Biology subjects

Gupta, S.

Publications and source records attributed to Gupta, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

In Vitro and Computational Evaluation of Thrombolytic Activity of Kinemakinase of Kinema, an Indigenous Fermented Food of Eastern Nepal

Kinema is a traditional fermented soybean food indigenous to the eastern Himalayan regions of Nepal and India. The fermentation process is primarily mediated by the bacterium Bacillus subtilis, which produces several bioactive compounds and enzymes with potential therapeutic applications. Considering the growing burden of cardiovascular diseases and the need for effective fibrinolytic agents for thrombolytic therapy, this study aimed to extract, partially purify, and evaluate the thrombolytic potential of kinemakinase derived from kinema prepared from white soybeans. Partial purification of the enzyme was achieved using ammonium sulfate precipitation. Thrombolytic activity was assessed in vitro using human blood clots, where three enzyme dilutions demonstrated clot lysis ranging from 66% to 68%, indicating considerable fibrinolytic potential. In silico analyses were also performed to investigate the structural and functional characteristics of the enzyme. The tertiary structure obtained from UniProt was modeled using the Robetta server and refined with GalaxyRefine. Docking with fibrin using ClusPro 2.0 and molecular dynamics simulations using iMODS confirmed favorable interaction and structural stability, while disulfide engineering enhanced protein stability. The findings suggest that kinema-derived kinemakinase may serve as a promising alternative thrombolytic agent, warranting further biochemical characterization and dosage optimization. Keywords: Kinema, Kinemakinase, Thrombolytic activity, Bacillus subtilis, Fermentation

microbiology↗

bioRxiv2026

SWI/SNF Alterations Define a Chromatin-Dependent Subtype of Urothelial Carcinoma

PurposeSWI/SNF (BAF) chromatin remodeling complex alterations are common in urothelial carcinoma, yet no biomarker-directed therapeutic strategies have been established for this population. We investigated whether BAF alterations delineate a biologically distinct, therapeutically actionable urothelial carcinoma subtype. Experimental DesignWe performed integrative genomic and transcriptomic analyses of 792 urothelial carcinoma tumors from the Oncology Research Information Exchange Network (ORIEN) and validated findings in the TCGA-BLCA cohort. Mechanistic studies incorporated RNA sequencing and ATAC-seq following histone deacetylase (HDAC) inhibition. Functional dependencies were assessed using patient-derived xenograft organoids and cell line models. Clinical relevance was explored in a biomarker-enriched investigator-initiated trial. ResultsApproximately half of urothelial carcinoma tumors exhibited BAF alterations, defining a previously unrecognized chromatin-altered molecular subtype characterized by activation of proliferative programs, loss of lineage identity, and altered metabolic signaling. This subtype was enriched for transcriptomic programs associated with HDAC inhibitor sensitivity and depleted of HDAC inhibitor resistance signatures. Mechanistically, HDAC inhibition induced widespread chromatin remodeling with reduced accessibility at AP-1 and TEAD-associated regions, and downregulation of E2F- and MYC-driven transcriptional networks. Functional studies confirmed enhanced HDAC inhibition sensitivity in ARID1A-mutated cell lines and a patient-derived organoid model. Early clinical observations demonstrated a durable responder treated with HDAC inhibitors and immunotherapy. ConclusionsBAF alterations define a chromatin-dependent tumor state in urothelial carcinoma that is selectively vulnerable to HDAC inhibition. Integrating genomic, epigenomic, functional, and early clinical evidence, these findings provide a rationale for biomarker-enriched clinical trials and HDAC inhibitor-based combination strategies in urothelial carcinoma.

cancer biology↗

bioRxiv2026

SketchDNA: A GUI-Enabled Toolkit for Multiscale Modeling of Topological DNA Structures

Computational modeling tools have enabled detailed exploration of the structural dynamics of nucleic acids at the nanoscale. Despite these developments, a unified platform for creating multiscale models of topological DNA structures, which are of fundamental importance across rapidly converging biological disciplines, is lacking. Here we present a GUI-enabled topological DNA design tool, SketchDNA (SDNA), that facilitates a user-friendly interface to create all-atom (AA) and coarse-grained (CG) models of DNA structures with tunable topological parameters. Building on the modular and open-source framework of SDNA, the software can be readily integrated with both emerging and existing DNA design platforms, such as oxDNA, MrDNA, etc. We demonstrate the utility of SDNA computational framework by simulating the conformational dynamics of three representative topological DNA structures, minicircles, catenanes, and Borromean rings. Analyzing AA and CG molecular dynamics (MD) simulations of these topological DNA systems using the AMBER and Martini DNA force fields, respectively, we characterize their equilibrium structural dynamics, fluctuations, and topological properties. While the AA simulations allow us to characterize the topology-dependent structural rearrangements at the nucleotide level, long-timescale CG simulations reveal supercoiling-induced conformational transitions. The multiscale SDNA toolkit broadens the applications of MD simulations by enabling in-situ characterization of the biophysical properties of topological DNA nanostructures. The GUI version of SDNA is available at https://sdna.biotech.iith.ac.in without registration, while the source code is accessible at GitHub.

biophysics↗

bioRxiv2026

STRIDE: Signal Transfer and Aligned Ion-Peak Discrimination for Peptidoform Evidence for Consistent Quantification of Site-Localized Post-Translational Modifications in Large-Scale DIA-MS

Data-independent acquisition (DIA) mass spectrometry has emerged as a powerful method for comprehensive post-translational modification profiling in proteomics, but faces challenges in consistently assigning peptidoforms to the same chromatographic peaks across multiple samples. This study introduces STRIDE to address this challenge by implementing dynamic programming alignment and propagating unique ion signature signals across runs. We benchmarked STRIDE using a synthetic phosphopeptide dilution series dataset, demonstrating an 11.3-fold reduction in false-negative rate compared to DIA-NN, from 68% with DIA-NN to 6% with STRIDE at 5% false-positive rate when requiring both correct site-localization and correct peak assignment. STRIDE also improved over the base OpenSWATH (IPF), reducing the false-negative rate from 14% to 6% for correct peak assignment. When applied to a phospho-enriched U2OS nocodazole dataset, our approach reduced missing values from 38.6% to 15.2% in a representative quantification matrix, increased highly reproducible phosphopeptide detections from 7,698 to 12,510 in nocodazole-treated samples, and identified 5,431 differentially abundant phosphopeptides compared to 2,588 with DIA-NN. This increased the number of enriched pathways recovered from differential phosphopeptide analysis, including pathways related to mitotic arrest, microtubule organization, chromatin regulation, RNA processing and kinase signalling. Lastly, we analyzed acetyllysine and glycopeptide-enriched DIA datasets, where STRIDE improved complete replicate detection, reduced missing quantification values, and increased site-specific glycan reproducibility. Together, these results demonstrate that STRIDE improves peptidoform proteomics through reduced false-negative rates, consistent peak assignment, and more complete quantification across runs.

bioinformatics↗

bioRxiv2026

Arsenophonus ghosts in bed bug genomes: multiple origins and 50 million years of persistence

Although numerous studies have documented horizontal gene transfer (HGT) from bacteria into insect genomes, they have been heavily biased toward the endosymbiont Wolbachia. In contrast, comparatively few studies have examined HGT from other symbionts and insect-associated bacteria. Moreover, research on these systems has primarily focused on transferred genes that retain function in the insect host, contributing to metabolism, symbiosis, detoxification, or host adaptation. Consequently, relatively few empirical systems have been available to investigate the persistence, degradation, diversification, and vertical inheritance of apparently non-functional HGT-derived sequences over macroevolutionary timescales. The widespread occurrence of putatively Arsenophonus-derived sequences across Cimicidae therefore provides a rare opportunity to extend these investigations beyond the Wolbachia model and to examine the long-term evolutionary fate of non-functional HGT-derived DNA across tens of millions of years of host diversification. We show that Arsenophonus-derived HGTs have multiple independent origins across Cimicidae but that one major HGT lineage has persisted through diversification of the subfamily Cimicinae for at least 50 million years. Phylogenetic and compositional analyses indicate that an ancestral Arsenophonus-derived genomic region has undergone progressive fragmentation, leaving numerous dispersed, apparently non-functional remnants while preserving a clear evolutionary signature. These results extend the study of bacterial HGT beyond the Wolbachia model and demonstrate that non-functional symbiont-derived DNA can persist over macroevolutionary timescales.

evolutionary biology↗

bioRxiv2026

A Synthetic Microbial Therapy Rewires Antitumor Immunity Across Multiple Cancer Types

Microbial immunotherapies show promise against cancer, yet broad efficacy and mechanistic insight remain elusive. Here, we introduce SPIKE 1.0 (S1.0), a metabolically engineered bacterium that converts tryptophan into immunomodulatory hydroxyindoles via tryptophan monooxygenase to remodel the tumor microenvironment (TME). A single systemic dose of S1.0 elicited potent, durable antitumor responses across multiple murine models, including humanized mice, with minimal toxicity. S1.0 enhanced inflammatory signaling, activated innate and adaptive immunity, and promoted T cell persistence, memory, and resistance to exhaustion. It outperformed checkpoint inhibitors and synergized with chemotherapy. Multi-omics profiling revealed that S1.0 rewired amino acid metabolism in tumor-infiltrating immune cells and disrupted immunosuppressive networks. These results establish S1.0 as a scalable, cost-effective microbial immunotherapy with broad translational potential for solid tumors.

immunology↗

bioRxiv2026

Biological recognition of mirror-image glycans

Recent synthesis of essential enzymes, such as DNA and RNA polymerases with opposite chirality, has boosted the feasibility of creating mirror-image life. Such life, if ever produced, will undoubtedly be coated by a dense display of glycans (glycoproteins, glycolipids, polysaccharides) built from enantiomers of common monosaccharides. Recognition of mirror-image glycans by extant glycan-binding proteins (GBPs) may be critical for colonization by or immune response to mirror life organisms. We evaluated recognition of enantiomers of common glycans by a diverse set of purified GBPs (plant and human derived), antibodies (including IgM from human plasma), mammalian cells (including immune cells), and organs in live animals. We found that GBP binding to enantiomers of naturally prevalent glycans is widespread. Notably, L-glucose and L-galactose interact with fucose-binding lectins, including DC-SIGN, a C-type lectin expressed on immune cells. These interactions can be inhibited by soluble "natural" glycan ligands and enantiomeric ones confirming specificity. Binding of L-glycans to diverse immune cell repertoires revealed preferences for specific glycan enantiomers. IgM antibodies from human serum showed donor-specific recognition of L-glycans. We propose that the recognition of L-glycans by extant GBPs arises from their co-evolution over millennia with the L-glycans that are present in the glycocalyx of many microorganisms.

biochemistry↗

bioRxiv2026

Identification of a new mycobacterial peptide that controls the activity of the iron-dependent regulator, IdeR.

Mycobacterium tuberculosis (Mtb) must regulate intracellular iron to survive in the host and cause disease. IdeR (Iron-dependent Regulator) is an essential Mtb protein that governs intracellular iron levels by regulating the expression of genes involved in iron metabolism. IdeR expression, however, is iron independent, and the mechanisms that regulate IdeRs function are not fully understood. Here, we report the discovery of a previously unrecognized Mtb peptide (PRI) that regulates IdeR activity. PRI is induced under iron limitation; it binds to IdeR and restricts its activity. In addition, we demonstrate that altering the balance between PRI and IdeR impairs iron homeostasis and intracellular replication of Mtb in macrophages. The findings reveal a new paradigm in mycobacterial iron regulation and open new avenues for targeting iron homeostatic mechanisms in Mtb, which are crucial for virulence and antibiotic resistance.

microbiology↗

bioRxiv2026

Biomolecular Condensation and L-Cysteine Signaling Activates Dormant Protease Activity of Papain Droplets: Implication toward Meat Tenderization

Proteases are an important class of proteolytic enzymes having great importance in both basic science and industrial applications. While cells tightly regulate the spatio-temporal activity of different proteases for cellular homeostasis, mis-regulation often leads to adverse effects. In this context, the protease activity of papain and its activation by L-cysteine is poorly understood in the literature. Herein, we discover that the protease activity of papain can be effectively regulated via a spontaneous liquid-liquid phase separation (LLPS) pathway. We show that papain undergoes biomolecular condensation via spontaneous LLPS under macromolecular crowding through the involvement of intermolecular hydrophobic interactions. Secondary structure analyses revealed a compact conformation of phase-separated papain with increased -helix content. Although native free papain is found to be active towards synthetic and protein substrates, the proteolytic digestion produces heterogeneous peptide aggregates. In contrast, we found that papain droplets remain dormant toward protein digestion due to the disulfide linkage of the active cysteine residue (Cys-25) in its compact conformational state. More importantly, we show that the protease activity of phase-separated papain can be reactivated in the presence of L-cysteine to produce uniform soluble peptide fragments. Our findings indicate that although disulfide linkages are not necessary for the phase separation of papain, upon phase separation, intermolecular interactions between phase-separated papain result in the formation of disulfide linkages involving active Cys-25 residues. The present discovery has tremendous technological importance to boost the efficacy of meat tenderization in the food industry.

biophysics↗

bioRxiv2026

Smad1 and Smad5 differentially transduce BMP signaling during in vitro differentiation of mouse embryonic stem cells into dorsal interneurons

A central unresolved question in development biology is how systems of overwhelming complexity arise from relatively few families of growth factors. Compounding this issue, signaling pathways often show signal convergence, where many ligands interact with fewer receptors, which then signal through a single second messenger complex. Here we investigate this question in the context of bone morphogenetic protein (BMP) signaling and its role directing dorsal spinal cord development, focusing on two receptor-regulated (R) Smads, Smad1 and Smad5. Multiple models have been proposed for their mode of action from acting redundantly through combined signal strength, to having distinct activities that drive different fate outcomes. We sought to distinguish between these models by generating CRISPR-edited Smad1 and Smad5 null mouse embryonic stem cell (ESC) lines to dissect the cell fate of activities of individual R-Smads, with a resolution not possible in vivo. Using a directed differentiation protocol for dorsal interneurons (dI), together with bioinformatic analyses, we have defined the roles of the R-Smads at key decision points along the dI specification timeline. Together, these findings support a model in which Smad1 and Smad5 play largely distinct roles in dorsal spinal cord development. While both R-Smads can activate canonical BMP-responsive transcriptional targets, they asymmetrically contribute to cell fate specification. Smad1 plays a restricted role, while Smad5 has a dominant role, regulating dorsal progenitor transcriptional dynamics and reiteratively directing the dorsal-most dI fates.

developmental biology↗

bioRxiv2026

Defining biologically directed therapy target signatures in urothelial carcinoma: A transcriptomic framework for precision therapy

IntroductionNectin-4 targeting antibody-drug conjugate (ADC) enfortumab vedotin (EV), in combination with pembrolizumab, is the first-line treatment for patients with locally advanced or metastatic urothelial carcinoma (UC). Optimal treatment strategies for patients who are non-responders or progress on EV with pembrolizumab remain an unmet clinical need. We sought to characterize ADC and immunotherapy (IO)-associated target expression profiles to identify candidate therapeutic vulnerabilities beyond EV. MethodsWe conducted a literature review to identify ADC and IO targets with approved or investigational relevance in UC. Unsupervised hierarchical clustering was used to identify clusters of target gene expression in RNA-seq data. Transcriptomic clustering analyses were performed in 434 patients from The Cancer Genome Atlas Bladder Urothelial Carcinoma cohort (TCGA-BLCA) and validated in an independent cohort of 478 patients from the Oncology Research Information Exchange Network (ORIEN) consortium. Proteomic interrogation of these targets was performed using mass spectrometry data from additional cohort of 116 patients. Differential gene expression analyses evaluated associations between target expression patterns, histologic variants, and consensus molecular subtypes of muscle-invasive bladder cancer (CMIBC). ResultsWe identified 13 ADC and 10 IO-associated targets with translational relevance in UC. Transcriptomic analyses revealed three reproducible clusters of overexpressed target genes across independent cohorts: 1) a luminal/epithelial-associated cluster enriched for VTCN1, SLITRK6, FGFR3, NECTIN4, TACSTD2, ERBB2, and ERBB3; 2) an immune target predominant cluster enriched for BTLA, LAG3, PDCD1, TIGIT, CTLA4, TNFRSF9, TNFRSF18, TNFRSF4; and 3) a basal/neuroendocrine-associated cluster characterized by CD274, F3, NT5E, EGFR, MET and DLL3. Similar clusters were largely conserved at the proteomic level. Adenocarcinomas overexpressed ERBB3 compared to neuroendocrine and squamous cell carcinomas. Pure squamous cell carcinomas overexpressed TACSTD2 compared to adenocarcinomas. In CMIBC subtypes, basal/squamous tumors expressed higher levels of CD274, EGFR, F3, LAG3, NT5E, and TNFRSF18, whereas luminal tumors demonstrated higher ERBB2 and ERBB3 expression. Neuroendocrine-like tumors showed higher DLL3 expression compared to all other subtypes. Tumors with low expression of NECTIN4, TACSTD2, and FGFR3 were enriched for alternative targets including DLL3, CD274, and CD276. Our findings provide a framework for hypothesis-driven therapeutic prioritization in advanced UC. Conclusions: UC is characterized by reproducible, biologically distinct patterns of ADC and IO target expressions. The degree of expression of NECTIN4 was positively associated with TACSTD2, FGFR3 and inversely associated with DLL3, CD276, and CD274, supporting alternative biologically informed treatment strategies besides EV . Histologic variants and molecular subtypes of UC also display distinct patterns of target expression. This study provides the first integrated transcriptomic framework linking ADC and IO target co-expression patterns for hypothesis-driven therapeutic prioritization. These findings provide a basis for rational ADC and immunotherapy development in advanced UC and support prospective proteomic validation in treatment stratified cohorts. Statement of Translational RelevanceEnfortumab vedotin plus pembrolizumab has redefined first-line therapy for advanced urothelial carcinoma, yet treatment selection following resistance or progression remains undefined. In this study, we integrate transcriptomic and proteomic analyses across independent cohorts to define reproducible patterns of antibody-drug conjugate (ADC) and immunotherapy target co-expression in urothelial carcinoma. We identify biologically distinct target-expression patterns that are associated with histologic and molecular subtypes and demonstrate coordinated and, in some cases, mutually exclusive relationships among therapeutically actionable targets. These findings have direct translational implications. First, they provide biologic rationale for rational sequencing and combination strategies based on co-expressed targets in NECTIN4-enriched tumors. Second, they identify alternative therapeutic vulnerabilities, including DLL3- and CD274-associated pathways, in tumors with low NECTIN4 expression, a population potentially enriched for resistance to EV-based therapy. Finally, this framework establishes a foundation for biomarker-driven clinical trials in urothelial carcinoma and supports the development of precision therapeutic approaches beyond current standards.

cancer biology↗

bioRxiv2026

Evidence of Filopodial translocation of Blastema associated microRNA rich Exosome like Extracellular Vesicles

Zebrafish (Danio rerio) possess remarkable regenerative capacity, making them an ideal model for studying the molecular mechanisms underlying tissue regeneration. In this article we report the identification of blastema linked exosome like extracellular vesicles (EVs) in zebrafish, that to the vesicles were plausibly being translocated in the proximo-distal axis through filipodia. We further thoroughly examined the exosome like EVs isolated from regenerating tissues of zebrafish caudal fins to characterize their nucleic acid cargo and evaluate their potential regulatory functions in regeneration. Caudal fins were amputated and allowed to regenerate and exosome like EVs isolated from blastema tissues displayed increased abundance compared to non-amputated controls. RNA sequencing identified a dynamic cluster of EV linked microRNAs (miRs). These differentially expressed miRs, including dre-miR-21, dre-miR-200b, dre-miR-218a and dre-let-7e were upregulated and associated with promoting proliferation, migration, differentiation, and tumour suppression pathways. Moreover, dre-miR-100, dre-miR-146a and dre-miR-200c regulated osteogenic differentiation, inflammatory signalling, epithelial-mesenchymal transition, and cell adhesion. Regeneration is generally believed to be coordinated only by local morphogen diffusion. Through this study it is indicative that filipodia bound EVs might have a pivotal role in long-range communication between blastema and the proximal tissues during the regeneration process. A detailed analyses of the miR targets and their validation could potentially lead to novel advancement and solutions in the field of regeneration and regenerative medicine in the near future.

Developmental Biology↗

bioRxiv2026

PAD4-mediated histone citrullination contributes to enhanced NETosis in type 2 diabetes but not in type 1 diabetes

Neutrophil extracellular traps (NETs) are web-like structures released by activated neutrophils through a process known as NETosis, which help immobilize, trap, and eliminate invading pathogens. While NETs play a critical role in host defense, excessive or dysregulated NET formation can contribute to chronic inflammatory diseases such as diabetes and its complications. Elevated levels of NETs and increased expression of its mediator, protein-arginine deiminase type 4 (PAD4), have been reported in diabetes. However, the underlying molecular mechanisms governing the enhanced NETosis in type 1 (T1D) and type 2 diabetes (T2D) remain unclear. Using mouse models and human patient samples, we show that neutrophils undergo enhanced NETosis in T1D and T2D through distinct pathways. We found enhanced NETosis in T1D occurs in a PAD4-independent manner, driven by robust cytosolic ROS production by NADPH oxidase (NOX). This leads to myeloperoxidase activation and chromatin decondensation. We further confirm the PAD4 independent mechanism in neutrophils from STZ-induced PAD4-/- mice. In contrast, in T2D, neutrophils undergo NOX-independent NETosis, which relies on calcium-mediated mitochondrial ROS production, PAD4 activation, and hyper-histone citrullination. Taken together, our findings reveal previously uncharacterized distinct mechanisms of enhanced NETosis in T1D and T2D.

Molecular Biology↗

bioRxiv2026

Improving splice site usage prediction with SPLAIRE

BackgroundAlternative splicing, the mechanism by which intronic sequences are excised from pre-mRNAs to produce mature mRNA, affects >95% of human protein-coding genes and is a major driver of human disease states. The spliceosome, a protein-RNA complex responsible for splicing pre-mRNA, identifies candidate splice sites partly through the recognition of characteristic sequence motifs at exon-intron junctions. Deep learning models that predict the presence of splice sites from pre-mRNA sequence have achieved breakthrough performance relative to previous machine-learning techniques, and these models have improved our ability to identify pathogenic genetic variants that alter splicing. ResultsWe show that, while overall performance measures from these models suggest near-perfect performance, substantial gaps in prediction remain, including the identification of splice sites with low usage rates and tissue-specific splice sites. We leverage one of the largest paired RNA and genotyping datasets used to date to train a novel splicing model optimized for a specific cell type, human airway epithelial cells. We trained a dilated convolutional neural network on data from cultured airway epithelial cells from 100 donors, and showed that this model outperforms current state-of-the-art models on splice site identification and splice site usage quantification, including on multiple tissues not included in the model training data. ConclusionsWe present the most comprehensive evaluation of state-of-the-art splicing models published to date, revealing reasonable performance across models for genetic variant effect prediction along with important performance gaps and insights into directions for future model development.

genomics↗

bioRxiv2026

HNSW-MS: Hierarchical Graph Indexing Enables Accurate Real-Time Mass Spectral Similarity Search at Repository Scale

Spectral similarity comparison is the basis of mass spectrometry-based metabolomics, underpinning library matching, molecular network construction, and repository searches such as MASST. Public repositories now exceed billion spectra, and exhaustive pairwise comparison is increasingly limiting. We introduce HNSW-MS, which implements Hierarchical Navigable Small World graph indexing natively for mass spectra. It operates directly on GC-MS and LC-MS/MS spectra without embedding, so scores are identical to those of exhaustive comparison and results are fully reproducible. On the benchmark dataset of 8.4 million LC-MS/MS spectra, HNSW-MS is up to 900-fold faster than linear search, recall@1 and recall@10 achieve over 90%, with the recall-speedup tradeoff tunable at query. Such rapid retrieval allows for any query spectrum to be near-instantaneously integrated into a global molecular network. We demonstrate specific examples of use of global networking for molecular discovery, including uncovering a new pathway of stenothricin biosynthesis and a new dipeptide conjugate bile acid.

bioinformatics↗

bioRxiv2026

A DNA-binding protein senses DNA superhelicity to switch between bridging and nucleoprotein filament formation

The torsional state of DNA encodes regulatory information beyond sequence, yet how chromosome-associated proteins read this mechanical signal to switch between distinct functional outputs remains poorly understood. Using real-time single-molecule fluorescence imaging on topologically constrained DNA, we demonstrate that DNA superhelical polarity is the primary determinant of H-NS binding mode with direct consequences for chromosomal domain organization and gene silencing. On positively supercoiled DNA, H-NS assembles into nucleoprotein filaments at AT-rich loci, acting as a topological barrier that confines plectoneme diffusion. On negatively supercoiled DNA, H-NS switches to bridging mode, immobilizing plectonemes at AT-rich sites. Upon changes in DNA topology, H-NS dynamically switches between binding modes within seconds. An oligomerization-deficient mutant retains bridging but cannot form filaments, confirming the mechanistic distinction. On multi-AT-locus DNA, helicity-driven mode selection produces self-organized "chromosomal" architecture: some sites stochastically capture the plectoneme and bridge, while remaining sites assemble insulating filaments. These findings establish DNA superhelical polarity as the master switch governing H-NS binding mode and "chromosomal" domain organization, with broad implications for transcriptional regulation in bacteria.

biophysics↗

bioRxiv2026

A Novel MicroRNA-Odorant Receptor Axis Governs Neural Progenitor Cell Proliferation in Zebrafish CNS

Spinal cord injury causes irreversible neurological deficits in mammals; yet zebrafish achieve complete functional recovery through molecular mechanisms that remain poorly defined. In this study we emphasized on a critical miRNA-mediated regulation of Ependymo-radial glial (ERG) cell proliferation in zebrafish spinal cord. Using next-generation sequencing we constructed a spatiotemporal miRNA profile across multiple post-injury time points and identified dre-miR-N1 as a novel injury-responsive miRNA involved in ERG proliferation among several differentially expressed novel miRNAs. Fluorescent in situ hybridization confirmed its robust lesion-site expression and gain-of-function analysis demonstrated that dre-miR-N1 significantly impaired functional recovery. Target prediction and validation unexpectedly identified the odorant receptor gene or42a1 as a high-confidence target and a combinatorial approach of miRNA gain-of-function and or42a1 loss-of-function showed that dre-miR-N1 modulates the proliferative behaviour of or42a1-expressing ERG cells under both homeostatic and injury conditions. These findings uncover a previously unrecognized miRNA-odorant receptor axis governing injury-induced ERG cell expansion establishing a novel molecular framework for endogenous neural regeneration in zebrafish.

neuroscience↗

bioRxiv2026

From APOE Genetics to AI-Designed Drug Candidates: An Integrated Pipeline for Oral, Brain-Penetrant ACAT1 Inhibitors in Alzheimer's Disease

For more than 55 million people living with dementia worldwide, no oral disease-modifying treatment is currently available. Alzheimers disease (AD) remains one of the most urgent unmet needs in neurology, with recent genetic evidence estimating that 72-93% of AD burden is attributable to common APOE allelic variation. Mechanistically, the APOE {varepsilon}4 isoform impairs cholesterol transport in the brain, promoting cholesteryl ester accumulation in microglia; these lipid-laden cells lose phagocytic capacity for amyloid-{beta} clearance and autophagy-mediated degradation of phosphorylated tau, linking a single upstream metabolic disruption to both hallmark pathologies of AD. ACAT1/SOAT1, the brains predominant cholesterol-esterifying enzyme, is an attractive therapeutic target: its inhibition reduces cholesteryl ester formation, restoring microglial A{beta} clearance, reducing A{beta} production, and promoting tau degradation via autophagy, supporting a multimodal mechanism not addressed by currently approved therapies. However, prior ACAT inhibitor programs were limited by isoform non-selectivity, excessive lipophilicity, and lack of CNS optimization. Addressing these constraints, we present the CuraGenAI Drug Discovery Platform for the design of oral, brain-penetrant ACAT1-targeted small-molecule candidates. The pipeline integrates scaffold-constrained generative chemistry with multi-objective ADMET optimization across more than 30 criteria. From approximately 6 million generated molecules, multi-stage filtering yielded approximately 7,300 CNS-optimized candidates with predicted oral brain-penetrant profiles distinct from prior ACAT clinical compounds. Three nominated lead candidates showed predicted BBB probabilities >0.93, favorable CNS drug-like physicochemical profiles, and stable ACAT1 binding poses, and are prioritized for synthesis and in vitro validation.

neuroscience↗