Biology subjects

Gupta, S.

Publications and source records attributed to Gupta, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2026

Ancient human genomes from Ladakh reveal Tibetan, South Asian, and Central Asian admixture over the last three millennia

The trans-Himalayan region of Ladakh occupies a strategic position at the crossroads of South Asia, the Tibetan Plateau, and Central Asia, with archaeological evidence pointing to long-term cultural exchanges across these regions. However, the human genetic history of Ladakh remains largely unexplored. We generated paleogenomic data from seven individuals recovered from two sites in Western Ladakh - the Old Lady Spider Cave and Hanu - of which six are dated to 531-585 CE and one to the 19th century CE. The older individuals share substantial genetic ancestry with Tibetan groups but also harbor major contributions from two additional sources: one corresponding to the currently-oldest observation of the Ancestral North Indian genetic component that characterizes several present-day populations in North India and Pakistan, and another related to ancient Central Asian groups, with admixture events occurring between [~]2,100-2,500 years ago. In contrast, the later individual falls within a previously described ancient northern Himalayan genetic cline based on [~]1,100-1,300-year-old individuals from Himachal Pradesh, with ancestries related to ancient Tibetan and Steppe-related sources. Stable isotope analysis suggests that these individuals were local to Ladakh in late life and practiced an agro-pastoralist subsistence. Our study establishes that Ladakhs central role in Eurasian economic and socio-cultural networks was shaped by dynamic and sustained gene flow linking high-altitude Himalayan groups with both lowland South Asia and Inner Asia.

genetics↗

bioRxiv2026

BiomarkerKB: FAIR and Integrated Biomarker Knowledge Connecting Biomolecular and Clinical Data Types

Biomarkers are essential tools for disease detection, risk assessment, therapeutic monitoring, and precision medicine. However, biomarker data are dispersed across heterogeneous resources, inconsistently reported in the literature, and rarely standardized for computational use. This fragmentation limits reproducibility, cross-study integration, and the discovery of novel biomarker and disease relationships. We developed BiomarkerKB, a knowledgebase designed to harmonize and integrate biomarker information under a standardized data model. The model follows the FDA-NIH BEST biomarker definition and captures both core fields (biomarker entity, disease/condition, exposure agent) and contextual metadata (specimen, biomarker role, evidence, provenance). Biomarker data and related annotations were either curated from publications or collected from public resources (e.g., OpenTargets, GWAS Catalog, ClinVar, CIViC, OncoMX) and were also contributed by the Common Fund Data Coordinating Centers and the Early Detection Research Network (EDRN). Standardization was achieved using ontologies and reference resources such as Disease Ontology, UBERON, UniProtKB, and HUGO Gene Nomenclature Committee (HGNC) gene symbols. BiomarkerKB data were ingested into a Neo4j-based knowledge graph and integrated with the Common Fund Data Ecosystem (CFDE) Knowledge Graph. The initial release of BiomarkerKB contains over 200,000 biomarker-disease associations spanning genes, proteins, metabolites, glycans, and chemical elements. The knowledge graph comprises more than 300,000 nodes and 1.2 million edges, enabling structured exploration of biomarker relationships within CFDE data as demonstrated through the knowledge graph query-based use cases presented in this study. A publicly accessible web portal (https://biomarkerkb.org) provides keyword search, filtering, data downloads, and access to graph visualization to support both researchers and computational analyses. BiomarkerKB addresses a critical gap in biomarker informatics by providing an integrated, FAIR (Findable, Accessible, Interoperable, and Reusable), and unified framework for biomarker knowledge exploration and discovery.

bioinformatics↗

bioRxiv2026

Reprogramming BCMA-Targeted CAR-T Cells through γ-Secretase Modulation Blocks Antigen Shedding and Extends CAR-T Longevity

B-cell maturation antigen (BCMA) shedding by {gamma}-secretase generates soluble BCMA (sBCMA), which diminishes membrane antigen density, and limits the durability of BCMA-directed immunotherapies in multiple myeloma (MM). Here, using AI-driven diffusion modeling, we identify peptide inhibitors that selectively block {gamma}-secretase-mediated BCMA cleavage, stabilizing membrane-bound BCMA (mBCMA) without compromising cellular viability. The lead peptide, P5, suppresses sBCMA and restores mBCMA in vitro and in xenograft models. To create sustained, cell-intrinsic inhibition, we engineered Shedding Deterrent Locked-in (SHEDLOCK) CAR-T cells that locally secrete {gamma}-secretase-modulatory peptides. SHEDLOCK-1 CAR-T cells secreting P5 exhibits enhanced cytotoxicity and persistence relative to conventional CAR-T cells. Building on this, SHEDLOCK-2 CAR-T cells were generated, which secrete a modified, naturally derived peptide (nxP) that directly engages the {gamma}-secretase catalytic site, concurrently preventing BCMA shedding, and surprisingly enhancing CAR-T cell longevity by preserving telomere integrity and metabolic fitness. In MM xenograft and patient-derived models, SHEDLOCK-2 CAR-T cells demonstrate durable antitumor activity with a favorable safety profile. Together, these findings establish SHEDLOCK as a next-generation CAR-T platform and provide a strong preclinical foundation for Phase I clinical evaluation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/700575v1_ufig1.gif" ALT="Figure 1"> View larger version (69K): org.highwire.dtl.DTLVardef@3cd38aorg.highwire.dtl.DTLVardef@164167borg.highwire.dtl.DTLVardef@1ffed21org.highwire.dtl.DTLVardef@1294d06_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2026

Distinct trajectories of urbanization shape the human gut microbiome across South Asia

Human gut microbiomes respond to lifestyle transitions, yet the extent to which these responses are conserved across spatio-cultural contexts remains undercharacterized. We present the South Asian MicroBiome ARray (SAMBAR), a population-scale 16S gut microbiome study of 575 adults from ten geographically and socio-culturally diverse South Asian communities. Each community was sampled in ancestral villages and urban centers, enabling controlled comparisons of geography and lifestyle. Relative to global cohorts, SAMBAR microbiomes occupy a distinct compositional space with stronger correlation to geography and community membership than lifestyle. Although urbanization is consistently associated with increased abundance of disease-linked taxa, microbiome responses to lifestyle transitions are largely community-driven, including the acquisition of wheat- and dairying-associated microbial modules in some communities that may facilitate non-genetic adaptation to lactase non-persistence. Microbiome responses to urbanization are heterogeneous even at regional scales, reflecting local culture and geography and underscoring the need for community-specific investigations of health impacts.

microbiology↗

bioRxiv2026

An Indole Dimer Antifungal Metabolite from a Rice Seed Endophyte Inhibits Ergosterol Biosynthesis in Fungal Pathogens

The increasing prevalence of fungal phytopathogens and the widespread emergence of fungicide resistance necessitate alternative antifungal strategies with reduced environmental impact. Here, we report the isolation and characterization of a novel antifungal metabolite, SM06, produced by the rice seed-associated endophytic bacterium Phytobacter sp. RSE02. SM06 exhibited broad-spectrum antifungal activity against plant and human pathogenic fungi, including Curvularia lunata, Fusarium oxysporum, and Candida albicans. In vitro assays and micromorphological analyses revealed that SM06, an indole dimer, disrupts fungal cell membrane integrity, while in planta experiments demonstrated significant suppression of brown leaf spot disease in tomato and rice. Molecular docking suggested that SM06 binds to lanosterol 14-demethylase (ERG11), a key enzyme in fungal sterol biosynthesis. Consistent with this prediction, LC-MS-based analyses confirmed a significant reduction in ergosterol content in SM06-treated fungal cells. Together, these findings identify SM06 as a biologically active antifungal metabolite produced by a plant-associated bacterium and highlight its potential application in sustainable fungal disease management. IMPORTANCEFungal diseases cause major losses in crop-production and contribute to the growing challenge of antifungal resistance, underscoring the need for sustainable alternatives to chemical fungicides. This study identifies SM06, a novel indole dimer produced by the rice seed endophyte Phytobacter sp. RSE02, with strong antifungal activity against economically important plant pathogens and clinically relevant fungi. Through integrated chemical, cellular, and in planta analyses, we demonstrate that SM06 disrupts fungal membrane integrity by inhibiting ergosterol biosynthesis. The compound is biocompatible, stable, and effective in plant disease suppression, highlighting its translational potential for crop protection. These findings reveal seed endophytes as an important yet underexplored sources of antifungal metabolites and provide a mechanistic foundation for developing eco-friendly biocontrol strategies with implications beyond agriculture. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC="FIGDIR/small/697688v1_ufig1.gif" ALT="Figure 1"> View larger version (86K): org.highwire.dtl.DTLVardef@c7da68org.highwire.dtl.DTLVardef@d3f1a1org.highwire.dtl.DTLVardef@9cab82org.highwire.dtl.DTLVardef@1c01141_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2025

STAT1-Mediated Regulation of IL-17A/CEBPB/NF-κB Axisin HIV-1 Infected Human Cerebral Organoids RevealsTherapeutic Targets for Neuroprotection

HIV-associated neurological complications remain a major concern in people living with HIV (PLWH), even under effective viral suppression with combination antiretroviral therapy (cART), underscoring unresolved mechanisms driving HIV-related neurodegeneration. To address this gap, we used HIV-1-infected human cerebral organoids (hCOs) containing microglia as a physiologically relevant three-dimensional model of the central nervous system (CNS) and validated key findings in simian immunodeficiency virus (SIV)-infected, cART-treated rhesus macaque model. Integrating single-nucleus RNA sequencing with ATAC sequencing, we identified cell-type-specific alterations in dual innate immune-driven inflammatory signaling pathways across neural and glial populations. Microglia were preferentially infected and activated, initiating IL-17A-mediated cascades involving IFN-{gamma}/STAT1, CEBPB, and NF-{kappa}B pathways, which converged with cGAS-STING and Notch-NEURL1 signaling, both associated with neuroinflammation and synaptic dysfunction. cART suppressed viral replication, reduced IL-17A-driven neuroinflammation, improved mitochondrial function, and restored synaptic gene expression. Collectively, these findings identify IL-17A, STAT1, CEBPB, and cGAS-STING as key molecular drivers of HIV-associated neuroinflammation and establish a novel mechanistic link between IL-17A signaling and microglial remodeling, providing a translational framework for the development of CNS-targeted therapies in PLWH.

neuroscience↗

bioRxiv2025

Rutin derived microbiota metabolite 3,4 dihydroxybenzoic acid restores antibiotic susceptibility in XDR Gram negative pathogens with a system wide susceptibility signature

Antibiotic potentiators are a practical route to extend the utility of existing drugs against multidrug resistant Gram-negative pathogens, but most natural compound combinations remain mechanistically under defined and are rarely mapped at the level of whole cell susceptibility determinants. Here we apply a metabolite guided potentiator strategy based on gut microbiota derived breakdown products of the dietary polyphenol rutin, and we integrate potentiation phenotyping with system wide genetic susceptibility mapping to guide combination design. Three rutin metabolites, 3,4 dihydroxybenzoic acid (DHBA), 2,4,6 trihydroxybenzoic acid (THBA), and 3,4 dihydroxyphenylacetic acid (DOPAC), were screened against a CDC ARLG reference panel composed of multidrug resistant and extensively drug resistant (XDR) isolates, identifying DHBA as the most broadly active candidate, including against XDR Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli. DHBA restored susceptibility to selected antibiotic classes in resistant Gram-negative pathogens, including an approximately sixteen-fold reduction in colistin MIC in mcr 1 positive E. coli. To better understand the potentiation mechanism, we performed a large-scale genetic susceptibility screen of 316 E. coli single gene knockouts and defined a DHBA response architecture enriched for envelope and transport determinants with additional contributions from central metabolism and information processing pathways. Comparative mapping under colistin exposure revealed a distinct susceptibility architecture with limited overlap, supporting the concept that both potentiators and antibiotics engage secondary cellular systems beyond canonical primary mechanisms. In a Mini Bioreactor Array gut community model, DHBA produced a more conserved community shift than colistin. Finally, DHBA-antibiotic combinations improved outcomes in infection relevant models, including improved survival in Galleria mellonella, reduced intestinal burden in Caenorhabditis elegans, and reduced bacterial burden in an ex vivo porcine burn wound infection model. Collectively, these findings support a systems-based framework for developing mechanistically informed potentiator antibiotic-combinations to extend the lifespan of existing antibiotics.

microbiology↗

bioRxiv2025

In vivo human embryonic spinal cord atlas validates stem cell-derived human dorsal interneurons and reveals ASD spinal signatures

Restoring somatosensory function after spinal cord injury (SCI) faces fundamental challenges: neuronal subtypes must match both axial position and circuit identity, yet the developmental patterning of human dorsal spinal interneurons (dIs) remains incompletely defined. Here, we integrate six single-cell transcriptomics datasets derived from human embryonic spinal cord tissue spanning gestational weeks 4-25 to generate a reference atlas of early human somatosensory circuit development. The atlas reveals molecular signatures underlying expansion and specialization of dI4 and dI5 interneuron populations associated with mechanosensory and nociceptive processing. Guided by this resource, we established a neuromesodermal progenitor-based differentiation approach that generates dorsal interneurons spanning anterior-posterior identities. Comparison of in vivo and in vitro dI4/dI5 subclasses identified conserved gene networks associated with sensory modalities and revealed enrichment of autism spectrum disorder-associated genes within mechanosensory interneuron populations. Together, these findings clarify how human dorsal spinal interneuron diversity is established.

neuroscience↗

bioRxiv2025

Mapping Gene Impact on Single-cell Transcriptomic Networks via Perturbation Response Scanning

Gene knockout experiments are essential for dissecting gene function, and CRISPR has made targeted gene disruption more accessible than ever. Single-cell CRISPR screening enables the construction of rich genetic perturbation landscapes, facilitating the identification of genes whose perturbation strongly reshapes cellular states. However, due to the nonlinear dependencies within gene networks, identifying the most impactful tangible genes remains challenging. Existing virtual knockout methods estimate downstream effects of single-gene deletions but do not evaluate whether such perturbations disrupt global information flow or compromise cellular robustness. To address this limitation, we adapt a perturbation-response framework originally developed for protein structural dynamics to identify gene modules most susceptible to perturbation. We introduce the single-cell Perturbation Impact Index (scPII), a data-driven metric derived from gene regulatory networks that quantifies system-level responses to gene perturbations, without using any CRISPR screening information. Our results demonstrate that scPII effectively identifies genes whose perturbation has the greatest system-wide impact. Analysis of single-cell RNA-seq data from the Cancer Cell Line Encyclopedia revealed a strong correlation between scPII-derived gene impact scores and gene effect scores from genome-wide CRISPR screens. These findings indicate that scPII provides a robust metric for quantifying gene knockout effects. More broadly, integrating perturbation response scanning with gene regulatory networks offers a powerful framework for advancing single-cell data analytics in biomedical research.

bioinformatics↗

bioRxiv2025

Global Hypomethylation in Cell-free DNA Enables Non-invasive Colorectal Cancer Screening: Results from a Retrospective Validation Study

Global DNA hypomethylation is a defining hallmark of colorectal cancer (CRC) but is poorly captured by existing cell-free DNA (cfDNA) technologies, which typically interrogate only a fraction of CpG sites and are biased toward CpG islands. We developed Asima Rev, an electrical-impedance cfDNA assay that can differentiate healthy individuals from those with cancer by measuring cfDNA aggregation patterns associated with methylation state, enabling functional detection of genome-wide hypomethylation. In a cohort of 46 treatment-naive CRC patients and 33 controls, Asima Rev achieved 96% sensitivity and 94% specificity, with longitudinal monitoring in six patients fully concordant with clinical outcomes. Interrogation of 11 public methylation array datasets showed that whole array analyses underestimate global changes. However, restricting analyses to OpenSea regions, where cfDNA is enriched, revealed patterns consistent with Asima Rev and a significant 5.8% global hypomethylation in CRC tissue compared to adjacent normal tissue. Hypomethylation was not observed in immune cell genomic DNA (gDNA), a major contributor to cfDNA, supporting a predominantly tumor-derived contribution to the observed cfDNA signal. Together, these results demonstrate that Asima Rev captures a cfDNA signal consistent with true global methylation loss and outperforms locus-specific assays by measuring structural consequences of pan-genomic epigenetic alterations.

bioinformatics↗

bioRxiv2025

Encoding and Decoding of Brain Dynamic Functional Connectivity for ADHD Diagnosis

Recent studies have demonstrated strong associations between the changes in dynamic functional connectivity (FC) and both behavioral and cognitive functions. The sliding window technique is the most widely used method for evaluating dynamic FC; however, it faces two key challenges: distributional shifts across windows and high dimensionality, as FC is computed across windows of the entire time series. To address these issues, we propose BRAINMAP (Bi-level Representation using Attention for INterpretability with Mamba-Aided Prediction) to model the dynamic FC of the brain. BRAINMAP employs the Optimal Transport technique to correct distributional shifts across sliding windows and leverages Graph Neural Networks (GNNs) in conjunction with a hybrid approach that integrates an attention mechanism and the Mamba block to effectively capture spatiotemporal features for functional MR images. Finally, we introduce a novel Top-K sliding window feature selection algorithm to induce sparsity in dynamic FC. We conducted an extensive evaluation of our model for diagnosing Attention Deficit Hyperactivity Disorder (ADHD) using three resting-state fMRI datasets: ADHD-200, UCLA, and CNI-TLC, which comprise a total of 447 subjects with ADHD and 845 typically developing controls. Our architecture outperformed existing state-of-the-art dynamic FC models in ADHD detection, achieving improvements ranging from 3% to 12% across the three datasets. We demonstrate that our proposed model produces robust biomarkers, most notably the connection between the dorsal attention network and the visual network. Using an association study, we further establish the clinical relevance of the identified biomarkers.

neuroscience↗

bioRxiv2025

Integrative Transcriptomics and Phytochemical Screening Reveal Pratenol B, Eriodictyol, Losbanine, and Isookanin, as Potential EGFR and HRAS Inhibitors in Indian Oral Squamous Cell Carcinoma Patients

Oral squamous cell carcinoma (OSCC) is the most common head and neck cancer, with India contributing nearly one-third of the global cases. Management of OSCC remains difficult due to increasing risk factors, limited therapeutic options, severe side effects, and rising drug resistance. Therefore, novel and safer treatment strategies are urgently needed. This study explores the potential of phytochemicals as targeted inhibitors of key dysregulated biomarkers in Indian OSCC patients. RNA sequencing and pathway analysis revealed significant alterations in the MAPK signaling pathway, highlighting EGFR and HRAS as crucial therapeutic targets. Given the limited clinical success of existing EGFR-targeted therapies and the scarcity of HRAS inhibitors, a natural product-based approach was adopted. Molecular docking of 17,000 phytochemicals identified Pratenol B, Eriodictyol, Losbanine, and Isookanin as promising inhibitors, with Pratenol B showing dual inhibition of EGFR and HRAS. These compounds exhibited strong binding affinities, favorable pharmacokinetic profiles, high bioavailability, and low toxicity. Molecular dynamics simulations confirmed the stability of Pratenol B with both target proteins, surpassing reference inhibitors. Utilizing vast medicinal plant diversity presents a cost-effective and low-toxicity avenue for OSCC therapy. Further in vitro, in vivo, and clinical studies are warranted to validate these phytochemicals as potential therapeutics.

cancer biology↗

bioRxiv2025

Upregulation of a novel lncRNA KLHDC7B-DT in Head and Neck Cancers: Implications for Prognosis and Molecular Mechanisms

Head and neck cancer (HNC), the seventh most frequent malignancy globally, is a serious health concern. Despite advancements in treatment, the overall survival rate for HNC remains around 50%, with even poorer outcomes in metastatic cases. This highlights the urgent need to explore the molecular mechanisms underlying HNC for improved diagnostics and therapies. The importance of non-coding RNAs, especially long non-coding RNAs (lncRNAs), in a variety of malignancies, including HNC, has been highlighted in recent researches. Although lncRNAs and mRNA are very similar, lncRNAs do not encode proteins and they are longer than 200 nucleotides. While studies have implicated lncRNAs in several cancers, including HNC, their exact role in HNC remains unclear, and there are currently no clinically established lncRNA biomarkers for diagnosis or therapy. Thus, this study aims to identify novel lncRNAs and investigate their significance in HNC. For this, we looked over recent research and discovered a new lncRNA called KLHDC7B-DT. The KLHDC7B-DT is significantly upregulated in HNC patient samples, and this result is corroborated by TCGA HNC data and cell lines. Further investigation suggests that KLHDC7B-DT may be linked with cancer stage, grade, age, and worse patient survival. We identified potential miRNAs that interact with KLHDC7B-DT and presented mRNA targets of these miRNAs, along with their possible functions. We propose that KLHDC7B-DT may regulate IL-6 expression in HNC, either through miRNAs or alternative mechanisms. Additionally, we found a significant positive correlation between the neighboring protein-coding gene KLHDC7B and the KLHDC7B-DT in HNC, both of which are significantly upregulated in HNC. In conclusion, KLHDC7B-DT is upregulated in HNC and holds promise as a prognostic and therapeutic biomarker. This study marks the first report highlighting the significance of KLHDC7B-DT in HNC.

cancer biology↗

bioRxiv2025

Biochemical characterization of ABHD14A, an outlying member of the metabolic serine hydrolase family

Certain uncharacterised members of the metabolic serine hydrolase enzyme family remain difficult to annotate due to poor tractability, context-dependent expression, and the absence of defined biochemical activities. Here, we provide the first functional characterization of the human enzyme ABHD14A. By engineering a soluble N-terminally truncated variant, we demonstrate by gel-based activity-based protein profiling and p-nitrophenyl-ester hydrolysis assays that ABHD14A is an active enzyme that preferentially turns over short-chain esters. Notably, ABHD14A exhibits a CoA-dependent enhancement of p-nitrophenyl-acetate hydrolysis, indicative of a ping-pong type acetyltransferase mechanism similar to that previously described for another homologous ABHD14-enzyme, ABHD14B. To investigate endogenous protein levels, we generated a high-affinity polyclonal antibody and found that ABHD14A is undetectable across a panel of immortalized mammalian cell lines and adult mouse tissues, challenging current transcriptomic database predictions. Upon heterologous expression in HEK293T cells, full length ABHD14A is catalytically active and localizes specifically to the Golgi apparatus, suggesting a specialized role in secretory pathway biology. Together, these findings establish the enzymatic activity, mechanistic features, and subcellular localization of ABHD14A, while providing essential biochemical and immunological tools that now enable the systematic discovery of its physiological substrates and regulatory contexts.

biochemistry↗

bioRxiv2025

Hippocampal CA3 Nex/Neurod6+ neuron-specific TNFR2 alleviates chronic neuropathic pain by sex-dependently engaging opioid and endocannabinoid pathways

Chronic neuropathic pain (CNP) develops as a result of persistent neuroinflammation and maladaptive synaptic plasticity in the central nervous system following nerve injury. While tumor necrosis factor receptor 2 (TNFR2) signaling has been extensively studied in pain resolution, the expression of this receptor on specific neuronal populations and molecular pathways involved in spontaneous pain recovery still remains poorly defined. In this study, we investigated the role of TNFR2 signaling within hippocampal Nex/Neurod6 pyramidal neurons in promoting recovery from chronic constriction injury (CCI), a well-established rodent model of neuropathic pain. To achieve neuron-specific deletion of TNFR2, we generated tamoxifen-inducible conditional knockout mice (NexCreERT2:TNFR2F/F). We demonstrate that knocking out TNFR2 from Nex neurons prevents spontaneous pain recovery in both males and females. Thus, establishing that a supraspinal TNFR2 neuroimmune axis is necessary for pain recovery. Exogenous administration of a TNFR2 agonist at 7, 10, and 13 dpi (i.p.) significantly improved mechanical withdrawal thresholds in both sexes of wild-type mice but did not alleviate pain in Nex-specific TNFR2 knockouts, indicating that neuronal TNFR2 expression is required for TNFR2-mediated analgesia. Bulk RNA sequencing of hippocampal tissue collected at six weeks after CCI revealed that TNFR2 activation upregulates genes such as Pomc, involved in the opioid pathway, and oleoyl-ACP-hydrolase (OLAH), involved in the endocannabinoid pathway. Consistent with these findings, immunostaining and Western blot analyses showed that TNFR2 agonism restored cornu ammonis (CA3) region POMC and {beta}-endorphin protein levels that were otherwise suppressed after CCI. Behavioral experiment demonstrated that systemic blockade of the {micro}-opioid receptor with naltrexone (administered daily from 7-21 dpi (s.c.)) completely prevented TNFR2-mediated pain recovery in males but only partially in females. In contrast, inhibition of cannabinoid 1 receptor (CB1R) signaling with AM251 (administered at 7, 14, and 21 dpi (i.p.)) abolished TNFR2-driven analgesia in both sexes. Together, these results reveal that hippocampal TNFR2 signaling in Nex/Neurod6 neurons is critical in recovery from chronic neuropathic pain. TNFR2 activation promotes analgesia by engaging endogenous {beta}-endorphin/{micro}-opioid and endocannabinoid pathways in a sex-dependent manner, establishing TNFR2 agonism as a promising non-addictive therapeutic approach for chronic pain resolution. SignificanceChronic neuropathic pain (CNP) results from persistent neuroimmune signaling and is driven by maladaptive circuit plasticity. Due to the complexity of factors contributing to CNP, it often leaves patients with few treatment options, which, unfortunately, are either temporary or might be addictive. We have characterized a novel supraspinal mechanism through which tumor necrosis factor receptor 2 (TNFR2) signaling, specifically in hippocampal Neurod6/Nex+ expressing pyramidal neurons, is necessary for pain recovery following nerve injury. Pharmacological activation of TNFR2 in these neurons alleviates pain by engaging both endogenous opioid and endocannabinoid signaling pathways. We specifically demonstrate that TNFR2 agonism upregulates proopiomelanocortin (POMC) expression and {beta}-endorphin levels in the hippocampus. We further identify that pharmacological inhibition of either the -opioid receptor or cannabinoid 1 (CB1) receptor is sufficient to impair the effectiveness of TNFR2 agonist mediated pain resolution. Our findings thus uncover a novel neuroimmune mechanism where the TNFR2 agonist, exogenously activating the pro-resolving TNFR2, mitigates CNP by releasing endogenous pain neuromodulators. Here, we highlight that TNFR2 agonism could serve as a non-addictive therapeutic strategy for the resolution of chronic neuropathic pain.

pharmacology and toxicology↗

bioRxiv2025

Master of none: GPRC6A gene loss is more widespread than previously known

GPRC6A encodes a class C GPCR that can be activated by multiple ligands and potentially acts as a central regulator of diverse metabolic processes by modulating endocrine pathways. Experimental studies have reported numerous distinct functions for GPRC6A, suggesting it may be a key drug target for several metabolic disorders. Yet, the actual function of GPRC6A has been the focus of considerable debate due to contradictory results and the prevalence of loss-of-function mutations in human populations, leading to the perception of GPRC6A as a "Master of none". Interestingly, a genome-wide screen for gene loss events in vertebrate species identified the disruption of the GPRC6A gene in toothed whales, in contrast to widespread conservation in the closely related Bovidae family. We employ a synteny-informed comparative genomic approach to demonstrate that the loss of the GPRC6A gene among mammalian species is more widespread than previously reported, encompassing the entire Bovidae group within Artiodactyla and other fully aquatic mammals, including those belonging to Sirenia. An in-depth search of the genomes and short and long-read sequencing datasets of monotremes, hystricomorphs, rhinolophoid bats, pika, koala, and two shrews (white-toothed pygmy shrew and Asian house shrew) reveals at least nine independent GPRC6A gene loss events in vertebrates, highlighting its lineage-specific dispensability and raising questions regarding its ubiquitous functionality. The evolutionary loss of GPRC6A likely represents a lineage-specific response to specialised diets and ecological niches, reshaping metabolic regulation and taste perception and illuminating how niche specialisation influences gene retention or loss within the GPCR landscape across species.

evolutionary biology↗

bioRxiv2025

Spatial transcriptomic profiling identifies lacrimal gland epithelial cell-driven mechanisms underlying autoimmunity in Sjogrens disease.

Sjogrens disease (SjD) is a second most prevalent rheumatic disease involving autoimmune pathology of tear-producing lacrimal glands that leads to a common clinical manifestation of chronic ocular surface disease. Despite advances in understanding of SjD, lacrimal gland pathology remains incompletely understood limiting diagnosis and treatment. Here we analyze spatial transcriptomic profile of lacrimal glands from wild-type (C57Bl/6) mice and Thrombospondin (TSP)-1-/- mice, a spontaneous mouse model of SjD. We uncover molecular mechanisms underlying functional loss of major epithelial cell subtypes - acinar, duct and myoepithelial cells. We identify potential early mechanisms and markers of glandular damage. By integrating spatial and cellular profiles, we uncover the presence of antigen presenting cells in the proximity of duct epithelial cells that were not described previously in lacrimal glands. We further identify role of epithelial cells as active participants in promoting or sustaining inflammation. Our findings help reveal potential molecular and cellular cues that drive periductal infiltrates containing B cells and Tfh cells that form germinal centers to facilitate local autoantibody production. Overall, our study can provide a framework for therapeutic targeting of epithelial cell types and multicellular interactions underlying autoimmune pathology. Significance StatementTears produced by lacrimal glands are critical for protecting the eye surface by preventing tissue dryness and maintaining normal vision. Disruption of this function due to autoimmune inflammation in Sjogrens Disease compromises the protection of the eye surface causing dryness, a condition with a potential for sight-threatening complications like infections and ulcers. Understanding cellular and molecular interactions that lead to functional loss and autoimmune inflammation of the lacrimal gland is critical for developing effective therapies. We have analyzed transcriptional profile of glandular cells in a tissue section where their morphology and interactions with surrounding cells is preserved. By comparing glands from normal and mice with Sjogrens disease we identify molecular mechanisms that can form the basis for new therapies.

immunology↗

bioRxiv2025

Spatial Transcriptomics Identify T Cell-Driven Mechanisms of Kidney Damage in Immune Checkpoint Inhibitor-Associated Acute Interstitial Nephritis

IntroductionImmune checkpoint inhibitor-associated acute interstitial nephritis (ICI-AIN) is the most common finding on histopathology among patients with ICI-associated acute kidney injury (ICI-AKI). Patients with ICI-AIN often have T cell-dominant infiltration of the kidney and high tissue levels of CXCR3 ligands like CXCL9, 10, and 11; however, the mechanisms of inflammation in ICI-AIN are not well-understood. MethodsWe applied a sub-cellular spatial transcriptomics platform (Xenium Prime 5K) to compare the cellular composition of kidney biopsy tissue from patients with ICI-AIN with ICI-treated patients with acute tubular necrosis (ICI-ATN). ResultsAcross 8 kidney biopsy specimens (4 with ICI-AIN, 4 with ICI-ATN), we analyzed 332,000 cells, comprising kidney parenchymal cells and infiltrating immune cells. Using a spatially-aware cellular neighborhood-based classification, we identified cellular niches corresponding to each part of the nephron, in addition to unique fibrotic and inflammatory niches. Gene pathway analysis identified interferon-gamma (IFN-{gamma})/STAT1 signaling as strongly increased in ICI-AIN compared to ICI-ATN. While all inflammatory niches were overrepresented in ICI-AIN, CD8+ T cell infiltration and proinflammatory myeloid cells were the dominant immune niches. Spatial niche crosstalk analysis revealed that CD8+ T cell-derived IFN-{gamma} likely induced a proinflammatory program in myeloid cells, with increased production of CXCL9, 10, and 11. Furthermore, IFN-{gamma} signaling in ICI-AIN was associated with reduced oxidative phosphorylation in kidney tubular niches. ConclusionsSpatial transcriptomics reveal novel insights into key differences in the pathophysiology of ICI-AIN versus ICI-ATN. IFN-{gamma}-producing CD8+ T cells are likely key drivers of ICI-AIN and should be investigated as future therapeutic targets. Translational StatementSpatial transcriptomics may provide insight into differences in the cellular and spatial composition of immune checkpoint inhibitor-associated acute interstitial nephritis (ICI-AIN) and ICI-associated acute tubular necrosis (ICI-ATN). Using the novel Xenium 5K platform, we demonstrate that IFN-{gamma}-producing CD8+ T cells are central to the pathogenesis of ICI-AIN, and that CD8+ T cell-derived IFN-{gamma} likely induces a proinflammatory state in myeloid cells, with increased tissue production of CXCL9, 10, and 11. We then show that a reduction in oxidative phosphorylation and an IFN-{gamma}-driven influx of proinflammatory myeloid cells may further drive kidney damage in ICI-AIN. Many of these pathways represent potential druggable targets, and our findings may therefore inform future therapeutic approaches for ICI-AIN.

immunology↗