Mitochondrial homeostasis, governed by the balance between biogenesis and mitophagy, is essential for steroidogenesis in adrenocortical cells. While the requirement of active mitochondria for steroid synthesis is well-established, the hormonal regulation of genes governing mitochondrial function remains poorly understood. This study investigated whether angiotensin II (Ang II) and the cAMP/PKA pathway modulate the expression of key regulatory factors involved in mitochondrial biogenesis and redox status in the human adrenocortical H295R cell line. Using real-time qPCR and Western blot, we show that Ang II and 8Br-cAMP, a permeant analogue of cAMP, modulate NRF-1, Nrf2, UCP2, and ANT1 impacting on mitochondrial biogenesis, antioxidant defense, and respiratory activity. These molecular changes correlated with increased mitochondrial membrane polarization, as confirmed by MitoTracker red staining. Interestingly, Ang II stimulation promoted a time-dependent increase in TFAM levels, a key transcription factor in mitochondria, which correlates with the increase in mitochondrial DNA (mtDNA) content. The rate of oxygen consumption (OCR) and mitochondrial parameters were determined, with results showing that Ang II led to a significant increase in basal and maximum respiration, ATP production, and proton leak. These findings suggest that hormone stimulation favors mitochondrial activity, thereby enhancing the bioenergetic capacity of adrenocortical cells. Furthermore, treatment with the uncoupler CCCP triggered a retrograde signaling response, upregulating nuclear-encoded mitochondrial genes to counteract mitochondrial membrane depolarization. Our findings demonstrate for the first time that hormonal signals directly modulate the mitochondrial genetic program in H295R human adrenocortical cells, optimizing the bioenergetic platform required for efficient steroidogenic function.
]]></description>
<dc:creator><![CDATA[ Belluno, M. A., Arona, F. G., Helfenberger, K. E., Rodrigo, M. A., Mori Sequeiros Garcia, M. M., Maloberti, P. M., Benzo, Y., Poderoso, C. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723032</dc:identifier>
<dc:title><![CDATA[Angiotensin II and cAMP signaling pathways regulate mitochondrial biogenesis and activity in human adrenocortical cells.]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.722930v1?rss=1">
<title>
<![CDATA[
Talin controls the spatial distribution of vinculin tension in focal adhesions 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.722930v1?rss=1
</link>
<description><![CDATA[
Cells transmit force between the extracellular matrix and the actin cytoskeleton through integrin adhesion complexes centred on talin and vinculin. Vinculin binds talin through -helical vinculin-binding sites (VBS) that are exposed when talin rod domains unfold under force. Dissecting the significance of this interaction has relied heavily on the A50I mutation in vinculin, which has been widely used as a talin-binding-null mutant. Here we show that although the A50I mutation abolishes binding to the -catenin VBS, it retains nanomolar-affinity binding to multiple talin VBS. We therefore designed an improved mutant, I12K/A50I, that eliminates this residual talin binding. Biochemical assays and single-molecule stretching experiments demonstrate that I12K/A50I VD1 fails to bind talin even when VBS are exposed by force. Using vinculin tension and conformation sensors, we show that talin binding is required for efficient recruitment of vinculin to focal adhesions and for establishing spatial gradients of vinculin tension. However, vinculin can still experience mechanical load in the absence of talin binding. These results demonstrate that A50I is not a talin-binding-null and reveal that, while talin is not required for vinculin loading, it is essential for organising the spatial distribution of mechanical load within adhesion complexes.
]]></description>
<dc:creator><![CDATA[ Kallem, T., Guo, Y., Reynolds, M. K., Ball, N. J., Athale, M., Baker, K. B., Mykuliak, V. V., Ek, F., Aldaz-Casanova, S., Turkki, P., Hytonen, V. P., Brown, N. H., Hoffman, B. D., Yan, J., Goult, B. T. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.722930</dc:identifier>
<dc:title><![CDATA[Talin controls the spatial distribution of vinculin tension in focal adhesions]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723282v1?rss=1">
<title>
<![CDATA[
Quantitative analysis of fibroblast migration reveals migratory states characterized by force generation, cell shape and motion 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723282v1?rss=1
</link>
<description><![CDATA[
Cell migration depends on coordinating cell shape changes with force generation, yet how these processes are integrated remains unclear. Here, we combine live-cell imaging with traction force microscopy and computational analysis to quantify cell morphology, motility and force generation in migrating fibroblasts. We find that traction force magnitudes display a multimodal distribution, suggesting discrete migratory regimes. Using a Hidden Markov Model, we identify distinct force states that exhibit differences in shape and motion metrics, and show that individual cells transition between force states over time. To test the role of cytoskeletal organization in establishing the identified states, we analyzed cells lacking Arpc2, which disrupts branched actin assembly. Despite reduced forces and altered morphology, these cells also exhibit three migratory states. State transitions occur more frequently in cells lacking Arpc2 and unlike normal cells their protrusion geometry is force dependent. Together, our findings show that cell migration is organized into discrete mechanical states that couple morphology, motility and force generation.
]]></description>
<dc:creator><![CDATA[ Davis, E. M., Hockenberry, M. A., Truscott, H. H., Shaul, N. J., Bear, J. E., Elston, T. C. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723282</dc:identifier>
<dc:title><![CDATA[Quantitative analysis of fibroblast migration reveals migratory states characterized by force generation, cell shape and motion]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723287v1?rss=1">
<title>
<![CDATA[
NO modulates human airway smooth muscle function by altering glucose-6-phosphate dehydrogenase effects on sGC function in asthma 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723287v1?rss=1
</link>
<description><![CDATA[
Since NO can modulate mesenchymal cell function, we posit that NO can modulate gene expression associated with excitation-contraction coupling. Our study shows that treating asthma-derived HASMCs with a low dose of NO plus sGC stimulator BAY-41, in most cases sensitized smooth muscle sGC towards activation via an elevated sGC heterodimer and in some cases also improved sGC{beta}1, catalase, Cyb5r3 or Trx1 expression (n=24 non-asthma and n=25 asthma). Interestingly we found that majority of asthma HASMCs showed a marked downregulation of G6PD expression inducing a low GSH/GSSG ratio in asthma, and these findings were replicated in murine lungs of allergic asthma (OVA and CFA/HDM). Studies with HEK/COS-7 cells showed G6PD synergizing with hsp90 in enabling sGC heme-maturation. G6PD overexpression in HASMCs enhanced the sGC heterodimerization while silencing of endogenous G6PD abrogated it. Complementation of these cellular results with whole animal models of G6PD deficiency or overexpression provided verification to our findings. Mouse lung tissue from the humanized variant of G6PD deficiency, V68M (G6PD A- deficiency) showed significant downregulation in the sGC heterodimer, with a concomitant reduction in its NO heme-dependent activity, thereby showing that G6PD deficiency lowers sGC heme. Conversely, G6PD overexpressing mouse lung tissue displayed an elevated sGC heterodimer and also showed a robust G6PD-sGC{beta}1 interaction, suggesting G6PD to be involved in the heme-maturation of sGC{beta}1. While G6PD maintains the cell redox by generating NADPH, its new role in regulating sGC maturation links sGC dysfunction in asthma to G6PD deficiency and may potentially uncover new targets for asthma treatment.
]]></description>
<dc:creator><![CDATA[ Ghosh, A., Sumi, M. P., Koziol-White, C., Tupta, B., Wang, L., Ghosh, C., Jester, W. F., Panettieri, R. A., Stuehr, D. J. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723287</dc:identifier>
<dc:title><![CDATA[NO modulates human airway smooth muscle function by altering glucose-6-phosphate dehydrogenase effects on sGC function in asthma]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723011v1?rss=1">
<title>
<![CDATA[
Compartmentalized glycolysis powers ATP production in primary cilia and engages mitochondria via the phosphoenolpyruvate cycle 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723011v1?rss=1
</link>
<description><![CDATA[
Primary cilia are antenna-like sensory and signaling organelles present on most mammalian cells, including glucose-sensing pancreatic {beta}-cells. Here, we show that the local energetic demands of primary cilia require the ATP-producing enzyme pyruvate kinase. Loss of PKm1, but not PKm2, impairs ciliary glycolytic flux. While the entire glycolytic machinery localizes to cilia, our data indicate that mitochondria are a critical source of phosphoenolpyruvate (PEP), the high-energy glycolytic intermediate that drives the pyruvate kinase reaction. Abolishing PCK2, the mitochondrial enzyme that generates PEP, prevents cilia from sensing not only glucose but also the amino acids glutamine and leucine. Finally, by experimentally mislocalizing glycolysis, we demonstrate that primary cilia can utilize ATP generated within the cell body when glucose is limited. These findings indicate that primary cilia possess the capacity for local ATP generation, and when necessary, leverage a ciliary-mitochondrial signaling axis to meet their bioenergetic needs.
]]></description>
<dc:creator><![CDATA[ Huang, S. M., Foster, H. R., Lee, E. Y., Jo, J. H., Dong, X., Cho, B.-K., Goo, Y. A., Hughes, J. W., Merrins, M. J. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723011</dc:identifier>
<dc:title><![CDATA[Compartmentalized glycolysis powers ATP production in primary cilia and engages mitochondria via the phosphoenolpyruvate cycle]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.07.723676v1?rss=1">
<title>
<![CDATA[
USP10 Facilitates Homologous Recombination-Mediated DNA Double-Strand Break Repair through Localization to the Nucleolus 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.07.723676v1?rss=1
</link>
<description><![CDATA[
Ubiquitin-specific protease 10 (USP10) is a multifunctional deubiquitinating enzyme that primarily regulates cellular stress responses, including the DNA damage response. Here, we show that USP10 is required for homologous recombination (HR)-mediated repair of DNA double-strand breaks (DSBs) and for the maintenance of genomic stability. USP10-depleted cells exhibit spontaneous micronuclei, impaired DSB repair following zeocin and camptothecin treatment, and reduced sister chromatid exchange. These cells are also more sensitive to irradiation and mitomycin C and display increased chromosomal abnormalities after mitomycin C treatment. Persistent RAD51 foci formation in USP10-depleted cells suggests that USP10 functions at a step downstream of RAD51 nucleofilament formation. This function of USP10 in facilitating HR repair depends on deubiquitinase activity but is independent of G3BP1/2 and PABP binding. In addition, a newly identified nucleolar localization signal is required for the function of USP10 in DSB repair. Together, these findings indicate that USP10 maintains genome integrity by localizing to the nucleolus and facilitating HR-mediated repair of DSBs.
]]></description>
<dc:creator><![CDATA[ Utani, K., Sakasai, R., Himeda, T., Okuwa, T., Iwabuchi, K., Higuchi, M. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.07.723676</dc:identifier>
<dc:title><![CDATA[USP10 Facilitates Homologous Recombination-Mediated DNA Double-Strand Break Repair through Localization to the Nucleolus]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723346v1?rss=1">
<title>
<![CDATA[
Dual role of the OMM E3 Ub ligase MARCH5 in de novo peroxisome biogenesis and mitochondrial quality control through direct regulation of Pex26. 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723346v1?rss=1
</link>
<description><![CDATA[
Recent evidence indicates that mitochondria, through the activity of the E3 Ub ligase MARCH5, are critical for de novo peroxisome biogenesis. Here we report that peroxisome biogenesis factor Pex26 is a MARCH5 client protein. In peroxisome-containing cells, MARCH5 interacts with Pex26 and facilitates the transfer of newly synthesized Pex26 from the OMM to peroxisomes. MARCH5 also controls peroxisomal delivery of other candidate peroxins in peroxisome-containing cells. On the other hand, in peroxisome-deficient cells, the turnover rate of Pex26 is dramatically increased, and MARCH5 targets this protein for p97-dependent proteasomal degradation. Both activities are mediated by MARCH5-dependent Pex26 ubiquitination. Knockout of Pex26 induces the accumulation of cells containing Tom20-positive, Catalase-deficient pre-peroxisomes. Further supporting the critical role of MARCH5 in peroxisome biogenesis, these structures are absent in Pex26/MARCH5 double knockout cells. The data support the model, where in peroxisome-containing cells, MARCH5 acts as a peroxisome biogenesis factor, while with defective peroxisome biogenesis, as in Zellweger syndrome cells, it protects mitochondria from potentially toxic accumulation of peroxins on the OMM.
]]></description>
<dc:creator><![CDATA[ Bhattacharjee, D., Bippes, C. C., ZHAO, G., Boyman, L., Weldemariam, M. M., Kane, M. A., Neutzner, A., Karbowski, M. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723346</dc:identifier>
<dc:title><![CDATA[Dual role of the OMM E3 Ub ligase MARCH5 in de novo peroxisome biogenesis and mitochondrial quality control through direct regulation of Pex26.]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723885v1?rss=1">
<title>
<![CDATA[
A cohesion optimum underlies chromosome segregation fidelity in oocytes 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723885v1?rss=1
</link>
<description><![CDATA[
Chromosome segregation is compromised in eggs from women of both early and advanced reproductive ages. Deteriorating cohesion causes premature separation of sister-chromatids in eggs from older females. We show that the converse is true for oocytes of adolescents, with excessive cohesion impeding segregation. Oocytes from juvenile mice show severe chromosome lagging in anaphase I, leading to nondisjunction or, in extreme cases, failure of the first meiotic division. These defects are suppressed by experimentally weakening cohesion or enhancing its resolution during anaphase I. By contrast, lagging and nondisjunction are rare in the oocytes of young adults because cohesion is inherently weaker. Thus, relative cohesion strength underlies both the frequency and type of segregation errors observed in eggs throughout the female reproductive lifespan.
]]></description>
<dc:creator><![CDATA[ Yun, Y., Ikami, K., Do, D., Guo, Z., Leem, J., Bekele, H., Mogessie, B., Hunter, N. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723885</dc:identifier>
<dc:title><![CDATA[A cohesion optimum underlies chromosome segregation fidelity in oocytes]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723328v1?rss=1">
<title>
<![CDATA[
Rap1 Activates Endosomal AC9 to Drive the Second cAMP Wave 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723328v1?rss=1
</link>
<description><![CDATA[
GsPCR signaling orchestrates cAMP production in three distinct spatial waves originating from the plasma membrane (PM), endosomes, and the nucleus. While the molecular drivers of the PM and nuclear waves are well defined, the regulation of the endosomal cAMP wave remains insufficiently understood. Here, we identify the small GTPase Rap1b as a direct activator of the endosomal adenylyl cyclase 9 (AC9), revealing a novel mechanism for intracellular cAMP synthesis. Like Gs, Rap1b-GTP interacts with the C2 domain of AC9, allosterically enhancing its catalytic activity both in vitro and in cells. Using AC9 mutations that selectively disrupt Rap1b versus Gs binding, we elucidate the role of the Rap1b-AC9 unit in mediating the endosomal cAMP wave, introducing a new layer of spatial regulation to cAMP signaling.
]]></description>
<dc:creator><![CDATA[ Zhang, X., Pizzoni, A., Pinto, A., Altschuler, D. L. ]]></dc:creator>
<dc:date>2026-05-11</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723328</dc:identifier>
<dc:title><![CDATA[Rap1 Activates Endosomal AC9 to Drive the Second cAMP Wave]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-11</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723281v1?rss=1">
<title>
<![CDATA[
Transcriptional remodeling of ubiquitin regulatory networks during trained immunity 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723281v1?rss=1
</link>
<description><![CDATA[
Background: Trained immunity is a durable functional reprogramming of innate immune cells characterized by enhanced responsiveness upon secondary challenge. While metabolic rewiring and epigenetic remodeling are well-established features of this process, the contribution of ubiquitin-mediated post-translational regulation remains poorly defined. Methods: We performed an integrative analysis of publicly available human transcriptomic datasets derived from monocytes, macrophages, and PBMCs exposed to established training stimuli ({beta}-glucan, Bacillus Calmette - Guerin [BCG], and hemin - {beta} - glucan) followed by secondary stimulation. A curated panel of deubiquitinating enzymes (DUBs) and E3 ubiquitin ligases with established immune functions was analyzed for differential expression. Gene Ontology (GO) and KEGG pathway enrichment analyses were conducted to evaluate higher-order convergence across independent datasets. Results: Across multiple trained immunity models, we identified reproducible transcriptional remodeling of ubiquitin-modifying enzymes. USP25, OTUB1, and TRIM25 were consistently upregulated following restimulation, whereas several chromatin- and cytokine-regulatory DUBs-including USP3, USP4, USP7, USP16, MYSM1, and USP38 - were downregulated. Normalization to RPMI-restimulated controls reduced many activation-associated signals; however, USP25 remained persistently elevated, suggesting a stable training-associated signature. Pathway enrichment analysis independently demonstrated significant engagement of ubiquitin-related functional categories across datasets, supporting coordinated reorganization of ubiquitin regulatory networks. Conclusion: These findings identify selective transcriptional remodeling of the ubiquitin-proteasome system as a recurring feature of trained immunity. Integrating ubiquitin signaling into the established metabolic-epigenetic framework expands the conceptual model of innate immune memory and suggests that ubiquitin-modifying enzymes function as modulatory rheostats shaping immune amplitude and stability. Future functional and proteomic studies are required to determine whether these transcriptional signatures directly mediate trained immunity phenotypes.
]]></description>
<dc:creator><![CDATA[ Santelices, J., Schaefer, Z., Gachunga, W., Celeste, C., Parker, I. K. ]]></dc:creator>
<dc:date>2026-05-10</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723281</dc:identifier>
<dc:title><![CDATA[Transcriptional remodeling of ubiquitin regulatory networks during trained immunity]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-10</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723297v1?rss=1">
<title>
<![CDATA[
Coordinated cell and chloroplast growth and its perturbation by chloroplast DNA replication inhibition in green algae 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723297v1?rss=1
</link>
<description><![CDATA[
Coordination among cell growth, chloroplast expansion, and organelle genome dynamics is fundamental to algal physiology, yet its regulation remains unclear. We used time-resolved single-cell analyses to examine scaling relationships among cell size, chloroplast volume, nuclear dynamics, and nucleoid organization in Desmodesmus communis and Chlamydomonas reinhardtii under normal conditions and after inhibition of chloroplast DNA replication with nalidixic acid (NAL). Under control conditions, both species showed coordinated scaling among cell, chloroplast, and nuclear size, while nucleoid dynamics were driven mainly by changes in number. NAL disrupted these relationships in a species- and time-dependent manner. In C. reinhardtii, prolonged treatment uncoupled chloroplast and nuclear growth from cell expansion and led to fewer, enlarged nucleoids, consistent with impaired replication. In contrast, D. communis largely maintained coordinated scaling, with effects mainly limited to reduced nucleoid proliferation and delayed division. Temporal analyses indicated that NAL primarily affected nucleoid replication and segregation, with secondary consequences for chloroplast growth and cell-cycle progression. These findings identify chloroplast genome dynamics as a regulatory link between organelle growth and cell division.
]]></description>
<dc:creator><![CDATA[ Kselikova, V., Vankova, A., Audoor, S., Bhattacharjee, B., LOUIS, F., Mora, M., Singh, R., Alvarez, A., Goksal, E., Bisova, K. ]]></dc:creator>
<dc:date>2026-05-10</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723297</dc:identifier>
<dc:title><![CDATA[Coordinated cell and chloroplast growth and its perturbation by chloroplast DNA replication inhibition in green algae]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-10</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.09.721064v1?rss=1">
<title>
<![CDATA[
In vivo base editing via single myotrophic adeno-associated viruses in dystrophic mouse muscle and satellite cells 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.09.721064v1?rss=1
</link>
<description><![CDATA[
Duchenne muscular dystrophy (DMD) is the most common, lethal X-linked neuromuscular disorder of childhood and is caused by mutations in the Dmd gene that disrupt dystrophin expression. Although adeno-associated virus-mediated gene therapies hold tremendous promise for DMD treatment, their clinical applications have been limited by dose-dependent vector and genome-level toxicities. Here, we developed and tested a single-vector adenine base editing strategy as a potentially safer genome editing approach to recode the pathogenic nonsense mutation into a benign missense mutation in mdx4cvDMD mouse model. Delivered using a muscle-tropic adeno-associated virus (MyoAAV) at a clinically-feasible dose (4E13 VG/kg), this strategy enabled detectable molecular recoding of the mdx4cv mutation in mice ranging in age from 3 days to 6 months. Yet, the overall efficiency and therapeutic impact of in vivo base editing with this system was highest in mice treated at the juvenile stage, with animals administered MyoAAV vectors at 3 weeks of age showing robust recovery of dystrophin expression and significant improvement in muscle contractile properties only one month later. Notably, introduction of adenine base editors either earlier in development, in neonatal mice, or later, in adulthood, yielded substantially lower editing efficiencies, particularly in muscle satellite cells whose editing is essential to ensure durable rescue of dystrophin expression in growing and regenerating muscle. Taken together, these results demonstrate the therapeutic potential of single-vector adenine base editing for DMD and underscore the importance of recipient age and disease stage in achieving optimal treatment outcomes for this and other genetic muscle disorders.
]]></description>
<dc:creator><![CDATA[ Lin, K.-H., Lam, A., Ooijen, S., Maier, M., Kassis, G., Ellis, R., Messemer, K., Martin, J., Khairallah, R., Wagers, A. J. ]]></dc:creator>
<dc:date>2026-05-10</dc:date>
<dc:identifier>doi:10.64898/2026.05.09.721064</dc:identifier>
<dc:title><![CDATA[In vivo base editing via single myotrophic adeno-associated viruses in dystrophic mouse muscle and satellite cells]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-10</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723240v1?rss=1">
<title>
<![CDATA[
Differential tolerance for SEA domain misfolding encodes a MAPK pathway-specific response 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723240v1?rss=1
</link>
<description><![CDATA[
Signaling pathways often share components yet produce highly specialized biological responses. How signaling specificity is achieved between pathways utilizing common components is a fundamental question. In budding yeast, the same transmembrane mucin, Msb2, regulates two Mitogen-Activated Protein Kinase (MAPK) pathways controlling filamentous growth (fMAPK) and the response to osmotic stress (HOG). How this shared sensor distinguishes between stimuli and regulates different pathways is not clear. Using structure-guided analysis, we identified a conserved SEA (Sea urchin sperm protein, Enterokinase, Agrin) domain in fungal mucins and found that mutations disrupting protein folding selectively impair one pathway (fMAPK) but were tolerated by another (HOG). Mechanistically, these differences revealed distinct modes of signal transmission. The fMAPK pathway required an intact SEA domain and the cytosolic tail, consistent with a cis signaling mechanism that required structural coupling across the membrane. In contrast, the HOG pathway functioned independently of the cytosolic tail and tolerated misfolded SEA domain variants, consistent with trans signaling mediated by extracellular domains of interacting partners. The HOG pathway may detect misfolding as part of its sensing mechanism, as stressors that induce protein misfolding required Msb2 for survival. This work reveals how differential tolerance to protein deformation confers signaling specificity and identifies sensor deformation as a general feature of mechanosensory pathways that respond to environmental stress.

HIGHLIGHTSO_LISignaling pathways differ in tolerance to misfolding of a sensory domain
C_LIO_LIMisfolded SEA domains retain function in a stress pathway (HOG) pathway but not a cell differentiation pathway (fMAPK)
O_LIMisfolded SEA domain variants showed altered protein levels, mis-localization in the secretory pathway, and turnover by ERAD
C_LIO_LINon-functional variants lacked residues that stabilize the structure through intramolecular bonds
C_LI
C_LIO_LIDifferential tolerance for misfolding revealed distinct modes of signaling
O_LITrans signaling predominated in the HOG pathway and did not require proper SEA domain folding or the mucin cytosolic tail
O_LIA dominant hyperactive variant next to the SEA domain revealed basal interactions with the CR domain of tetraspanin
C_LIO_LIAlphaFold modeling showed distinct interactions occur between the SEA domain and tetraspanin in the basal and activated states
C_LI
C_LIO_LICis signaling predominated in the fMAPK pathway
O_LIRequired a properly folded SEA domain and conformational coupling to the cytosolic tail
C_LIO_LIYapsin processing was required for SEA domain activation and turnover of the mucin cytosolic tail
C_LI
C_LI
C_LIO_LIHOG pathway may sense protein misfolding as part of its activation mechanism.
C_LIO_LISEA domains are conserved throughout fungal mucins and mammalian glycoprotein sensors suggesting a generalizable mechanism
C_LIO_LIProtein deformation may provide information to survival pathways about environmental stress.
C_LI

GRAPHICAL ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=167 SRC="FIGDIR/small/723240v1_ufig1.gif" ALT="Figure 1">
View larger version (50K):
org.highwire.dtl.DTLVardef@20aorg.highwire.dtl.DTLVardef@85f517org.highwire.dtl.DTLVardef@9dd3f1org.highwire.dtl.DTLVardef@1c5976f_HPS_FORMAT_FIGEXP  M_FIG C_FIG Signaling pathways often share components yet activate different effector processes through mechanisms that remain unclear. The same mucin regulates two MAPK pathways (red and green), and the discovery of a conserved SEA domain provided insights into specificity mechanisms. In the fMAPK pathway that regulates filamentous growth, the mucin works in a classical manner, where an external signal (in this case underglycosylation by glucose limitation) transduces a signal to the cytosolic domain in cis. By comparison, the HOG pathway that responds to osmotic stress displayed a remarkable tolerance for mucin and SEA domain deformation. Protein variants that caused SEA domain misfolding, mislocalization, and degradation by ERAD retained function in the HOG pathway. Truncations that removed the cytosolic tail and transmembrane anchor were also functional. These phenotypes support a trans activation mechanism with external partners that was preferential for activation of the HOG pathway. SEA domain deformation may be induced by environmental stress as a trigger for the HOG pathway. Cells may detect misfolding of protein domains to gain information about environmental stress.
]]></description>
<dc:creator><![CDATA[ Priyadarshini, A., Cullen, P. J. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723240</dc:identifier>
<dc:title><![CDATA[Differential tolerance for SEA domain misfolding encodes a MAPK pathway-specific response]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723202v1?rss=1">
<title>
<![CDATA[
ILK binding to β1 Integrin is indirectly mediated by Kindlin-2 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723202v1?rss=1
</link>
<description><![CDATA[
Integrin-linked kinase (ILK) and kindlin-2 (K2) are key components of focal adhesions (FAs) that regulate cell-matrix adhesion and integrin signaling. Both proteins directly bind each other, but how they influence each others localization to FAs and binding to integrins remains a subject of ongoing debate. Here, we establish a sensitive workflow to study protein-protein interactions in cells by combining methods from biochemistry, cell biology and super-resolution microscopy. Together with an analytical framework this approach allowed us to distinguish direct from indirect molecular interactions and construct detailed interaction networks. Disrupting the ILK-K2 interaction reduced ILK localization to FAs and compromised integrin function, whereas K2 recruitment was unaffected. Our interdisciplinary approach also revealed that ILK does not directly bind {beta}1-integrin cytosolic domains in vitro and in cells. Instead, ILK was recruited to integrins exclusively through a K2-dependent mechanism, primarily via K2 bridging ILK and {beta}1 integrins. These findings define the hierarchical relationship between ILK and K2 in FAs and highlight the essential role of K2-mediated ILK recruitment for integrin adhesion and signaling.

Significance StatementHow cells anchor to their environment is a fundamental question in biology. Integrins provide such a connection by bridging the extracellular matrix and the cytoskeleton. A central regulator of the integrin machine is Integrin-linked kinase (ILK). How ILK is recruited to {beta}1 integrins is hotly debated since its discovery more than 30 years ago. By integrating cell biology and biochemistry with super-resolution DNA-PAINT microscopy and a novel spatial analysis framework, we demonstrate that ILK does not bind directly to integrin cytoplasmic tails. Instead, we found that ILK is recruited by kindlin-2 (K2) to active, adhesion plaque-resident integrins. This work resolves a long-standing controversy in cell biology and establishes a versatile workflow for distinguishing direct from indirect protein-protein interactions in situ.
]]></description>
<dc:creator><![CDATA[ Reinhardt, S., Boettcher, R., Brod, F., Speidel, J., Jungmann, R., Faessler, R. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723202</dc:identifier>
<dc:title><![CDATA[ILK binding to β1 Integrin is indirectly mediated by Kindlin-2]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.08.723607v1?rss=1">
<title>
<![CDATA[
Selective Elimination of TP53 Mutant Cells by Transcript-Activated Chromatin Shredding 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.08.723607v1?rss=1
</link>
<description><![CDATA[
Genetic mutations that drive cancer often occur in tumor suppressor proteins, including the p53 transcription factor which is altered in [~]40-50% of cases1,2. However, current therapies fail to target most such mutations because the mutant proteins typically lack defined drug-binding pockets, and restoring the endogenous function has proven challenging. Here, we programmed CRISPR-Cas12a2, an RNA-guided nuclease with trans-nucleolytic cleavage activities3,4, to selectively kill cancer cells by targeting cancer-specific transcripts. This approach eliminates cells by inducing trans chromatin cleavage, triggering DNA damage and cell death. Unlike existing methods, RNA-guided Cas12a2 senses cellular RNA signatures to shred chromatin, enabling precise targeting of undruggable mutations. Transcript-activated chromatin shredding provides an innovative paradigm to develop precision disease treatments for undruggable targets.
]]></description>
<dc:creator><![CDATA[ Zeng, J., Cheng, Z., Chen, H., Thompson, J., Crosby, K. T., Hang, H., Ngo, W., Xia, C., Rosas-Rivera, D., Kang, M. H., Mao, Y., Lee, G., Diffley, J. F. X., Song, Y., Qiu, L., Krah, N. M., Murthy, N., Jackson, R. N., Liu, Y., Ashworth, A., Doudna, J. A. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.08.723607</dc:identifier>
<dc:title><![CDATA[Selective Elimination of TP53 Mutant Cells by Transcript-Activated Chromatin Shredding]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723114v1?rss=1">
<title>
<![CDATA[
NFIC-mediated CaSR endocytosis defines a hyperactive TOMM20highCHGAhigh oxyphil cell state as a pathological driver of autonomous secondary hyperparathyroidism 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723114v1?rss=1
</link>
<description><![CDATA[
Secondary hyperparathyroidism (SHPT) is a debilitating complication of chronic kidney disease. Its clinical management is frequently compromised by calcimimetic resistance, a refractory state primarily driven by the progressive downregulation of the membrane calcium-sensing receptor (CaSR). Despite the centrality of CaSR as a therapeutic target, the mechanisms governing its aberrant expression and membrane localization remain incompletely elucidated. Here, we generated a single-cell transcriptomic atlas of human parathyroid tissues from SHPT and primary hyperparathyroidism (PHPT) patients, uncovering a unique stromal-immune niche that is specifically induced by uremic stress in SHPT. Our data also observed a striking dissociation between CaSR mRNA and its abundance as a membrane protein in SHPT tissues. Pseudotime trajectory analysis showed a progressive decline in CaSR pathway activity and a concomitant increase in endo-lysosomal activity along the trajectory, terminating in pathological oxyphil cells as the endpoint of chief cell differentiation in SHPT. Mechanistically, Nuclear Factor I C (NFIC) transcriptionally activated clathrin light chain B (CLTB) and Ras-related protein Rab7a (RAB7A) to trigger clathrin-mediated endocytosis and lysosomal degradation of CaSR, thus reducing its membrane abundance. This degradative program was further validated by multiplex immunofluorescence in TOMM20highCHGAhigh pathological oxyphil cells in human SHPT tissues. To translate these mechanistic findings into a clinically actionable strategy, we repurposed the clinically approved lysosomal inhibitor hydroxychloroquine (HCQ) to block this CaSR degradation pathway. In patient-derived xenograft (PDX) mouse models, co-administration of HCQ and cinacalcet acted synergistically to restore membrane CaSR expression, normalize serum PTH and calcium levels and suppress parathyroid tumor growth more effectively than monotherapies. Collectively, our single-cell-guided study identifies an NFIC-driven endo-lysosomal program as a previously unrecognized mechanism underlying CaSR downregulation and calcimimetic resistance in SHPT, and establishes HCQ repurposing as a clinically tractable therapeutic strategy for patients with refractory SHPT.
]]></description>
<dc:creator><![CDATA[ Yang, Q., Liu, J., Wang, Y., Zhao, R., Li, H., Yao, Y., Xu, C., Kou, B., Lei, M., Zhao, Q., Chen, X., Li, H., Zhao, R., Cui, R., Wang, M., Li, M., Yao, X., Bai, Y., Xia, F., Zhang, S., Liu, X., Li, X., Hou, P. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723114</dc:identifier>
<dc:title><![CDATA[NFIC-mediated CaSR endocytosis defines a hyperactive TOMM20highCHGAhigh oxyphil cell state as a pathological driver of autonomous secondary hyperparathyroidism]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723191v1?rss=1">
<title>
<![CDATA[
Septin crosstalk with microtubules and actin is regulated by a GSK3-dependent phosphoswitch 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723191v1?rss=1
</link>
<description><![CDATA[
Septins are cytoskeletal filaments that associate with the actin and microtubule cytoskeleton, but the mechanisms that govern septin crosstalk and function with these networks are largely unknown. Here, we show that glycogen synthase kinase 3 (GSK3) directly phosphorylates septin-9 (SEPT9), acting as a molecular switch that bidirectionally controls septin distribution between actin and microtubules. We show that GSK3 inhibition redistributes endogenous SEPT9 toward microtubules in multiple cell types. Phosphomimetic mutations at serines 82 and 85 reduce microtubule binding and enhance actin association in cells and in vitro, while phosphonull mutations promote microtubule binding and growth. In primary hippocampal neurons, GSK3{beta} inactivation promotes SEPT9-microtubule association, and phosphomimetic mutations impair asymmetric neurite growth during neuronal polarization. These findings reveal a phosphorylation-dependent mechanism of septin partitioning between actin and microtubules, placing the cytoskeletal functions of septins under the control of GSK3 - a kinase linked to multiple signaling pathways of cell physiology and metabolism.

HighlightsO_LIGSK3{beta} phosphorylates SEPT9, and its activity gates septin-cytoskeleton association
C_LIO_LIS82/S85 phosphorylation reduce microtubule binding and increase actin localization
C_LIO_LIUnphosphorylated SEPT9 binds preferentially to microtubules, promoting their growth
C_LIO_LIGSK3{beta} inactivation drives SEPT9 to microtubules to establish neuronal polarity
C_LI
]]></description>
<dc:creator><![CDATA[ Alam, M. N. A., Holt, T. C., Schaefer, A. W., Mayca-Pozo, F., Reghunathan, S., Butts, S. M., Bhakt, P., Kesisova, I. A., Spiliotis, E. T. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723191</dc:identifier>
<dc:title><![CDATA[Septin crosstalk with microtubules and actin is regulated by a GSK3-dependent phosphoswitch]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.06.723184v1?rss=1">
<title>
<![CDATA[
Temporal regulation of G2 phase avoids therapy-induced senescence caused by DNA replication stress-inducing drugs and provides synergistic cytotoxicity 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.06.723184v1?rss=1
</link>
<description><![CDATA[
The cellular response to DNA replication stress (DRS) provoked by anticancer drugs involves activation of the G2/M checkpoint (which promotes transient cell cycle arrest at G2 phase) and DNA repair, followed by induction of apoptosis or senescence. Here, we activated the p53-p21 pathway and ATR using DRS-inducing drugs, and found that that the transition to senescence depends on the duration of the G2 phase. Shortening of G2 duration by G2/M checkpoint inhibitors led not only to a switch in cell fate from senescence to mitotic entry, but also to effective cell death through carry-over of chromosomal aberrations (generated by DRS-inducing drugs) into mitosis and subsequent mitotic progression. Such enhanced cell death was also observed in p53 deficient cells, which do not normally undergo senescence. Thus, we propose that temporal regulation of G2 phase is an approach to enhancing the effects of DRS-inducing drugs in a manner that is independent of p53 status.
]]></description>
<dc:creator><![CDATA[ Nonaka, K., Wakasa, T., Ochiiwa, H., Kataoka, Y., Ando, K., Oki, E., Yoshizumi, T., Maehara, Y., Kitao, H., Iimori, M. ]]></dc:creator>
<dc:date>2026-05-09</dc:date>
<dc:identifier>doi:10.64898/2026.05.06.723184</dc:identifier>
<dc:title><![CDATA[Temporal regulation of G2 phase avoids therapy-induced senescence caused by DNA replication stress-inducing drugs and provides synergistic cytotoxicity]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-09</prism:publicationDate>
<prism:section></prism:section>
</item>
</rdf:RDF>
