Biology subjects

De, S.

Publications and source records attributed to De, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Dynamin-2 is a fission factor for SNX1-coated endosomal tubules

Sorting nexins of the SNX-BAR family shape endosomal membranes into tubular carriers that route cargo away from the degradative pathway, but we poorly understand the machinery that cuts these tubules to release transport carriers. Here we show that Dynamin-2 (Dyn2) is a fission factor for SNX1-coated tubules. In HeLa cells, Dyn2 localises to SNX1 tubules, and its knockout lengthens them and produces interconnected networks. Dyn2 binds SNX1 directly, and this interaction requires both the proline-rich domain of Dyn2 and the N-terminus of SNX1. On supported lipid bilayers, SNX1 alone tubulates membranes, and adding Dyn2 and GTP converts the emergent tubules into vesicles, indicating that no additional factors are needed for this activity. In cells, loss of Dyn2 redirects the retromer cargo GLUT1 to a LAMP1-positive compartment and reduces glucose uptake. These findings identify Dyn2 as the enzyme that releases SNX1 carriers and link this scission step to cargo recycling.

cell biology↗

bioRxiv2026

Two functionally distinct amyloid-β intermediates link aggregation pathways to pathology in Alzheimer's disease

Neurotoxic intermediates formed during amyloid-{beta}(A{beta}) aggregation are thought to drive Alzheimer's disease, yet their rarity and transient nature have obscured their identity and formation pathways in the human brain. Here, we combine super-resolution microscopy and single-molecule functional imaging with kinetic modelling to classify A{beta} intermediates according to their membrane-disrupting activity and connect their damaging effects to the aggregation pathways that generate them. Applying this framework to recombinant A{beta}42, an Alzheimer's knock-in mouse model and human Alzheimer's brain tissue, we find two functionally distinct intermediate populations present across all systems. They form through separate aggregation routes and differ in their neurotoxicity, abundance and susceptibility to inhibition. Both damage neurons and glia, but only one converts into plaques. Suppressing these intermediates pharmacologically in mice reduces plaque burden, attenuates astrogliosis and improves cognition. Our findings connect model systems with human pathology and identify intermediate formation pathways as actionable therapeutic targets across protein-misfolding disorders.

biophysics↗

bioRxiv2026

Reduced LANCL1-AS1 in old human skeletal muscle diminishes mitochondrial activity, shortens mt-mRNA poly(A) tails, and suppresses myogenesis

Regeneration of skeletal muscle preserves muscle mass and function, which decline with age. Here, we sought to identify long noncoding (lnc)RNAs involved in skeletal muscle myogenesis and potentially relevant to muscle aging. Cross-sectional analysis of skeletal muscle transcriptomes from healthy 22-through 89-year-old individuals revealed lncRNA LANCL1-AS1 among the top declining transcripts. Conversely, LANCL1-AS1 increased robustly during skeletal myogenesis and promoted myogenic differentiation in culture. Affinity pulldown by ChIRP followed by mass spectrometry revealed that LANCL1-AS1 associated with the mitochondrial protein LRPPRC, enhancing the formation of the chaperone complex LRPPRC-SLIRP, which maintains longer poly(A) tails of mitochondrial (mt-)mRNAs and stabilizes mt-mRNAs. Importantly, while myoblasts from old rhesus monkey muscle expressed lower levels of LANCL1-AS1 and mt-mRNAs, and displayed lower mitochondrial activity than young monkey myoblasts, overexpressing LANCL1-AS1 in old myoblasts restored mitochondrial activity and myogenesis. We propose that the age-associated reduction in LANCL1-AS1 contributes to impaired mitochondrial function and reduced myogenic capacity in aging skeletal muscle.

molecular biology↗

bioRxiv2026

Distinct clonal dynamics and interactions within the microenvironment near tumor stroma interfaces in rare histologic variants of bladder cancer

Rare histologic variants of bladder carcinoma, such as squamous cell and neuroendocrine carcinoma, generally have a worse prognosis compared to pure urothelial carcinoma (PUC), but the underlying molecular determinants are not well understood. We developed a novel computational genomics framework to characterize the dynamics of tumor-stroma-immune interactions at tumor borders and interiors from spatial transcriptomics data. We profiled bladder carcinoma samples of different histological variants on the Visium platform. Differences in clonal phylogeny and spatial heterogeneity of major subclones between the samples suggested disparate clonal spatiotemporal dynamics and interaction with stromal and immune compartments - which was notably prominent at the tumor-stroma interface. Our framework captured immune heterogeneity in the tumor microenvironment, including variations in the presence and architecture of tertiary lymphoid structures. Our analyses further indicated that there are substantial histology-specific differences in cell type composition, clonal spatial heterogeneity, inter-cellular signaling, and cellular processes. These variations collectively suggest divergent mechanisms of microenvironment remodeling across bladder cancer histologies. Cell-free DNA profiling from liquid biopsy captured tumor and microenvironment signatures from tumor boundaries and interiors, potentially allowing for tracking clonal dominance non-invasively. Our method tracks the trajectory of neoplastic disease in bladder cancer samples while identifying aggressive features.

genomics↗

bioRxiv2026

METTL16 promotes taxane resistance in Triple-Negative Breast Cancer through m6A-dependent translational upregulation of ABCB1

Triple-negative breast cancer (TNBC) commonly develops resistance to taxane-based chemotherapy, resulting in recurrence and poor clinical outcomes. Defining the molecular mechanisms that sustain chemoresistance is essential for improving therapeutic efficacy. Using unbiased insertional mutagenesis, we identified the RNA methyltransferase METTL16 as a previously unrecognized epi-transcriptomic driver of taxane resistance. METTL16 overexpression conferred resistance to docetaxel and paclitaxel across multiple TNBC models, and METTL16 expression was elevated in paclitaxel-resistant cells. Genetic depletion of METTL16 in paclitaxel-resistant cells restored taxane sensitivity. Because enhanced drug efflux is a well-established mechanism of taxane resistance, we investigated whether METTL16 regulates the multidrug transporter ABCB1 (P-glycoprotein). Paclitaxel-resistant TNBC cells exhibited elevated METTL16 and ABCB1 expression compared to parental cells. METTL16 binds to ABCB1 mRNA and catalyzes its N6-methyladenosine (m6A) modification, promoting increased ribosome loading and translational upregulation without altering transcript abundance. Inactivation of METTL16 impaired ABCB1 polysome association and restored paclitaxel sensitivity, demonstrating that the methyltransferase activity is essential for resistance. Consistent with this mechanism, METTL16 overexpression increased ABCB1 protein levels, whereas METTL16 down-regulation increased intracellular paclitaxel accumulation. Analysis of TNBC patient datasets revealed a positive correlation between METTL16 and ABCB1 expression, supporting the clinical relevance of this mechanism. Antisense-mediated inhibition of METTL16 using a translation-blocking Vivo-Morpholino reduced the survival of resistant TNBC cells and suppressed tumor growth in vivo. Surprisingly, genetic ablation of METTL16 caused profound loss of TNBC cell viability, while having only modest effects on nonmalignant mammary epithelial cells, indicating a cancer-selective dependency. Collectively, these findings define a METTL16-ABCB1 interaction that drives taxane resistance and establish METTL16 as a therapeutic target in TNBC.

cancer biology↗

bioRxiv2026

SenCat: Cataloging human cell senescence through multiomic profiling of multiple senescent primary cell types

There is an urgent need to comprehensively catalog senescence markers across cell types in an organism in order to characterize senotypes and senescent cell heterogeneity. Here, we profiled the transcriptomes and proteomes in 14 different primary human cell types undergoing over 30 senescence paradigms to create a senescence catalog we termed SenCat. We found that, while senescent cells from all primary tissue types did not share a single unique marker, they did activate shared specific metabolic and damage-response pathways implicated in tissue repair. Machine learning analysis of the SenCat transcriptomic and proteomic datasets successfully identified independent sets of senescent human cells, and senescent-like cells in mouse lung and kidney. In sum, SenCat represents a much-needed resource to identify senescent cells across tissues in the body. HIGHLIGHTSO_LIIdentifying senescent cells in organisms in vivo remains a challenge C_LIO_LIWe created SenCat: a catalog transcriptomes and proteomes of senescent primary cells C_LIO_LIMachine learning (ML) analysis of SenCat identified robust senescence scores C_LIO_LIML-derived senescence scores uncovered senescent-like cell dynamics in vivo C_LI

molecular biology↗

bioRxiv2026

Metabolite profiles distinguish exposure to Dengue and Zika flaviviruses in human induced pluripotent stem cells (hiPSCs)

Flaviviruses such as Dengue virus (DENV), Zika virus (ZIKV) and West Nile Virus (WNV) pose growing public health threats, exacerbated by asymptomatic infections and non-vector transmission. Current diagnostics, including nucleic acid tests and serology, are limited by transient viremia and antibody cross-reactivity, underscoring the need for improved detection methods. We utilized untargeted metabolomics to differentiate DENV and ZIKV infections in human induced pluripotent stem cells (hiPSCs), a model relevant to study virus-host interactions, drug screening, and therapeutic safety. LC-MSbased profiling revealed virus-specific metabolic reprogramming during both acute and long-term infections. DENV3 induced early and sustained activation of metabolic features, while ZIKV-MR766 triggered transient suppression. Long term infection with DENV2, DENV3, or ZIKV-PRV resulted in distinct metabolic signatures in hiPSCs, with ZIKV-PRV showing the greatest divergence. Shared metabolite changes across conditions included amino acids (e.g., tryptophan, glutamate), lipids, and nucleosides. Functional studies demonstrated that tryptophan metabolism regulates infection dynamics: inhibiting serotonin biosynthesis reduced viral load, whereas blocking kynurenine synthesis enhanced viral replication. These findings position metabolomics as a viable approach for flavivirus detection and highlight metabolic pathways as potential therapeutic targets. ImportanceAccurate diagnosis of flavivirus infections remains challenging due to short periods of viremia, high rates of asymptomatic infection, and extensive serologic cross-reactivity among related viruses such as DENV and ZIKV. These limitations complicate clinical diagnosis, surveillance, and screening of human donor tissues. In this study, we demonstrate that untargeted metabolomics can distinguish ZIKV and DENV infections by identifying virus-specific host metabolic signatures during both acute and long-term infection. Using human induced pluripotent stem cells as a clinically relevant model, we show that metabolic reprogramming persists even in the absence of overt cytopathic effects and can reveal functional pathways that regulate viral replication. Our findings highlight host metabolite profiling as a complementary diagnostic strategy that may improve detection of flavivirus exposure, particularly in asymptomatic individuals or settings where conventional molecular and serologic tests are insufficient.

microbiology↗

bioRxiv2026

Small molecule inhibition of the mitochondrial lipid transfer protein STARD7 attenuates influenza viral replication

The increasing appearance of drug-resistant and zoonotic influenza strains highlights an urgent need for host-directed antivirals that offer broad-spectrum activity and a higher barrier to resistance. Here, we describe the characterization of M4, a small-molecule identified from a high-thoughput screen that potently inhibits influenza A and B viruses. Mechanistic studies reveal that M4 suppresses replication by causing nuclear retention of the viral ribonucleoprotein (vRNP) complex. Chemoproteomic profiling identified the lipid transfer protein STARD7 as the primary cellular target, and genetic depletion of STARD7 phenocopies the antiviral effects of M4. Additional studies localizes the M4 binding site to cysteine 302 within the lipid-binding domain of STARD7, supporting a model in which STARD7-dependent lipid transfer activity promotes efficient vRNP trafficking and nuclear export. Combining M4 with baloxavir enhances antiviral efficacy in a murine infection model, providing in vivo support for a host-directed strategy. Together, these results identify STARD7 as a metabolic checkpoint licensing vRNP export and establish a proof of concept for therapeutic intervention with small molecule inhibitors. Author SummaryInfluenza viruses continue to cause widespread illness and pose an ongoing pandemic threat, in part because existing antiviral drugs can lose effectiveness as the virus evolves resistance. To address this challenge, we focused on identifying therapies that target host cell processes required for viral replication, rather than viral proteins themselves. In this study, we describe a small molecule, M4, that inhibits replication of both influenza A and B viruses by blocking a critical step in the viral life cycle. We found that M4 acts on a host protein called STARD7, which helps move certain lipids to the right places inside cells. When STARD7 is inhibited, the influenza viral ribonucleoprotein (vRNP) complex becomes trapped in the cell nucleus and cannot reach the cytoplasm, preventing the virus from completing its replication cycle. Disrupting STARD7 genetically produces the same effect, confirming that this host protein is important for influenza replication. These findings point to a previously unrecognized host gating mechanism that controls nuclear export of the vRNP complex during infection. Although M4 alone showed limited activity in animals, combining it with an existing antiviral drug strongly improved antiviral efficacy. Together, our results reveal a new host pathway that influenza viruses rely on and support host-directed combination approaches to strengthen antiviral treatment and help counter drug resistance.

microbiology↗

bioRxiv2026

Interplay of microRNA-20a and HuR in the regulation of beclin1 during Withaferin-A mediated impaired autophagy in Breast Cancer cell-line, MCF-7

Dysfunctional autophagy is connected to multiple diseases. Withaferin-A, a biologically active withanolide, has been shown to impair autophagy in the breast cancer cell line, MCF-7; however, the underlying mechanism remains unclear. Here while uncovering the mechanism, it was found that treatment of MCF-7 cells with WA declines Beclin1 protein synthesis by restricting association of its mRNA to polysomes with no significant reduction in its mRNA level. Reporter assay established that WA-treatment enhanced the expression level of mature miR-20a which directly interacts with the 3UTR of beclin1 mRNA. RNA affinity chromatography revealed association of mRNA stabilizing protein, HuR with the 3UTR of beclin1 mRNA. Additionally, co-immunoprecipitation assay determined the interactions of GW182, an essential component of GW-body and Ago2, a crucial member of miRNA related silencing complex (RISC) with the 3UTR of beclin1 mRNA. In parallel, it was found that knocking down of HuR eliminates the interaction of GW182 with beclin1 3UTR, while the association of miRNA (Ago2) remains unaffected. These results suggested simultaneous association of HuR, GW182 and microRNA with beclin1 mRNA which could be pivotal for sequestration of beclin1 mRNA in GW-bodies. Thus, our findings culminated in identifying an unconventional mechanism of regulation of Beclin1 expression by the dual interplay between hsa-miR-20a and HuR during WA-induced impaired autophagy in MCF-7 cells.

molecular biology↗

bioRxiv2025

Myeloid cells contribute to bystander CD8 T cell accumulation in metabolic-associated steatohepatitis and are sufficient for fibrosis

Background & AimsMetabolic dysfunction-associated steatohepatitis (MASH) is a leading cause of liver fibrosis, morbidity, and mortality. It is characterized by the accumulation of fat in hepatocytes causing cell death followed by stromal cell activation and scar deposition at later disease stages. While a marked accumulation of cytotoxic CD8 T cells is observed in MASH in humans and mice, the role of adaptive immune cells in fibrosis progression remains debated. MethodsTranscriptional data were curated from human datasets and preclinical mouse models, including mice deficient in TCR{beta} T cells (Tcrb-/-), which lack conventional CD4 helper and CD8 cytotoxic T cells, in diet-induced fibrosing MASH for single-cell RNA-sequencing. Specific human and mouse liver transcriptomic immune signatures were corroborated by flow cytometry and immunofluorescence. ResultsxMASH was associated with the expansion of lymphocytes and myeloid cells. Auto-aggressive CXCR6+PD1+FASLG+ CD8 T cells were enriched in livers of high-fat-diet-fed mice and correlated with MASH and fibrosis. Notably, Tcrb-deficient mice in a chemical and dietary preclinical MASH model developed fibrosis to the same extent as wild-type mice and exhibited exacerbated pro-fibrotic type 3 inflammation. Loss of conventional CD8 T cells neither impacted myeloid cell number nor phenotype within the fibrotic niche. Neutrophil- and non-conventional lymphocyte-derived GM-CSF and IL-17A were central drivers in the fibrotic niche composed of pathogenic macrophages and activated myofibroblasts. Furthermore, myeloid cells were identified as the main source of CXCL16 in diseased livers, retaining auto-aggressive CD8 T cells to the scar and contributing to their accumulation. Thus, non-adaptive cells, including scar-associated macrophages (SAMs) and myofibroblasts, are sufficient for MASH-driven fibrosis in this model. ConclusionsTargeting myeloid cells and fibroblasts should be prioritized as anti-fibrotic therapies for MASH. Graphical abstractThe immunopathological niche in fibrosis O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/694415v3_ufig1.gif" ALT="Figure 1000"> View larger version (47K): org.highwire.dtl.DTLVardef@122a24org.highwire.dtl.DTLVardef@8ca28borg.highwire.dtl.DTLVardef@103907borg.highwire.dtl.DTLVardef@10280e3_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsAdaptive and innate immune cells localize to the human MASH fibrotic niche CD8 T cells increase with MASH severity while MHC-II+ cells accumulate with fibrosis CXCR6+FASLG+ CD8 auto-aggressive T cells accumulate at the scar CXCL16+ scar-associated macrophages retain CD8 T cells in the fibrotic MASH niche Liver injury, inflammation and fibrosis persist in the absence of conventional T cells Impact and implicationsMetabolic dysfunction-associated steatohepatitis (MASH) is a progressive liver disease in which fibrosis is a key determinant of patient outcomes and survival. This study demonstrates that fibrosis in MASH can develop independent of conventional T cell immunity, driven by myeloid cells including scar-associated macrophages (SAMs) and neutrophils within a localized hepatic niche in mice and humans. This highlights the need to prioritize innate immune mechanisms as key drivers of chronic (late-stage) liver disease for researchers and clinicians developing anti-fibrotic therapies.

immunology↗

bioRxiv2025

Proteus mirabilis entails a switch from commensal to pathogen - Genomic insights from a blaTEM-1B-harboring novel isolate from India.

BackgroundProteus mirabilis is an opportunistic pathogen that can shift from gut commensalism to causing severe urinary tract and bloodstream infections. The genomic basis of P. mirabilis transition from commensalism to severe infection is not well understood, especially in Indian clinical isolates. This study aimed to explore the genomic attributes of a prevalent P. mirabilis strain in India. MethodsWhole-genome sequencing (WGS) was performed on a clinical P. mirabilis isolate by using NGS. AMR genes and virulence factors were identified using ResFinder/CARD and VFDB, respectively. Sequence typing was performed with MLST. Protein-protein interactions and noncoding RNAs were analyzed using STRING, CytoHubba, and BAKTA. ResultsWGS confirmed the multidrug-resistant nature of the isolate, revealing a broad repertoire of AMR genes, including blaTEM-1B, conferring resistance to beta-lactam antibiotics. Several resistance determinants were co-localised with insertion sequences on a single contig, indicating a high potential for horizontal gene transfer. The virulome profile highlighted multiple fimbrial genes, hemolysins, and toxin-encoding loci, together supporting strong adhesion, biofilm formation, and host tissue damage. Comparative analysis revealed unique genomic signatures not present in reference isolates, including additional virulence determinants and mobile genetic elements. MLST profile of the isolate is closely related to ST-675 & ST-797, while virulence typing (vST-1157) indicated a novel profile distinct from reference ST-675 strains. Insilico protein-protein interaction of this isolate revealed a highly interconnected virulence network, with urease, fimbrial adhesins, and hemolysins emerging as central functional modules driving pathogenicity. In addition, a noncoding RNA was identified within the genomic context of the ure operon (Contig 10), suggesting a possible regulatory role in urease expression. PPI network analysis highlighted urease, fimbrial proteins, and hemolysins as central hubs contributing to pathogenicity. ConclusionsThis study presents the first genomic characterization of a multidrug-resistant P. mirabilis clinical isolate from India, designated as P. mirabilis strain Indica. The convergence of AMR genes, virulence factors, MGEs and a novel noncoding RNAs linked to urease expression highlights the emergence of a high-risk variant in India. These findings have significant implications for infection control and emphasize the need for genomic surveillance to guide therapeutic strategies and develop new interventions globally.

microbiology↗

bioRxiv2025

Complex HPV-human DNA structures revealed by large-scale DNA analyses in an HPV-cancer derived cell line

Most human papillomavirus (HPV)-associated cancers harbor viral DNA integrated into the human genome as extrachromosomal circles, intrachromosomal segments, or both. Distinguishing intrachromosomal from identical-sequence extrachromosomal DNA (ecDNA) by sequencing alone is challenging, and the architecture of large-scale HPV-human DNA structures remains incompletely understood. To address this, we applied complementary genomic tools, spanning single-nucleotide to megabase resolution, to the HPV16-positive oropharyngeal cancer-cell line UM-SCC-47. These revealed that an initial integration event formed a 23 kb extrachromosomal heterocatemer circle comprising 7.5 kb of HPV16 DNA and 16 kb of the human TP63 gene. Subsequent genomic rearrangements generated heterocatemer tandem arrays extending to 0.6 megabases, plus additional large-scale rearrangements involving the HPV-TP63 structures, as revealed by long-read DNA sequencing and optical genome mapping. Fluorescent in situ Hybridization (FISH) showed that the heterocatemers were intrachromosomally localized at chromosome 3 at the TP63 locus in 100% of the cells. Long-read RNA sequencing further showed that these intrachromosomal templates produced spliced, polyadenylated transcripts. A subset of cells also harbored HPV16 ecDNA derived from the intrachromosomal HPV-TP63 DNAs. These findings define previously unrecognized higher-order architecture of integrated HPV DNA and highlight the power of FISH for distinguishing intrachromosomal from extrachromosomal DNA structures. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/680684v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@f3d584org.highwire.dtl.DTLVardef@470811org.highwire.dtl.DTLVardef@162c88corg.highwire.dtl.DTLVardef@5c80dd_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2025

Tretinoin, a Vitamin A derivative, exerts antiviral effects against Chikungunya virus infection through the nuclear retinoid receptor signaling pathways

Chikungunya virus (CHIKV) has reemerged as a global pathogen causing serious public health threat and socio-economic damage, particularly due to the absence of specific antivirals. Retinoids have been reported to exhibit antiviral potentials against multiple viral infections. In the present study, Tretinoin (TR) or all-trans retinoic acid (ATRA) was evaluated for its anti-CHIKV activity using in vitro, in vivo, and ex vivo models. TR was able to impede CHIKV infection efficiently in Vero and physiologically relevant muscle cells, C2C12, with drastic reduction in the viral RNA, proteins and progeny formation. The IC50 of TR against CHIKV was estimated to be 20.05 {micro}M and 19.71 {micro}M in Vero and C2C12 cells, respectively. In addition, the inhibition was effective during entry as well as in the early stages of post CHIKV-infection. Notably, TR showed remarkable virucidal activity. Next, global transcriptome profile demonstrated that the signaling by Retinoic acid pathway was highly upregulated after CHIKV infection and Retinoic acid receptors (RAR), specifically RAR-{beta} are the key mediators for the anti-CHIKV effect of TR. Further, the drug was also capable to modulate CHIKV-induced inflammatory responses by reducing the phosphorylated MAPKs, NF-{kappa}B and proinflammatory cytokines. Interestingly, TR protected C57BL/6 mice from CHIKV challenge by diminishing viral burden leading to reduced clinical scores and better survival. Similar observation was also noticed in the ex vivo model of CHIKV-infected hPBMC-derived monocyte-macrophage populations. In conclusion, this work demonstrated the repurposing potential of TR against CHIKV infection for the first time, encouraging its clinical validation towards future therapeutics. IMPORTANCECHIKV is an arbovirus causing Chikungunya Fever (CHIKF) leading to debilitating arthralgia and myalgia. The unavailability of licensed antivirals or potent vaccines against CHIKV encourages extensive research to find a therapeutic cure. Retinoids, the derivatives of vitamin A, have been repurposed as antiviral agents against a broad range of viruses; however, their efficacy against alphaviruses remains unexplored. Accordingly, Tretinoin (TR), also known as all-trans retinoic acid (ATRA), was validated for anti-CHIKV efficacy in this investigation. Significant perturbation of CHIKV titer through the modulation of Retinoic acid receptors (RAR) was observed upon drug-treatment. Further, TR reduced major inflammatory markers (MAPKs and proinflammatory cytokines) supporting its anti-inflammatory effect. Further, its anti-CHIKV potential was validated in the mouse model and hPBMC-derived monocyte-macrophage cells, indicating the pre-clinical efficacy. Finally, this is the first study to report the anti-CHIKV property of Tretinoin using in vitro, in vivo, and ex vivo approaches suggesting its repurposing potential.

microbiology↗

bioRxiv2025

Structural Studies of Nedicistrovirus IRES-Driven, Initiation Factor-independent Translation Shed Light on Key Steps of Eukaryotic Translation Elongation

We utilized the Nedicistrovirus (NediV) intergenic region (IGR) IRES-mediated, initiation factor-independent translation initiation system and determined high-resolution structures of 80S ribosome complexes with the NediV IRES in various functional states, including binary complexes, aminoacyl-tRNA-bound complexes, and complexes with elongation factor eEF2. In binary complexes, the NediV IRES primarily occupies the ribosomal P site, exhibiting conformational flexibility and engaging the ribosome at multiple interaction sites. Upon translocation, the IRES undergoes structural rearrangements, including destabilization of its PKI domain, facilitating the transition to canonical elongation. Crucially, we captured an eEF2-bound complex, along with an eEF1A-bound post-proofreading complex featuring a mismatched tRNA, the latter representing the first instance of a canonical elongation complex visualized in the presence of a natural, hydrolysable nucleotide and without the addition of any trapping agents. These findings provide a comprehensive structural overview of IGR IRES-mediated translation initiation and its transition to elongation, revealing key mechanistic details of viral translation and proofreading.

biochemistry↗

bioRxiv2025

Secondary nucleation of α-Synuclein drives Mitochondria dysfunctions and Lewy body formation in Parkinson's Disease

The seeding of -Synuclein (Syn) is a key driver of Lewy pathology propagation in Parkinsons disease (PD) and forms the basis for recent diagnostic advances. However, it remains unclear how the structural and biochemical features of Syn seeds dictate their propagation efficiency, capacity to induce Lewy body formation, and resulting cellular toxicity. Using genetic and idiopathic PD cell models, we map the pathogenic cascade beginning with the seed-driven conversion of endogenous Syn, followed by impaired degradation, mitochondrial dysfunction, and ultimately Lewy body formation. By coupling kinetic modelling of aggregation with functional readouts, we identify secondary nucleation as the predominant mechanism generating toxic Syn aggregation intermediates, identifying the critical process that links seeding to pathology. Extending this framework to PD brain, we quantitatively correlate seeding capacity with the spatiotemporal spread and severity of Lewy pathology, revealing a mechanistic connection between Syn aggregation dynamics and disease progression at molecular, cellular, and anatomical levels. By unifying molecular mechanism with clinicopathological progression, our work identifies catalytic Syn fibrillar seeds as tractable targets for both disease-modifying therapy and biomarker development in PD. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=193 HEIGHT=200 SRC="FIGDIR/small/676873v1_ufig1.gif" ALT="Figure 1"> View larger version (79K): org.highwire.dtl.DTLVardef@1eca299org.highwire.dtl.DTLVardef@a714d4org.highwire.dtl.DTLVardef@1486db5org.highwire.dtl.DTLVardef@1aa895_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LISyn fibril-oligomer interplay drives mitochondrial abnormalities and Lewy pathology C_LIO_LIFibrillar Syn catalyse toxic aggregate formations via secondary nucleation C_LIO_LIPhosphorylated Syn evades lysosomal clearance and drives enhanced dysfunctions C_LIO_LISeeding capacity of Syn predicts Lewy pathology burden and disease progression C_LI

neuroscience↗

bioRxiv2025

Unraveling proteomic chaos by independent component analysis - ClpX proficiency promotes the iron and oxygen limitation responses of Staphylococcus aureus and affects the intracellular bacterial behavior

In the opportunistic pathogen Staphylococcus aureus, protein homeostasis is largely mediated by the Caseinolytic protease (Clp) system. The proteases ClpXP and ClpCP are crucial for general and targeted proteolysis, which rely on the unfoldases ClpX and ClpC interacting with specific targets. However, the global effect on the proteome especially under infection-relevant stresses is not well-understood. To assess the effect of ClpX deficiency during infection-related processes, mass spectrometry-based global proteome profiles of S. aureus HG001 wild-type, an isogenic {Delta}clpX mutant, and a clpX complemented strain were recorded under control conditions as well as iron and oxygen limitation. The proteomic profiles revealed specific ClpX- and stress-dependent changes. A set of 24 robust stress-independent ClpX modulated proteins was identified and the stress-dependent influences were unraveled by independent component analysis (using the iModulon approach). These analyses revealed a role of ClpX in e.g., cell division, cell envelope homeostasis, the quinone stress response and prophage activation. Moreover, ClpX-dependent stress-specific effects were observed in the {Delta}clpX mutant, e.g. reduced induction of the heme uptake system under iron limitation and a dampened Rex-controlled oxygen limitation response. This revealed in particular that ClpX is central for heme homeostasis in S. aureus. Furthermore, in a Galleria infection model, the S. aureus {Delta}clpX mutant was attenuated compared to the wild-type HG001. This is consistent with a drastically reduced intracellular replication of the {Delta}clpX-mutant in cell culture-based infection experiments, however, high intracellular persistence of the {Delta}clpX mutant was also observed. This highlights the relevance of ClpX for bacterial fitness and virulence. ImportanceDuring infection processes, pathogens cope with host-mediated stressors. In response to those stressors, bacteria adapt their gene expression as well as their proteome profile. In the pathogen Staphylococcus aureus, protein homeostasis is mainly controlled by the Clp system. In particular, ClpX is the most conserved Clp unfoldase and is involved in overall regulation of virulence and bacterial fitness. However, the majority of ClpX targets remains elusive in S. aureus. With our proteomics approach and in depth data analysis, we provide a resource for global insight into ClpX-dependent adaptation of S. aureus physiology under infection-relevant conditions. Based on this, we uncover ClpXs role as a central player in the iron and oxygen limitation response. In addition, we demonstrate the importance of ClpX in S. aureus bacterial fitness in infection processes. However, reduced levels of ClpX lead to high intracellular persistence, which questions ClpXs suitability as a therapeutical target.

microbiology↗

bioRxiv2025

Mechanism-aware inference of response to targeted cancer therapies

Targeted therapies like small-molecule inhibitors often work by blocking proteins that cancer cells rely on for survival. Omics based modeling of drug sensitivity alone lack mechanistic grounding. We propose FORGE (Factorization Of Response and Gene Essentiality) a simple yet powerful joint matrix factorization framework that co-models drug response and target gene essentiality, enabling the stratification of promising treatment groups for targeted therapy consideration. FORGE also provides Benefit Score -- a predictive score that estimates treatment efficacy from basal gene expression profiles. We validated the predictive performance of FORGE across multiple targeted therapies, including Erlotinib (EGFR inhibitor) and Daporinad (NAMPT inhibitor). Our meta-analysis of large scale in-vitro studies underscores FORGEs ability to identify common determinants of drug vulnerabilities and target gene essentiality. Such convergences were not observed when treatment vulnerabilities and gene essentialities were modeled independently. We also demonstrated the universality of Erlotinib Benefit Scores by transferring transformations learned from high-throughput drug response studies across other published datasets, including the TAHOE-100M single-cell perturbation atlas and patient-derived xenograft studies. FORGE successfully identified key regulators within the molecular pathways targeted by these therapies, reinforcing its potential for mechanistically grounded treatment stratification.

cancer biology↗

bioRxiv2025

Senolytic treatment with fisetin reverses age-related endothelial dysfunction partially mediated by SASP factor CXCL12

BackgroundAdvancing age is the strongest risk factor for cardiovascular diseases (CVDs), primarily due to progressive vascular endothelial dysfunction. Cellular senescence and the senescence-associated secretory phenotype (SASP) contribute to age-related endothelial dysfunction by promoting mitochondrial oxidative stress and inflammation, which reduce nitric oxide (NO) bioavailability. However, the molecular changes in senescent endothelial cells and their role in endothelial dysfunction with aging remain incompletely unclear. As such, in this study we sought to identify the endothelial cell senescence-related signalling pathways, endothelial-derived SASP factors, and their impact on endothelial function with aging. MethodsSingle-cell transcriptomics was performed on aortas from young (6 months) and old (27 months) mice with and without in vivo senolytic treatment with fisetin (100 mg/kg/day administered in an intermittent dosing paradigm) to characterize endothelial cell senescence and transcript expression changes. Circulating levels of SASP factors were measured to validate transcriptional changes. Plasma exposure and protein addition and inhibiton experiments were conducted in isolated mouse arteries and cultured human endothelial cells to determine the causal role of the circulating SASP milieu and specific SASP factors in mediating endothelial dysfunction and underlying mechanisms-of-action. ResultsSenescent endothelial cells exhibited elevated expression of SASP factors, particularly Cxcl12, which was reversed by fisetin supplementation, with responses also reflected in circulating CXCL12 concentrations. Plasma from old mice impaired endothelial function by inducing vascular cell senescence, reducing NO, increasing mitochondrial oxidative stress, and promoting endothelial-to-mesenchymal transition--effects partially driven by CXCL12 and prevented by fisetin. ConclusionsThese results identify the SASP and CXCL12 as drivers of age-related endothelial dysfunction and establish mechanisms of senolytic intervention with fisetin supplementation. NOVELTY AND SIGNFICANCEO_ST_ABSWhat is known?C_ST_ABSO_LIAdvancing age is the primary risk factor for cardiovascular disease, in part due to progressive endothelial dysfunction. C_LIO_LICellular senescence contributes to age-related endothelial impairment through the secretion of a pro-inflammatory milieu known as the senescence-associated secretory phenotype (SASP), which can affect neighboring cells and tissue function. C_LIO_LISenolytic compounds selectively eliminate senescent cells and improve vascular function in preclinical models of aging. C_LI What new information does this article contribute?O_LIEndothelial cells are highly susceptible to senescence with aging in vivo and are selectively cleared by senolytic treatment with the natural compound fisetin. C_LIO_LISingle-cell transcriptomic profiling identifies CXCL12 as the most highly upregulated SASP factor in senescent endothelial cells and in circulation with aging, both of which are reversed by senolytic treatment with fisetin. C_LIO_LIWe identify a specific circulating SASP factor, CXCL12, as a partial mediator of endothelial dysfunction by inducing mitochondrial oxidative stress, impairing nitric oxide bioavailability, and promoting endothelial-to-mesenchymal transition (Endo, which is restored by senolytic treatment with fisetin. C_LI SummaryThis study provides novel mechanistic insight into how senescent endothelial cells and their secretory products--particularily CXCL12--contribute to age-related endothelial dysfunction. It further demonstrates that senolytic treatment with fisetin reverses these effects, highlighting a promising translational strategy for targeting vascular aging and preserving endothelial health. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=68 SRC="FIGDIR/small/670216v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@b49a8org.highwire.dtl.DTLVardef@110da92org.highwire.dtl.DTLVardef@17dbd54org.highwire.dtl.DTLVardef@1906c6_HPS_FORMAT_FIGEXP M_FIG C_FIG

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