Biology subjects

Kumar, M.

Publications and source records attributed to Kumar, M..

At least 19 recordsLinked to original sources

bioRxiv2026

A cellulose synthase interactome uncovers BAG proteins as regulators of cellulose synthase homeostasis

Cellulose synthase complexes build the load-bearing cellulose microfibrils of plant cell walls, yet how the abundance of their catalytic CELLULOSE SYNTHASE A (CESA) subunits is maintained remains unclear. Here, we used multi-bait TurboID proximity labelling with ten cellulose-synthesis-associated baits and six subcellular controls to define a high-confidence cellulose synthase neighbourhood. Stringent spatial and recurrence-based filtering yielded a core network of 119 interactions among 44 proteins and identified three members of the conserved Bcl-2-associated athanogene (BAG) family as previously unrecognised regulators of cellulose synthase homeostasis. BAG1-3 associated with primary-wall CESAs in reciprocal proximity-labelling experiments. Arabidopsis bag mutants showed reduced cellulose accumulation, hypersensitivity to cellulose-synthesis inhibitors, and markedly decreased CESA protein abundance without corresponding changes in CESA transcript levels. Loss of BAG function also increased the accumulation of CESA6 in vacuolar compartments. These findings identify BAG proteins as previously unrecognised regulators of cellulose synthase homeostasis and link a conserved proteostasis-associated protein family to plant cell wall biosynthesis. More broadly, the study establishes multi-bait proximity labelling, combined with cell location-specific controls, as a strategy for resolving dynamic protein networks whose components traffic through multiple subcellular compartments.

plant biology↗

bioRxiv2026

A New aspect of the pathology of brain tuberculosis: Mycobacterium tuberculosis infects and alters human neural progenitor cells

Brain tuberculosis remains associated with high mortality, and many survivors exhibit cognitive impairments. Progress in understanding the disease is hindered by the lack of human models. In this study, human neural organoids were infected, revealing that a subpopulation of neural progenitor cells (NPCs) is directly infected by apoptotic cell receptors expressed by NPCs, mediating bacterial uptake. Phagocytosed bacteria were localized in late endosomes, lysosomes, and the cytoplasm. Cytoplasmic bacteria frequently formed cords, indicating limited control of bacterial expansion. Immunostaining demonstrated that infected NPCs produce a type I interferon (IFN) response, corroborated by increased expression of type I IFN and IFN-regulated genes detected by RNA sequencing. Pathways related to innate immune response, cell death, and proliferation were also activated following Mycobacterium tuberculosis (Mtb) uptake by NPCs. The addition of color-coded microglia and monocytes to 3D neural organoids and NPCs revealed cross-infection of NPCs and other phagocytes by Mtb, suggesting a mechanism by which NPCs may access the bacteria. Infection of NPCs resulted in increased cell death, inhibition of neural differentiation, and reduced proliferation, effects that were partially mitigated by anti-IFN treatment. Differentiated neurons were not infected. These findings indicate that brain organoids and NPC-based in vitro platforms provide a novel approach for studying brain tuberculosis. Decreased NPC function may contribute to brain tuberculosis-induced cognitive disease.

immunology↗

bioRxiv2026

Malnourishment and expanded microbiome maintain superior efficacy of the recombinant tuberculosis vaccine BCG::ESAT-6-PE25SS

Tuberculosis (TB) remains the leading cause of infectious mortality. The limited efficacy of the only TB vaccine, Bacille Calmette-Guerin (BCG) against pulmonary disease necessitates improved vaccines. Host factors such as malnutrition and microbiome composition shape immune responses in humans, though these factors are overlooked in preclinical vaccine evaluation. Here we show that a recombinant BCG strain, BCG::ESAT-6-PE25SS, confers superior protection compared to BCG across murine models of malnutrition, antibiotic-induced dysbiosis and environmentally enriched microbiota. Unexpectedly, malnourished mice displayed reduced Mycobacterium tuberculosis (Mtb) burden, associated with altered host metabolism and immune composition. Microbiome disruption increased TB susceptibility, whereas diversification of microbiota enhanced resistance and immune heterogeneity. Vaccine efficacy correlated with enrichment of known immunomodulatory microbial taxa. These findings suggest diet-microbiome-immunity interactions as potential key determinants of TB pathogenesis and provide evidence for the importance of vaccine candidate evaluation under physiologically relevant co-morbid conditions

microbiology↗

bioRxiv2026

Chemical Rescue Serves as a Predictive Proxy for Glycosynthase Activity on Glycosidic Bonds via a Shared Glycosyl Oxocarbenium Transition State

Engineered glycosynthases (GSs) are powerful biocatalysts for custom glycan synthesis, yet their optimization via directed evolution is severely constrained by bottlenecks in high-throughput screening for activated azido-sugar donors. Here, we demonstrate that chemical rescue (CR)--the azide-mediated restoration of hydrolytic activity in nucleophile-deficient mutants--serves as a predictive, high-throughput proxy for glycosynthase activity. Applying an azide-responsive Escherichia coli biosensor screen to a site-saturation mutagenesis library of Thermotoga maritima -L-fucosidase (TmAfc), we established a strong rank-order correlation between CR and GS activities in both crude lysates ({rho} = 0.73) and purified enzymes ({rho} = 0.95). Transition path sampling and QM/MM umbrella sampling revealed that both pathways proceed through a shared oxocarbenium-ion-like transition state ({Delta}G{ddagger} {approx} 8.7 kcal/mol), providing a structural and thermodynamic rationale for using CR to select for transition-state-stabilizing mutations. Biochemical characterization of top-performing variants yielded an engineered fucosynthase (TmAfc_D224G_N70D_T392S) exhibiting a nearly 100-fold enhancement in Vmax alongside altered regioselectivity. This two-tiered screening framework leverages cost-effective chemical rescue assays to streamline glycosynthase engineering for tailored glycans synthesis.

biochemistry↗

bioRxiv2026

Exosome-Derived Proteomic Signatures Highlight Pathogenic Mechanisms in Moyamoya Disease

BackgroundThe etiology and molecular mechanisms of Moyamoya disease (MMD) remain unclear. Exosomes, as carriers of bioactive molecules, may reflect disease-specific alterations and serve as potential biomarkers. This study aimed to investigate disease mechanisms using proteomic profiling of serum-derived exosomes (SDEs) in MMD. Materials and MethodsPeripheral blood each from 15 MMD patients and 15 healthy-controls were used to isolate SDEs via ultracentrifugation. Proteins from pooled SDEs were extracted, digested, and analyzed by LC-MS/MS. Differentially expressed proteins were examined using MetaboAnalyst, DAVID, Enrichr, STRING, and Cytoscape. Key targets were validated at transcript and protein levels using RT-qPCR and ELISA in independent cohorts. ResultsA total of 2,554 proteins were identified, with 213 showing differential expression (118 upregulated, 95 downregulated; p [≤] 0.05). Functional and pathway analyses revealed enrichment in angiogenesis, cytoskeletal remodeling, and endothelial signaling. PRKG2 and MYC were upregulated, while RHOA was downregulated, highlighting their involvement in focal adhesion and PI3K-AKT pathways. Validation confirmed these findings. ConclusionDysregulated proteins were linked to RHOA-ROCK and PI3K-Akt signaling, suggesting their role in driving VSMC phenotypic switching, contributing to vascular occlusion. These findings indicate that altered exosomal-proteins may participate in maladaptive vascular remodeling, although the initial trigger for VSMC transition remains unknown.

systems biology↗

bioRxiv2026

Immunoproteasome Deficiency Impairs Microglial Clearance and Worsens Tau and Amyloid Pathology

Immunoproteasome induction is prominent in Alzheimers disease (AD), but whether it protects proteostasis or amplifies neuroinflammation remains unresolved. Here, we generated immunoproteasome-deficient PS19 tauopathy and APP/human tau double-knock-in mice by crossing each disease model with L7M1 mice lacking two immunoproteasome catalytic subunits. Immunoproteasome deficiency increased phospho-tau burden, exacerbated amyloid-{beta} pathology and heightened microglial reactivity without suppressing constitutive 26S proteasome activity. In primary microglia and longitudinal two-photon imaging, immunoproteasome-deficient microglia engaged and engulfed tau aggregate-bearing material but failed to resolve internalized cargo, revealing a post-engulfment degradative checkpoint. Single-nucleus transcriptomics identified a remodeled P2ry12low/Trem2high microglial state with impaired phagolysosomal and mitochondrial programs. Reanalysis of human single-nucleus transcriptomic datasets showed that reduced microglial immunoproteasome expression was associated with cargo-processing gene-program changes similar to those observed in immunoproteasome-deficient mouse microglia. Together, these findings identify immunoproteasome biogenesis as a protective glial stress response that supports microglial aggregate clearance in AD.

neuroscience↗

bioRxiv2026

SentryPath: a mechanistic protocol-ranking simulator with leave-one-trial-out cross-validation across 13 phase-III oncology randomised controlled trials and a pre-registered prospective forecast

BackgroundPivotal oncology trials cost a median of {approx}$19 million each (oncology often $45 million or more) and contribute to a capitalised cost of {approx}$2.6 billion per approved drug, yet most candidate protocols never reach trial. Existing in-silico screening tools either rely on closed proprietary PK/PD modelling or require patient-level data; a transparent, cohort-level, cross-validated mechanistic alternative is missing. MethodsSentryPath is a physics-based stochastic differential equation simulator built on a Gompertzian tumour-growth term with Emax pharmacodynamic kill and Bliss-independence combination modelling, scored at the cohort level. Validation against 13 published phase-3 randomised controlled trials covering six cancer types uses the 2-year overall-survival (OS) rate ratio as the primary endpoint, cross-checked against ClinicalTrials.gov posted results. For cancer types with [≥]2 trials we apply leave-one-trial-out crossvalidation: two shared efficacy scalars per cancer type are fit on training trials and used to predict the heldout trial cold. ResultsWith the per-drug efficacy proxies held fixed from the literature, two shared cancer-type scalars fit on the training trials transfer to the held-out trial with a mean held-out error of 3.7 % (range 0.7-7.3 %) on 2-year OS rate ratios across three NSCLC trials; extending the same method to RCC, HCC, and ESCC yields a 5.4 % aggregate across nine folds (per-fold range 0.2-11.2 %), reported with per-cancer stratification. We are explicit that only the two scalars are held out -- the per-protocol efficacy proxies underneath are literature-anchored to drug classes that include the benchmark trials, so this is a test of scalar transfer, not of the whole engine cold. Cross-validation improves on the same engine without it (16.4 % with production cancer priors; 21.9 % with no efficacy modifiers); a matched in-sample fit of the same two-scalar model gives 4.4 %, slightly below the 5.4 % held-out, the expected direction. Two prospective forecasts are preregistered on the Open Science Framework with falsification envelopes and pre-readout bias disclosure. The first forecast (NCT04770896) reaches its primary data cutoff on 2026-06-30; the observed outcome and its mapping to the pre-committed interpretation will be reported in a versioned update to this preprint. ConclusionA transparent mechanistic simulator, with a literature-anchored efficacy library and only two cross-validated scalars per cancer type, transfers those scalars across held-out NSCLC trials at 3.7 % mean error (range 0.7-7.3 %) and extends to other cancers with documented per-cancer stratification. The validation is pilot-scale (3-9 folds) and the scalars sit on a fixed, trial-informed substrate; its distinguishing contribution is less the error magnitude than the public predict-verify-disclose cycle that goes beyond retrospective fit.

bioinformatics↗

bioRxiv2026

RubiSpec-MS: Determination of Rubisco CO 2 /O 2 Specificity Factor using Liquid Chromatography-Mass Spectrometry

O_LIThe specificity factor(SC/O) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is a key kinetic parameter quantifying the ratio of carboxylation to oxygenation reactions. Current methods to measure SC/O are expensive, labor-intensive, require specialized expertise, and/or rely on radiolabeled compounds, hindering Rubisco research. C_LIO_LIWe present RubiSpec-MS (Rubisco Specificity by LC-MS/MS), a method to determine the SC/O of purified Rubiscos using liquid chromatography-coupled mass spectrometry (LC-MS/MS) for separation and sensitive detection of non-radioactive reaction products. C_LIO_LIWe benchmark RubiSpec-MS using multiple model Rubisco enzymes. We find that the assay accurately reproduces corresponding literature values for each enzyme and performs similarly in a side-by-side comparison using an established radiometric method. C_LIO_LIRubiSpec-MS provides a faster, more accessible, and highly parallelizable method to measure Rubisco SC/O using equipment common in academic core facilities and industrial laboratories. C_LI

Plant Biology↗

bioRxiv2026

Generative design of programmable asymmetric β-barrel nanopores

Protein nanopores are powerful tools for molecular sensing, sequencing, and separation, but designing pores with programmable function remains challenging. Native homo-oligomeric transmembrane {beta} barrels (TMBs) are used for these applications, but their uniform lumens limit spatial resolution and analyte discrimination. Although monomeric TMBs can be designed using energy-based methods, these approaches remain highly manual and limited to structural design rather than function. Here, we present a generative AI framework for TMB design, with backbone and sequence design models trained on a curated distillation set. We characterized 48 designs spanning 0.7-1.5 nm in diameter, diverse lumen chemistries, and hydrophobic thicknesses. Crystal structures closely match the design models. We demonstrate that our method produces customizable nanopores for ion sensing, DNA translocation, and transport across polymer membranes.

biochemistry↗

bioRxiv2026

Dihydroartemisinin inhibits Epstein-Barr virus reactivation and replication targeting lytic proteins: insights for drug repurposing

Epstein-Barr virus (EBV) is an oncogenic virus which is responsible for various malignant as well as non-malignant diseases and leads to about 200,000 deaths each year. Despite efforts, there are no FDA-approved drugs targeting EBV. Reactivation of EBV plays a critical role in the transition from latency to lytic cycle, leading to viral replication and disease progression, and is primarily regulated by the transactivator BZLF1. In this study, we combined computational screening with experimental validation to identify repurposing drugs that inhibit EBV reactivation and replication. FDA-approved compounds predicted using in-house AI/ML-based model (Anti-EBV) and miRNA-seq and RNA-seq analyses, were selected for further evaluation. Molecular docking against BZLF1, supported by in silico alanine scanning to identify critical DNA-binding residues, led to the selection of seven candidate drugs. Among these, an antimalarial drug, dihydroartemisinin (DHA), showed the strongest inhibitory activity in vitro, with an IC99 of 1 {micro}M and an SI Index of 113.5. DHA reduced both EBV viral copy number and the expression of early and late lytic genes. Molecular docking and simulation studies demonstrated stable binding of DHA within the BZLF1 DNA-binding pocket, inhibiting the key residues involved in BZLF1 activation and DNA binding. Analysis at the gene level confirmed its inhibitory effect on EBV replication, while expression analysis at the transcriptional and protein levels, along with immunofluorescence analysis, indicated its inhibitory effect on EBV reactivation and virion assembly. These findings suggest DHA as a promising repurposing antiviral candidate targeting EBV lytic proteins and offers an effective target-based therapeutic strategy. ImportanceThis study identifies a repurposed small-molecule inhibitor of EBV reactivation and replication. Here, we proposed target-based therapy, integrating computational and experimental approaches to target the EBV lytic transactivator BZLF1. Since early lytic EBV protein BZLF1 plays a critical role in viral reactivation and replication, inhibition of its activation and DNA-binding function represents a promising therapeutic approach to prevent EBV infection. Molecular docking and simulation studies revealed stable binding of DHA within the BZLF1 DNA-binding pocket. Furthermore, in vitro analyses demonstrated significant inhibition of viral gene copy number and reduced mRNA and protein levels of key lytic proteins. Thus, this study demonstrated DHA as a safe and effective repurposed therapeutic candidate against EBV infection.

microbiology↗

bioRxiv2026

Accelerated amyloid neurodegeneration in HIV-1-infected APP-KI Alzheimer's disease mice

STRUCTURED ABSTRACTO_ST_ABSINTRODUCTIONC_ST_ABSA higher incidence of dementia, including Alzheimer s-like pathology, is observed in aged people living with HIV-1. However, mechanisms linking HIV-1 to Alzheimers disease (AD) pathology remain unclear, due to the lack of animal models that allow for concurrent study. METHODSWe created a novel APP knock-in (KI) AD mouse, NOG/APPKM670,671NL/IL-34 (hNAIL) that permits study of progressive brain HIV-1 replication. The mice harbor human microglia-like cells. Four-month-old CD34+ human cell reconstituted mice infected with the HIV-1ADA strain facilitated studies of HIV-1 replication on AD pathologies. RESULTSHIV-1 replication increased A{beta} levels and reduced synaptic and neuronal integrity. Spatial transcriptomics demonstrated distinct A{beta} and HIV-1 transcriptional patterns, whereas dual diseased combinations amplified AD pathology. Neurons showed highest transcriptional change, with genes linked to neuroinflammation, protein trafficking, and synaptic dysfunction. DISCUSSIONThe hNAIL mice enable interrogation of HIV-AD comorbidities, with a future potential for the development of novel therapeutic interventions.

neuroscience↗

bioRxiv2026

Computational drug repurposing identified Artemisinin and Mebendazole as potential inhibitors of virulence-associated proteins SKSR and essential kinases CpCDPK1 of Cryptosporidium parvum

Cryptosporidium parvum is a protozoan parasite responsible for cryptosporidiosis, significantly threatening immunocompromised individuals, particularly HIV/AIDS patients, by causing severe diarrhea and potential mortality. Current treatments are largely ineffective, prompting investigations into new therapeutic options. This study evaluated two antiparasitic drugs: Mebendazole, used for helminth infections, and Artemisinin, used for malaria. The SKSR gene family encodes virulence factors in C. parvum, and Calcium-dependent protein kinase1 (CpCDPK1) regulates the life cycle of C. parvum; targeting these proteins may reduce growth and infection in hosts. In the current study, molecular docking was conducted taking Mebendazole and Artemisinin drugs as ligands, SKSR gene family and CpCDPK1 proteins as drug targets. Results with SKSR showed binding energy of -4.9 kcal/mol, -6.72 kcal/mol for Mebendazole and Artemisinin, respectively. Whereas, with CpCDPK1, the binding energies were -6.44 kcal/mol, -9.18 kcal/mol for Mebendazole and Artemisinin, respectively. Docking of Nitazoxanide (an in-use drug for C. parvum) with SKSR and CpCDPK1 revealed binding energies -4.2 kcal/mol, -4.81 kcal/mol, respectively. The stability of the proteins (targets) upon binding to the ligands was assessed by performing all-atom MD simulations for 100ns using the GROMACS package. No major variations were observed upon binding of Artemisinin and Mebendazole to SKSR and CpCDPK1. The findings of MD simulations imply that both proteins maintain their stability upon binding of Artemisinin and Mebendazole. Molecular Docking and MD simulation studies suggest that Artemisinin and Mebendazole are potential candidates for repurposing in the treatment of C. parvum infections, with recommendations for in vitro studies to validate these findings.

molecular biology↗

bioRxiv2026

Comprehensive identification and characterization of candidate effector proteins in Puccinia triticina reveals insights into the wheat leaf rust pathogenesis

The biotrophic pathogen Puccinia triticina is the causative agent of the most vulnerable foliar disease, namely leaf rust disease of wheat. The pathogen-secreted effectors are essential in modulating fungal virulence and host immune responses. Despite their significance, potential effectors and their underlying mechanisms governing host susceptibility remain elusive. In the present study, we employed an in silico approach to identify and characterise effector proteins from the P. triticina proteome. Later, performed temporal expression profiling to prioritise effector candidates associated with rust disease. Here, a total of 273 high-confidence effector candidates were identified and analysed their physicochemical properties, domains, motifs, and functional annotations, to assess their conservation and dynamics. Although most of the effectors were uncharacterised, the conserved motif virulence-associated [YFW]xC was notably enriched in the effector repertoire. Comparative PHI-base annotation highlighted similarities with known fungal virulence factors involved in host susceptibility. Effectors harbouring CAZyme activity indicate involvement in host cell wall modification. Promoter analysis identified multiple stress- and defence-related transcription factor binding sites, suggesting regulated expression during infection. Transcriptome analysis revealed that 20 effector genes were significantly upregulated during P. triticina infection. qRT-PCR validated the expression of 4 highly induced effector transcripts following P. triticina infection in susceptible wheat variety. Specifically, two of these candidates demonstrated biphasic expression pattern that aligns contrasting PTI- and ETI-mediated defense mechanisms critical for sustained virulence. Overall, this study provides a comprehensive framework for identifying functionally relevant P. triticina effectors and offers insight for future effector-target studies and effector-based leaf rust management strategies.

plant biology↗

bioRxiv2026

All-trans retinoic acid recalibrates macrophage transcriptional responses to drug-resistant Mycobacterium tuberculosis through strain-specific immunometabolic reprogramming

Tuberculosis (TB) caused by Mycobacterium tuberculosis (M.tb) remains the leading infectious cause of death worldwide, with multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains presenting an escalating therapeutic crisis. All-trans retinoic acid (ATRA), an active vitamin A metabolite with established immunomodulatory properties, has emerged as a candidate host-directed therapy, yet its genome-wide transcriptional effects on macrophage across strains of differing drug-resistance profiles remain uncharacterized. We performed RNA sequencing of murine peritoneal macrophages infected with drug-susceptible H37Rv, MDR-2261, or XDR-MCY M.tb strains, with and without ATRA treatment, validated by RT-PCR and flow cytometry. All three strains activated a conserved pro-inflammatory program dominated by Nos2, Il1b, and Acod1 induction with progressive suppression of host translational and mitochondrial machinery. XDR-MCY uniquely induced Ifnb1/IFN-{beta}, suppressed Prdx1 and Gpx4, the latter shared with MDR-2261 and showed selective downregulation of MHC-II processing genes, suggesting a multilayered immune evasion strategy. ATRA activated canonical retinoid signaling across all infection states and consistently induced Arg1-mediated resolution signaling, with Nos2 suppression observed in H37Rv- and MDR-infected macrophages. ATRA selectively restored Epas1/HIF-2 in H37Rv- and MDR-infected macrophages without disrupting the HIF-1-driven antimicrobial program or itaconate biosynthesis. ATRA responsiveness progressively attenuated with drug resistance, with XDR-MCY-infected macrophages largely refractory to transcriptional reprogramming. These findings provide a transcriptional rationale for evaluating ATRA as a host-directed adjunct in drug-resistant TB and identify Gpx4 suppression in drug-resistant strains and XDR-specific Ifnb1 induction as vulnerabilities necessitating further investigation.

microbiology↗

bioRxiv2026

TSC2 acting as a transcription factor for miR-514b-3p and regulating PI3K-AKT-MTOR pathway via nucleus

The PI3K-AKT-MTOR signalling pathway is pivotal in regulating cell survival, proliferation, and growth. TSC2 or tuberin acts as a key negative regulator of this pathway by forming a cytoplasmic complex with TSC1 and TBC1D7. While the cytoplasmic role of TSC2 is well established, emerging evidence suggests its nuclear functions. Previously, we have identified TSC2 as a transcriptional repressor of the EREG gene. Building on this foundation, the present study has investigated the transcriptional role of TSC2 in miRNA gene regulation. A genome-wide miRNA microarray profiling of TSC2-depleted SCC131 cells identified 19 upregulated and 24 downregulated miRNAs. Of these, miR-514b-3p emerged as one of the most upregulated miRNAs. The RT-qPCR analysis showed that TSC2 knockdown results in a robust miR-514b-3p upregulation, whereas TSC2 overexpression suppresses its expression in SCC131 cells. Moreover, TSC2 negatively regulates the MIR514B promoter activity in an NLS-dependent manner. Our ChIP analysis confirmed the direct binding of TSC2 to the MIR514B promoter, establishing miR-514b-3p as a transcriptional target of TSC2. We further identified TSPAN9 as a direct downstream target of miR-514b-3p. TSC2 positively regulates TSPAN9 levels by repressing miR-514b-3p, thereby establishing a novel TSC2/miR-514b-3p/TSPAN9 regulatory axis. We further uncovered a crosstalk between TSC2/miR-514b-3p/TSPAN9 axis and the canonical PI3K-AKT-MTOR signalling pathway, where miR-514b-3p positively and TSPAN9 negatively regulates this pathway. Interestingly, AKT functions as an upstream regulator of this axis by modulating the nuclear localization of TSC2. Collectively, this study provides new insights into the non-canonical, nucleus-dependent transcriptional functions of TSC2, thus expanding its role beyond cytoplasmic signalling regulation and underscoring its significance in the cellular signalling networks. Graphical abstractThe diagrammatic representation of the TSC2/miR-514b-3p/TSPAN9 axis and its interaction with the canonical PI3K-AKT-MTOR pathway. Abbreviations: PI3K, Phosphatidylinositol-3 kinase; TSC1, Tuberous sclerosis complex subunit 1; TSC2, Tuberous sclerosis complex subunit 2; TBC1D7, Tre2-Bub2-CDC16 domain family member 7; RHEB, Ras homolog enriched in brain; GTP, Guanosine tri phosphate; GDP, Guanosine di phosphate; MTORC1, Mechanistic target of rapamycin kinase complex 1; S6K1, p70 ribosomal protein S6 kinase; 4E-BP-1, eIF4E-binding protein 1; and TSPAN9, Tetraspanin 9. This figure was created in BioRender (https://app.biorender.com/).

cancer biology↗

bioRxiv2026

TOXsiRNA: A web server to predict the toxicity of chemically modified siRNAs

Small interfering RNAs (siRNAs) are largely modified with chemical molecules to enhance their properties for use in molecular biology research and therapeutic applications. Toxicity effects may arise due to these chemical moieties as well as sequence based off-targets at cellular level. Enormous resources are required to experimentally design and test the toxicity of these chemical modifications and their combinations on siRNAs. To address this problem, we developed TOXsiRNA web server to computationally predict the toxicity of chemically modified siRNAs and their off-targets. We selected 2749 siRNAs with different permutations and combinations of 21 different chemical modifications engineered on them. Next, we used Support Vector Machine (SVM), Linear Regression (LR), K-Nearest Neighbor (KNN) and Artificial Neural Network (ANN) machine learning applications to develop models. Best performance was displayed by mononucleotide composition-based model developed with SVM, offering Pearson Correlation Coefficient (PCC) of 0.91 and 0.92 on training testing and independent validations respectively. Other sequence features like dinucleotide composition binary pattern and their combinations were also tested. Finally, three models of chemically modified siRNAs were implemented on the web server. Other algorithms that include predicting normal as well as chemically modified siRNA knockdown efficacy, off target etc. are also integrated. The resource is hosted online for scientific use freely at url: http://bioinfo.imtech.res.in/manojk/toxsirna.

bioinformatics↗

bioRxiv2026

PEPsRNA: A computational resource of experimentally tested peptide based siRNA delivery

In siRNA-based applications, cellular delivery remains one of the main hurdles. Many formulations are tested for the same and peptides came up as one of the optimal options. The latter have various advantages like natural biological presence, high specificity, and natural metabolism etc. siRNA in conjugation with peptides have exhibited enhanced mRNA silencing. Peptides aid siRNAs in condensation to smaller volumes, enhance nuclease protection, increase half-life, promote cell specific binding as well as endosomal escape and release in cytosol. Despite its prime importance, no resource is available for the peptide-based delivery of siRNAs, therefore to fill the gap we developed PEPsRNA web server. It includes 2266 entries of 270 different kinds of peptides, 106 different types of siRNAs and shRNAs along with more than 80 conjugate molecules targeting 55 different genes, experimentally tested for the delivery of the siRNAs. To provide the detailed insights of the procedure, we have incorporated analysis of the peptides (e.g. secondary structure, amino acid composition, polarity, hydrophobicity etc.), siRNAs (e.g. secondary structures with minimum free energies etc.) and associated conjugate molecules (e.g. structure, SMILES, Inchl). We have derived these values using various other tools and resources to make the web server comprehensive. We further compared various physicochemical properties with the efficacy of the peptide based on the target gene silencing, but these properties do not shown any distinct conclusive relationship. The data is available for browsing, searching and downloading freely on the web server with URL: http://bioinfo.imtech.res.in/manojk/pepsirna. Highlights PEPsRNA is the first database of experimentally tested peptides for siRNA delivery It comprised of 2266 entries with 270 peptides and about 80 conjugate molecules Analysis of peptides, siRNAs and details of conjugate molecules are provided Browse, search and various tools are incorporated for data retrieval and usage

bioinformatics↗

bioRxiv2026

Stem cell control and cancer initiation by an autocrine, injury-activated Igf complex

Stem cells rapidly proliferate after injury to repair damaged tissue, and chronic injury predisposes to cancer. However, injury-activated mitogens, the mechanisms that keep them inactive until injury, and their role in cancer are not understood. Here we identify Igf2 as the injury-activated mitogen for neuroendocrine stem cells, a facultative airway stem cell and origin of small cell lung cancer. Igf2 is constitutively produced by the stem cells but sequestered in inactive form by co-expressed Igf binding proteins. Injury releases Igf2 and induces proliferation by activating its receptors and repressing Rb tumor suppressor, which normally enforces stem cell quiescence. Persistent pathway activation initiates oncogenesis. Thus, in addition to its classical hormonal roles in physiology, growth, and aging, Igf operates locally with Igf binding proteins and Rb to control injury-induced stem cell activation and cancer. This pathway may also control related stem cells and cancers of the body and brain.

cell biology↗