Biology subjects

Kumar, A.

Publications and source records attributed to Kumar, A..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

GeneAutomate: A Browser-Based, Integer-Indexed Platform for Dual-Gene-List Functional Annotation and Interactive Network Visualization

Comparative interpretation of two gene lists, for example, two treatment arms, two tissues, or a discovery and a validation cohort, is a routine task in functional genomics. While several tools offer dual-list comparison (e.g., EnrichmentMap, RRHO packages), they typically require local software installation, R/Bioconductor, or manual reconciliation of separate single-list outputs. Most widely used web-based enrichment tools (DAVID, g:Profiler, Enrichr, ShinyGO, WebGestalt) are built around the analysis of a single gene list at a time, and those that support comparison often lack interactive, publication-ready visualization or depend on server-side query latency. Here we present GeneAutomate, a browser-based tool purpose-built for side-by-side comparison of two gene lists. GeneAutomate performs Over-Representation Analysis (ORA) against Gene Ontology (GO) and Reactome using an exact hypergeometric test with Benjamini-Hochberg false discovery rate correction, and Gene Set Enrichment Analysis (GSEA) when ranked (log2 fold-change) input is supplied, alongside Protein-Protein Interaction (PPI) subgraph extraction from BioGRID physical interactions. All reference data (Gene Ontology, Reactome, BioGRID, and NCBI/Ensembl identifier cross-references) are pre-compiled offline into a single integer-indexed database of approximately 32 MB for Homo sapiens, in which every gene identifier Ensembl ID, Entrez ID, official symbol, or alias is resolved to one canonical integer prior to any user query. This design removes live database round-trips from the runtime path, enabling fast, at-your-desk enrichment without installation or a server-side per-query bottleneck. The tool renders thirteen interactive, D3.js- and Cytoscape.js-based comparative visualizations, including a Rank-Rank Hypergeometric Overlap (RRHO) heatmap, a GO-slim "Radar/Spider" functional fingerprint, and chord/edge-bundled cross-talk diagrams that are, to our knowledge, not offered as an integrated set by any existing academic or commercial ORA/GSEA platform. GeneAutomate is an unfunded, individual student project developed with feedback from a professor, and is in its final stage of development. It requires no installation or login. We describe the tools architecture, statistical methods, and comparative feature set relative to established academic tools (DAVID, ShinyGO, g:Profiler, Enrichr, WebGestalt, STRING, PANTHER, GeneMANIA, Cytoscape, clusterProfiler, GSEA, Metascape) and commercial platforms (IPA, MetaCore, Pathway Studio, iPathwayGuide, Partek Pathway), and we state candidly the current versions limitations, which are planned to be the added in next version: single-species (human-only) coverage, no upstream regulator analysis, and comparison currently limited to two (occasionally three) concurrent lists. GeneAutomate is available at https://geneautomate.tech/.

bioinformatics↗

bioRxiv2026

A Charge-Encoded Rheostat Permits Helix Nucleation but Limits Propagation in Skp1

Molecular recognition by intrinsically disordered regions (IDRs) is widely thought to involve coupled folding and binding, yet the sequence features that regulate this transition remain underexplored. Here we show that helix 8 (H8), a disordered C-terminal segment of the SCF ubiquitin ligase adaptor Skp1, is intrinsically prevented from forming a stable helix by its own sequence grammar. Using an integrative approach to dissect its conformational dynamics, we find that H8 frequently nucleates helical structure but rarely propagates into a fully formed stable helix, populating instead a shallow metastable basin of helical intermediates. Contrary to conventional models of helix-coil exchange, where nucleation is rate limiting, helix initiation in H8 is readily accessible, while propagation is selectively suppressed by a glutamate-rich acidic patch. This acidic segment acts as a charge-sensitive conformational rheostat that limits helix extension and maintains H8 in a predominantly disordered state. As a result, H8 transiently samples partially helical conformations on the microsecond timescale without committing to a stable fold. We propose that this propagation-limited mechanism preserves conformational flexibility while maintaining recognition competence across a structurally diverse family of F-box binding partners. More broadly, our findings suggest that charged- hydrophobic-charged sequence patterning can encode conditional, context-dependent structure as a general organisational principle in intrinsically disordered proteomes.

biophysics↗

bioRxiv2026

Structural and Energetic Determinants of Monobody Recognition of Oncogenic KRAS Variants

Monobodies are engineered binding proteins that recognize extended protein surfaces and offer advantages over small-molecule inhibitors for targeting challenging KRAS oncoproteins. Monobody 12D4 exhibits high affinity and selectivity for the oncogenic KRAS(G12D) mutant, but the molecular determinants governing its recognition and the basis for its mutant selectivity remain poorly understood. Here, we combined molecular dynamics simulations and energy calculations to characterize the interactions between monobody 12D4 and WT KRAS as well as four clinically relevant oncogenic variants (G12C, G12D, G12V, and G12R) in both GTP- and GDP-bound states. Our simulations revealed that 12D4 recognition depends on a conserved hydrophobic interaction network centered on the monobody FG loop (residues L77, F78, and W79). This network forms stable contacts with KARS Switch II and 3-helix. The energy calculations also showed that residue K75 of 12D4 formed a mutation-specific electrostatic interaction with KRAS G12D. This interaction contributed significantly to the affinity of 12D4 toward this mutant, whereas this interaction was absent in other variants. No monobody currently exists for targeting KRAS G12R in either nucleotide state, and no monobody selectively targets KRAS G12C and G12V in the GDP-bound inactive state. To address these, we performed computational redesign at residues 75. We identified mutations (K75Q, K75Y, and K75M) that enhanced predicted binding to G12C, G12R, and G12V variants through reorganization of interfacial contacts. Our work establishes a structural framework for understanding KRAS-monobody recognition and provides a rational foundation for engineering variant-selective monobodies with improved affinity toward previously untargetable KRAS mutants. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/737552v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1e3cbf1org.highwire.dtl.DTLVardef@62684borg.highwire.dtl.DTLVardef@ad2ab0org.highwire.dtl.DTLVardef@cdc29a_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

A single chromosome 3p break initiates clear cell renal cell carcinoma evolution

Clear cell renal cell carcinoma (ccRCC) is initiated by chromosome 3p loss, yet chromosome losses impose a profound fitness burden on normal cells. How renal epithelial cells tolerate this deleterious aneuploidy during early tumorigenesis remains unclear. Analysis of 949 ccRCC genomes reveals two major classes of chromosome 3p alterations: simple deletions and complex rearrangements surrounding a terminal breakpoint - a pattern we term breakpoint-confined chromothripsis. We modeled both alterations in non-transformed human renal proximal tubule epithelial cells by introducing a single DNA double-strand break on chromosome 3p. Despite an initial fitness disadvantage, chromosome 3p loss drives adaptive genomic evolution that recapitulates recurrent ccRCC-associated aneuploidies, including 5q gain and 14q loss. These alterations alleviate the fitness constraints of 3p loss and promote metabolic reprogramming, clonal expansion, and malignant transformation, producing tumors with features of ccRCC. Thus, a single chromosome break initiates the evolutionary trajectory of ccRCC by creating a fitness bottleneck that selects for recurrent aneuploidies.

cancer biology↗

bioRxiv2026

The anaerobic cryo-EM structure of the methanogenic Mtr complex reveals a nitrogenase-like cluster bound to its active site

Methanogenic archaea conserve energy by coupling methyl-group transfer to the generation of a chemiosmotic sodiumion (Na+) gradient. This central energy-conserving step is catalyzed by the membrane-bound N5-methyl-H4MPT:coenzyme M methyltransferase (Mtr). Here, we present high-resolution cryo-electron microscopy structures of the Mtr complex from Methanosarcina mazei determined under strictly anaerobic conditions. The structures reveal an unexpected, electron-dense metallocluster embedded within the central cavity of the MtrCDE trimer in the membrane plane. Based on the unique topology and density we modeled it as an [Fe8S9C] L-type cluster. It is positioned adjacent to both the coenzyme M substrate and the corrinoid cofactor of MtrA in the MtrA-MtrCDE engaged state, thereby being located right at the catalytic core of the enzyme. We could further show that binding of MtrA to MtrCDE triggers rearrangements within the interface of MtrDE that widen a putative ion-conduction pathway. The proximity of the conserved sodium-binding site to the catalytic center suggests a putative link between methyl-transfer chemistry and Na+ translocation. In a broader context, these findings improve our understanding of how methyl transfer, analogous to redox chemistry, can drive chemiosmotic energy conversion.

biochemistry↗

bioRxiv2026

Simultaneous Functional Ultrasound, Intrinsic Optical Signal and Widefield Calcium Neuroimaging

Functional ultrasound (fUS) maps cerebral blood volume (CBV) but lacks molecular and neuronal specificity. By simultaneously integrating fUS with optical imaging, we show that fUS-derived CBV correlates with both optically measured hemoglobin and neuronal calcium activity in awake mice. We further derive hemodynamic response functions linking calcium activity to CBV during spontaneous and sensory-evoked activity. Application to a mouse glioblastoma model demonstrates utility for studying neurovascular dysfunction in complex neuropathologies.

neuroscience↗

bioRxiv2026

Matrix remodeling plays an etiological role in driving laminin-α2 deficient pathology

Laminin-2 (gene: LAMA2) is a key protein in the basement membrane of muscle and Schwann cells. A complete lack of this protein results in LAMA2-related congenital muscular dystrophy (LAMA2-RD), a severe muscle disease characterized by progressive muscle weakness, respiratory insufficiency, failure to thrive and shortened life span. One key signature of this disease is early onset of fibrosis coupled with poor muscle growth. We previously showed that TGF-{beta} and its activator, integrin-V, are elevated in dystrophic fibers of DyW mice, a mouse model of LAMA2-RD. Other than activating TGF-{beta}, integrin-V is also known to facilitate the transdifferentiation of various cell types to myofibroblasts. In this study we present evidence for transcriptional dysregulation of genes driving myofibroblast transdifferentiation and extracellular matrix (ECM) remodelling during the early development of DyW mice that is also reflected in muscle biopsies from young LAMA2-RD patients. We hypothesize that the early ECM remodelling, seen in both DyW mice and LAMA2-RD children, may explain the congenital onset of fibrosis with poor muscle growth seen in the disease. Summary StatementCharacterization of fibrogenic pathways that play potentially etiological roles in in LAMA2-related congenital muscular dystrophy

neuroscience↗

bioRxiv2026

CDK1 facilitates RAD51-mediated DNA repair to protect dictyate stage arrested oocytes from genotoxic stress

Oocytes arrested at the dictyate stage of meiosis I must maintain genomic integrity for prolonged periods to preserve female fertility. During this extended arrest, DNA lesions arising from endogenous and exogenous sources threaten oocyte survival, yet the molecular mechanisms coordinating DNA repair in dormant oocytes remain poorly understood. Here, we identify cyclin-dependent kinase 1 (CDK1) as a critical regulator of the oocyte DNA damage response and homologous recombination (HR) repair under genotoxic stress. Using cisplatin-induced DNA damage models in fetal goat ovaries and neonatal mouse ovaries, we investigated repair mechanisms operating within the ovarian reserve. Label-free proteomic profiling revealed significant enrichment of DNA damage response pathways following cisplatin exposure, with CDK1 emerging as one of the most prominently upregulated kinases. Pharmacological inhibition of CDK1 had little effect on follicle survival under physiological conditions but aggravated oocyte and follicle loss following DNA damage, indicating a stress-dependent role for CDK1 in preserving ovarian follicle pool integrity. Mechanistically, DNA damage activated a Chk2-dependent signaling pathway that promoted p63 phosphorylation and altered the WEE1-CDK1 regulatory axis, resulting in reduced inhibitory CDK1 phosphorylation (Thr14/Tyr15) and increased activating phosphorylation (Thr161). Activated CDK1 was associated with enhanced RAD51 phosphorylation and accumulation at DNA damage foci, supporting homologous recombination (HR)-mediated repair in dictyate-arrested oocytes. In contrast, CDK1 inhibition reduced phospho-RAD51 levels, impaired RAD51 localization, increased persistent {gamma}H2AX accumulation, and elevated oocyte apoptosis. Notably, suppression of CDK1 was accompanied by increased expression of the non-homologous end joining (NHEJ) marker Ku80 and the nucleotide excision repair (NER) factor XPA, suggesting increased engagement of alternative DNA repair pathways. Furthermore, inhibition of Chk2 abolished the DNA damage-associated CDK1 activation signature and restored WEE1 expression, supporting a model in which CDK1 functions downstream of Chk2 signaling during the oocyte DNA damage response. Collectively, our findings identify a previously unrecognized Chk2-CDK1-RAD51 signaling axis that coordinates homologous recombination repair in dormant oocytes and safeguards ovarian follicular pool integrity under genotoxic stress. These findings provide new mechanistic insight into how dictyate-arrested oocytes maintain genome stability during prolonged meiotic arrest.

Cell Biology↗

bioRxiv2026

Electron confurcation drives photosynthetic H2 production in cyanobacteria

Cyanobacteria are major contributors to global photosynthesis and are intensively studied for sustainable green H2 production. Central to this process is the bidirectional [NiFe]-hydrogenase HoxEFUYH, yet its physiological redox partners have remained unresolved. Ferredoxin, NAD(H), and NADP(H) have been proposed as partners, but the lack of active enzyme preparations has prevented a definitive assignment. Here, we purified the intact HoxEFUYH complex from Synechocystis sp. PCC 6803 under strictly anaerobic conditions and reveal its function as both a bifurcating and confurcating hydrogenase. During H2 uptake, HoxEFUYH utilizes NAD+ and oxidized ferredoxin, whereas H2 production strictly requires both NADH and reduced ferredoxin; NADPH does not support either reaction. Combining high-resolution cryo-electron microscopy with biochemical and spectroscopic analyses, our data reveal that an flavin-containing reductase module is electronically connected to the catalytic [NiFe]-hydrogenase core through an extended chain of iron-sulfur clusters, defining the structural basis for bifurcating and confurcating electron flow. These findings fundamentally revise the physiological role of HoxEFUYH by showing that photosynthetic H2 production does not rely solely on photosynthetic electrons but instead couples reduced ferredoxin from the light reaction with NADH derived from "dark" carbohydrate oxidation. This requires reassessment of current strategies for green H2 production in cyanobacteria.

biochemistry↗

bioRxiv2026

Basal BNIP3/NIX mitophagy is controlled by selective protection of sentinel receptors

Mitochondrial homeostasis is maintained by multiple quality control pathways, including mitophagy, which targets dysfunctional mitochondria for degradation. During receptor-mediated mitophagy, the outer membrane proteins BNIP3 and NIX directly recruit autophagy machinery to the mitochondrial surface, though their precise regulation is still unclear. In recent years, new BNIP3- and NIX-interacting proteins have been identified that influence mitophagic flux. PPTC7 and FBXL4 target BNIP3 and NIX for proteasomal turnover to keep levels of the receptors low, whereas TMEM11 is proposed to spatially control mitophagy by interacting with receptors at active mitophagy sites. However, it is unclear how each of these interactions is controlled and how they interplay with each other. Here, we identify a repressor of mitophagy, ARMC1, which forms a complex with TMEM11, BNIP3, and NIX. During mitophagy activation, ARMC1 dissociates from the complex, freeing the receptors to initiate mitophagy. We find that TMEM11 then acts in an antagonistic relationship with PPTC7, protecting the receptors from proteasomal degradation. Our data are consistent with a two-stage model. At steady state, a population of sentinel receptors is repressed and primed to respond to mitochondrial dysfunction. Once mitophagy is activated, TMEM11 protects BNIP3 and NIX, ensuring a sustained mitophagic response. Our findings provide a framework for understanding how two key regulatory pathways intersect to modulate receptor-mediated mitophagy.

Cell Biology↗

bioRxiv2026

The structure of a 2-MDa chloroplast RNA polymerase reveals unexpected evolutionary complexity

Transcription in chloroplasts depends on the Plastid-Encoded RNA polymerase (PEP), a bacterial-derived enzyme whose catalytic core remains encoded by the highly reduced genome inherited from the cyanobacterial ancestor. In land plants, PEP has roughly doubled in size, expanding into a [~]1 MDa multisubunit machinery through the acquisition of numerous nuclear-encoded subunits. Based on phylogenetic analyses, this added complexity has been widely attributed to the demands of plant terrestrialization. Contrary to this view, we show that in the unicellular green alga Chlamydomonas reinhardtii, PEP assembles into an even larger [~]2 MDa complex containing twelve previously uncharacterized nuclear-encoded subunits (PEPS1-12), representing an RNA polymerase architecture of unprecedented size. A cryo-EM structure at 2.7 [A] resolution reveals that several of these subunits occupy positions analogous to those in land plant PEP, and that metabolic enzyme folds have been repurposed as structural scaffolds stabilizing the highly expanded plastid-encoded core. Despite this, most of the newly identified PEPS subunits lack detectable sequence or structural similarity to their land plant counterparts. These findings demonstrate that PEP complexity is not a hallmark of land plant evolution and may instead reflect, at least in part, the evolutionary entrenchment of additional subunits around an expanded plastid-encoded core. More broadly, they suggest that essential organellar machines can acquire substantial structural complexity that leaves little trace in sequence-based analyses, a pattern consistent with constructive neutral evolution.

molecular biology↗

bioRxiv2026

piRNAs from Y chromosomal protein coding, noncoding and endogenous retrovirus homologous repeat families regulate autosomal gene expression in mouse testis

BackgroundHeterochromatic long arm of mouse Y chromosome harbors the multicopy species-specific sequences Ssty, Sly, Asty and Orly that are transcribed in testis and have known functions in male fertility. Of these Ssty and Sly encode proteins - yet all the transcripts are not translated. To investigate the roles of these Y-heterochromatic transcripts further, we analyzed them. MethodsMice with 2/3rd deletion of the Y-chromosome (XYRIIIqdel) and its wild type (XYRIII) were used in this study. Bioinformatic approaches, small RNA northern blots, Electrophoretic Mobility Shift Assays, Luciferase reporter assays, dPCR analysis, RT-qPCR assays and western blotting techniques were used to identify piRNAs that regulate autosomal genes. ResultsWe demonstrate that the multicopy gene families from mouse Y-long arm generate piRNAs predominantly in testis. We observed sequences homologous to these piRNAs in the UTRs of a few autosomal genes, which are differentially expressed in the sperms of XYRIIIqdel mice. Furthermore, the Endogenous Retrovirus Element (ERV) LTR, found in the Orly1 transcript identified piRNAs in the database, showed homology to UTRs and associated genomic regions of a few autosomal genes. Orly1 showed a reduction in genomic copy number by digital PCR in XYRIIIqdel mice. One of the four autosomal genes containing the ERV segment in their UTRs, showed a differential testicular protein expression in the mutant mice. ConclusionsThus, we further elucidate that different classes of repeats from Y-chromosome regulate autosomal gene expression via piRNAs. Besides, this study also identified novel roles for a Y-derived ERV in autosomal gene regulation in testis.

Molecular Biology↗

bioRxiv2026

Inhibitory Gain and Hub Architecture Confer Dynamic Resilience to Microcircuit Degeneration

Neurodegeneration progressively removes synapses and neurons, yet neural circuits can retain stable collective dynamics despite substantial structural loss. Which structural principles confer this resilience remained unclear. Using large-scale spiking networks spanning empirical and synthetic microcircuit architectures, we systematically compared synaptic and neuronal modes of degeneration under controlled pruning strategies. We found that resilience was not determined by connectivity loss alone, but by how inhibitory gain was embedded within circuit architecture. Networks in which inhibitory neurons occupied structurally central positions robustly maintained health-like firing rates, levels of synchrony, and informational bandwidth across degeneration stages, whereas architectures lacking such embedding exhibited amplified dynamical disruption. Across regimes, the evolution of activity was organized by a compact set of weight-aware structural descriptors that generalized across network sizes and classes, with total effective synaptic coupling providing a dominant organizing axis. These results identified inhibitory architecture as a mechanistic determinant of circuit resilience and provided a predictive framework linking structural degeneration to collective dynamics.

neuroscience↗

bioRxiv2026

Lipid Droplets Direct Amyloid Assembly and β-Cell Stress through Interfacial Control of Amylin Aggregation

Islet amyloid polypeptide (IAPP, amylin) aggregation is a central pathological feature of type 2 diabetes, yet the cellular factors governing its conformational conversion remain incompletely understood. Here, we identify lipid droplets (LDs) as active modulators of amylin structure, aggregation, and {beta}-cell stress. Using artificial and native LDs, we show that amylin binds LD surfaces with high affinity and undergoes accelerated conversion into {beta}-sheet-rich conformations. LDs promote rapid nucleation while constraining fibril elongation, yielding shorter and morphologically distinct amyloid assemblies. Residue-resolved NMR mapping reveals a conserved N-terminal interaction interface, which is amplified upon removal of LD surface proteins, indicating that the LD proteome modulates peptide engagement and aggregation pathways. In {beta}-cells, lipid loading drives intracellular colocalization of amylin with LDs and reshapes transcriptional stress responses, attenuating ER stress and apoptosis while altering markers of {beta}-cell identity. Finally, systemic lipidomic profiling reveals coordinated remodeling of neutral lipid species across dysglycaemic states, linking intracellular LD dynamics with whole-body lipid metabolism. Together, our findings establish lipid droplets as dynamic scaffolds that reshape amylin aggregation pathways and associated {beta}-cell stress responses, providing a mechanistic bridge between lipid dysregulation and islet amyloidosis in diabetes. Significance StatementLipid droplets accumulate in pancreatic {beta}-cells during metabolic stress, yet their role in amylin aggregation remains unclear. Using structural, biophysical, and cellular approaches, we show that lipid droplet interfaces directly bind human amylin, reshape its aggregation pathway, and alter fibril morphology. Native lipid droplets and their associated surface proteins further modulate aggregation kinetics and peptide conformations. In {beta}-cells, lipid loading enhances amylin colocalization with lipid droplets and modifies stress and survival responses. Complementary lipidomic profiling reveals systemic remodeling of neutral lipid species across dysglycaemic states. These findings identify lipid droplets as active regulators of amyloid-associated proteostasis, linking lipid dysregulation to {beta}-cell dysfunction in type 2 diabetes.

Cell Biology↗

bioRxiv2026

Estrogen-related receptor signaling counters sarcopenia and preserves exercise fitness in naturally aged mice

BackgroundEstrogen-related receptor gamma (ERR{gamma}) drives an exercise mimicking aerobic gene program in the skeletal muscle that could be beneficial in aging. We have investigated the effect of chronic ERR{gamma} activation on minimizing sarcopenia. MethodsExperiments were performed in muscle specific ERR{gamma} transgenic (TG) mice and wild type (WT) littermates, at young (4-5 months) and old (24-26 months) age. In the skeletal muscle, global gene expression changes, as well as myofiber histological changes in fiber type, size, vascular supply and neuromuscular junction (NMJ), and mitochondrial content were measured. Functional analysis was performed using in vivo muscle contraction assay. Exercise fitness was measured using treadmill sprint and endurance test. Gene and protein expression was measured using QPCR and Westerns, respectively. ResultsERR{gamma} activates a pan-ERR aerobic program in the skeletal muscle to increase expression of 574 genes including ERR, mitochondrial homeostasis (e.g. Mfn1, Opa1, Drp1, Fis1, and Tfam), vascularization (e.g. Vegfa, Angpt1, Fgf1), and neuromuscular junction (NMJ) (e.g. Nrp1, Aspa, Ptprm, Cxcr4), simultaneously suppressing the expression of atrophy related genes (e.g. Atrogin1, Traf6, Nedd4, Myd88, p21). ERR{gamma} increases mitochondrial content [Mitochondrial area: old TG vs. WT, 2.00 fold; young TG vs. WT, 1.32 fold], oxidative capacity [NADH-TR activity: old TG vs. WT, 1.20 fold; young TG vs. WT, 1.22 fold] and myofiber type [2a: old TG (687{+/-}258) vs. WT (252{+/-}71); young TG (797{+/-}168) vs. WT (440{+/-}76); 2x: old TG 1348{+/-}87 vs. WT 976{+/-}219; young TG 1131{+/-}135 vs. WT 936{+/-}84; 2b: old TG (798{+/-}103) vs. WT (1628{+/-}148); young TG (967{+/-}133) vs. WT (1623{+/-}189)], and capillarity [capillary-to-myofiber ratio: old TG (3.25{+/-}0.19) vs. WT (2.41{+/-}0.16); young TG (3.41{+/-}0.21) vs WT (2.59{+/-}0.2)] and [NMJ number [old TG (67{+/-}8) vs. WT (40{+/-}9); young TG (77{+/-}11) vs WT (77{+/-}7)], mitigating age-related loss of NMJ and myofiber cross-sectional area [old TG (1570{+/-}147{micro}m2) vs. WT (1692.5{+/-}208{micro}m2) WT; young TG (1828.15{+/-}132.8{micro}m2) vs. WT (2109.7{+/-}296.8{micro}m2)]. ERR{gamma} overexpression preserves muscle contractility with aging [Fatigue resistance: 22.72% reduction in force in old vs. young WT; 3.11% reduction in force between old vs. young TG]. Furthermore, ERR{gamma} maintains exercise fitness in old mice [Running: old TG (2964.52{+/-}405m) vs. old WT (910.75{+/-}6034m); young TG (2232.43{+/-}193.64m) vs. young WT (1366.76{+/-}60.76m)]. ConclusionsERR{gamma} drives a pan-ERR and counter sarcopenic gene program enhancing oxidative myofiber type, mitochondrial content, vasculature, and NMJ in aging muscle. Consequently, ERR{gamma} minimizes myofiber atrophy, preserves contractility, and improves exercise fitness in old mice. Therefore, ERRs are potential translational targets for combating sarcopenia.

physiology↗

bioRxiv2026

Hepatic Differentiation of Human Pluripotent Stem Cells into Functional In Vitro Models Recapitulating Native Liver Complexity for MASLD Modelling

Human in vitro hepatic models that accurately recapitulate liver function are essential for fundamental and translational research; however, currently utilised models for disease modelling and drug discovery lack physiological fidelity and require prolonged culture time. Here, we present a streamlined 10-day protocol for efficient and reproducible differentiation of human pluripotent stem cells into hepatocyte like cells (HLCs) and hepatic liver organoids (HLOs). Both models exhibited mature hepatocyte differentiation, as evidenced by albumin secretion and CYP3A4 metabolic activity. Interestingly, HLOs display enhanced multicellular complexity incorporating endothelial, stellate, and macrophage populations along with hepatocytes, thereby more closely recapitulating the native liver microenvironment than HLCs. Here, steatosis was induced in both platforms, which resulted in triglyceride accumulation and upregulated lipogenic markers (DGAT1, DGAT2). However, only HLOs recapitulated advanced disease characteristics, including inflammatory (IL-10) and fibrotic (alpha SMA, COL1A1) responses. Resmetirom, a thyroid hormone receptor-{beta} agonist, significantly reduced steatosis and restored molecular signatures in both models. Additionally, transplanted HLOs demonstrated prolonged survival and displayed host-derived vascularization, thereby validating in vivo maturation. Collectively, HLCs provides a rapid and physiologically relevant liver model, with HLOs offering superior utility for disease modelling, therapeutic evaluation, and regenerative applications due to their enhanced functional and physiological relevance.

cell biology↗

bioRxiv2026

Hypothalamic MCH neurons links tau pathology to sleep disruption

Sleep disruption is an early and pervasive feature of Alzheimers disease (AD), yet the circuit mechanisms linking tau pathology to sleep-wake dysfunction remain unresolved. Here, we identify melanin-concentrating hormone (MCH) neurons in the lateral hypothalamus (LH) as a critical node disrupted in tauopathy. Longitudinal EEG/EMG recordings in PS19 mice reveal progressive impairments in sleep architecture and homeostasis. Histological analyses revealed significant degeneration of both MCH neurons and the neighboring hypocretin (Hcrt) neuronal population in the LH at late stages of tauopathy. In vivo fiber photometry recordings demonstrated a selective functional impairment of MCH neurons, characterized by reduced activity during REM sleep, whereas Hcrt neuronal activity remained largely preserved. In addition, cell-autonomous expression of mutant tau in MCH neurons recapitulates sleep disruption, establishing a causal role. Despite tauopathy-induced neuronal loss and reduced endogenous activity, optogenetic and chemogenetic activation show that MCH neurons retain functional capacity, and their activation restores sleep in aged PS19 mice. Together, these findings define a circuit mechanism linking tau pathology to sleep and identify MCH neurons as a tractable therapeutic target.

neuroscience↗

bioRxiv2026

In silico characterization of unique fungal modular rhodopsin expands the horizon of novel optobiological and biomedical applications

Microbial modular rhodopsins, in which light-sensing rhodopsin domains are fused with effector modules, have emerged as promising tools for optogenetic regulation in algae and other systems. However, the diversity and potential regulatory roles of fungal modular rhodopsins remain largely unexplored. Here, we performed a comprehensive in-silico analysis to identify previously uncharacterized fungal modular-rhodopsins that pair a conserved light-sensing core with diverse effector domains, including RPEL-motif, NADP-binding Rossmann fold domain, MCM (Mini-Chromosome Maintenance) domain, and GC-cAT (Carnitine O-Acetyltransferase) modules. In Aureobasidium pullulans, the representative modular rhodopsin (ApRh-RPEL) contains RPEL-motif associated with actin-related and transcriptional regulatory processes, suggesting light-driven fungal signaling pathway involved in transcriptional and cellular regulation, respectively. Rhodopsins fused with NADP-binding Rossmann fold and MCM domains further indicate possible applications in light-programmable metabolic and cell-cycle signaling. Genome mining additionally revealed that A. pullulans harbours a diverse but underexplored array of biosynthetic gene clusters (BGCs), raising the intriguing possibility that light perception may regulate secondary metabolite pathways. Supporting this, multisource protein-protein interaction network analysis links ApRh-RPEL to enzymes involved in terpenoid and sphingolipid biosynthesis, indicating potential cross-talk between light-sensing module and metabolic regulation. These findings outline a computationally derived model in which fungal modular rhodopsins (ApRh-RPEL) function as opto-synthetic regulators of biosynthetic processes. Structural predictions confirmed conserved Schiff-base lysine and retinal-binding pocket, highlighting functional diversity across fungal rhodopsins. Together, these findings expand the optogenetic toolkit and provide a framework for engineering light-driven signaling in fungi, with applications in optobiological and biomedical applications.

bioinformatics↗