Biology subjects

Singh, P.

Publications and source records attributed to Singh, P..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Metatranscriptomic analysis reveals toxin-antitoxin system shifts in caries-associated oral microbiomes

BackgroundToxin-antitoxin (TA) systems are bacterial regulatory elements involved in persistence, dormancy, and biofilm stability under stress conditions such as limited nutrients, acid exposure, and antimicrobial treatment. Although their roles are well-characterized in isolated pathogens, TA systems remain largely unexamined in the context of the oral microbiome. Given their potential involvement in microbial adaptation during oral diseases and treatment, we aimed to identify transcriptionally active TA systems and evaluate their condition-specific expression patterns across oral health states using metatranscriptomic data. ResultsWe re-analyzed two publicly available supragingival metatranscriptomic datasets: a longitudinal dataset by Carda-Dieguez et.al. (Dieguez) capturing treatment transitions (fluoride (Fl), fluoride-arginine (Fl-Ar)) in caries-active and caries-free individuals, and a cross-sectional dataset by Ev et.al., (Ev) profiling caries-active lesions (CA), caries-inactive (CI), and caries-free (CF) states. After quality control, host sequences removal, rRNA removal, and HUMAnN3-based functional profiling, we identified 1,189 unique UniRef90 gene clusters with known TA associations. Differential analysis using ANCOM-BC revealed 197 TA-related gene clusters as significantly modulated in at least one condition. Of these, 22 gene clusters were differentially expressed in Dieguez, 38 in Ev, with 77 shared across both datasets. Distinct TA expression signatures were observed across healthy (CF, CI, baseline), lesion types (CAa, CAi, CAs), and treatment stages (Fl, Fl-Ar), suggesting condition-specific regulatory activity. Functional annotation of differentially expressed TA genes using the Toxin-antitoxin system database, eggNOG-mapper, and InterProScan, revealed 18 genes with strong evidence for their involvement in key bacterial stress-response pathways. These included members of the ParD-ParE, RelE/StbE-RelB/StbD, FitA, and YafQ families, which are annotated with mRNA catabolism, transcriptional repression, and prokaryotic defense-associated pathways. ConclusionThis is the first metatranscriptomic analysis profiling functional shifts in toxin-antitoxin systems across oral healthy, caries, and treatment states. TA systems showed dynamic and condition-specific expression, with pathway annotations suggesting their roles in microbial persistence, stress adaptation, and ecological remodeling during caries progression. These findings open new avenues for targeting microbial stress modules in precision microbiome therapeutics.

bioinformatics↗

bioRxiv2025

A Shared Amyloid Architecture in Cardiac Fibrils from Three Neuropathy-Associated ATTR Variants

ATTR amyloidosis results from the systemic accumulation of wild-type (ATTRwt) or mutant (ATTRv) transthyretin amyloids, leading to multi-organ dysfunction and death. The disease exhibits variable pathology and penetrance, and its relationship with the amyloid structure remains unclear. Patients carrying the neuropathy-associated variants ATTRvI84S and ATTRv-V122{Delta} present polymorphic ATTR fibrils, in contrast to the consistent morphology reported for most ATTR fibrils to date. Here, we aim to elucidate a potential link between neuropathic symptomatology, distinct mutations, and amyloid structural diversity, using cryo-EM. We determined the ex-vivo fibril structures from the variants ATTRv-P24S, ATTRv-A25S, and ATTRv-D38A, whose patients presented variable clinical manifestations, including neuropathy. Our findings revealed that, despite differences in mutations and diverse clinical phenotypes, these variants share a common amyloid core previously identified in ATTRwt and several other cardiac ATTRv. This structural consistency is significant for the development of structure-guided diagnostic tools capable of addressing the diverse spectrum of ATTR amyloidosis. HighlightsO_LIDetermines transthyretin amyloid structures of three human ATTRv by cryo-EM C_LIO_LIDetermines the structure of three ex-vivo ATTRv fibrils linked to polyneuropathy. C_LIO_LIReveals structural similarities of ATTRv amyloid cores. C_LIO_LIReveals a common fold despite the different mutations and symptomatology. C_LIO_LIContributes to the understanding of transthyretin aggregation in patients with diverse phenotypes C_LI

biophysics↗

bioRxiv2025

Reversible therapeutic resistance in EGFR mutant lung cancer caused by RB1 loss induced lineage plasticity

Treatment with EGFR tyrosine kinase inhibitors (EGFR TKI) is standard of care for patients with lung cancer initiated by activating mutations in the EGFR gene. While EGFR TKI treatment is effective, virtually all patients progress on therapy. Loss of function mutations in the RB1 tumor suppressor gene are associated with poor EGFR TKI outcomes, but underlying mechanisms remain unclear. Analysis of genetically engineered experimental models and human clinical specimens described here demonstrates RB1 loss has minimal impact on initial cell cycle responses to EGFR TKI but markedly accelerates acquired resistance over time. RB1 loss promotes transcriptional plasticity that enables rare cells to spontaneously assume reversible, resistance-conferring lineage states. These findings highlight the importance of non-genetic adaptations in driving acquired EGFR TKI resistance, advance our understanding of how RB1 impacts cancer, and identify opportunities to improve EGFR TKI treatment outcomes by targeting reversible epigenetic resistance mechanisms.

cancer biology↗

bioRxiv2025

Structural polymorphism of ex-vivo ALECT2 amyloid fibrils revealed by cryo-EM

ALECT2 amyloidosis is a rare systemic disease characterized by the pathological deposition of leukocyte cell-derived chemotaxin-2 (LECT2) as amyloid fibrils, primarily affecting the kidneys and liver. The molecular mechanisms underlying LECT2 aggregation remain poorly defined, hindering diagnostic and therapeutic development. Here, we present cryo-electron microscopy structures of ex-vivo ALECT2 fibrils extracted from a patients kidney. We identified three fibril polymorphs: a predominant single-protofilament morphology and two minor double-protofilament morphologies. The dominant single-protofilament morphology comprises the full-length 133-residue LECT2 protein and retains all three native disulfide bonds. Low-resolution reconstructions of double-protofilament morphologies suggest they adopt a similar fold to the single protofilament morphology, but form paired assemblies with different inter-filament interfaces. Mass spectrometry also reveals acetylation within the fibrils. These findings offer critical insights into the structural basis of ALECT2 amyloid formation and identify molecular features that could inform future diagnostic and therapeutic approaches.

biophysics↗

bioRxiv2025

Structural variability of apolipoprotein A-I amyloid fibrils across organs, mutations, and clinical presentations, revealed by cryo-EM

Hereditary apolipoprotein A-I (AapoA-I) amyloidosis is a rare systemic disease caused by the deposition of amyloid fibrils formed by apolipoprotein A-I in multiple organs, leading to severe clinical outcomes. With no available therapies or diagnostic tools, defining the structure of AApoA-I fibrils is crucial to understanding disease mechanisms and guiding intervention. Using cryo-electron microscopy, we analyzed AApoA-I fibrils from the heart, kidney, liver, and spleen of patients carrying G26R, L90P, and R173P mutations. G26R fibrils, regardless of organ, exhibited untwisted morphologies and could not be resolved structurally. Conversely, L90P and R173P fibrils displayed a compact diabolo-shaped conformation in all organs analyzed. Their high-resolution maps enabled visualization of cis-Proline 66, which may represent a potential conformational switch during fibril formation. Our findings suggest that mutation-driven polymorphism may influence organ tropism and clinical presentation. This work advances our understanding of AapoA-I fibril assembly and provides insights toward developing targeted clinical tools.

biophysics↗

bioRxiv2025

Highly modular genomic architecture underlies combinatorial mechanism of speciation and adaptive radiation

Hybridisation can fuel rapid adaptive radiation, but how it enables the formation of phenotypically highly dimensional species-rich radiations remains unclear. We investigated this by analysing genotype-phenotype associations for 14 ecological (trophic, body patterns) and mating traits (nuptial colour) across 107 species of Lake Victoria cichlid fishes. We find weak trait covariance across the radiation, with many different trait combinations constituting different species. Across the radiation, polygenic, redundant, and lowly pleiotropic genomic architectures of hybrid origin underlie the repeated evolution of key traits. Such independent genomic modules can be reshuffled and recombined like Lego bricks, generating diverse trait combinations from a finite number of elements. During speciation, dispersed oligogenic trait modules become coupled through long-range linkage disequilibrium. We propose that this genomic and phenotypic modularity emerged from repeated cycles of past hybridisation, enabling superfast adaptive radiation through combinatorial speciation.

evolutionary biology↗

bioRxiv2025

Behavioral variation affects persistence of an experimental food-chain

Intraspecific behavioral variation in prey could alter predator-prey interactions, yet its effects on temporal dynamics and food-web persistence remain underexplored. Pea aphids (Acyrthosiphon pisum) exhibit dropping behavior in response to predators like the seven-spot ladybird (Coccinella septempunctata). This response could be an effective anti-predator defense but could be costly in terms of energy expenditure and time not available for feeding. To investigate the impact of behavioral variation on food-chain persistence and dynamics, we used a tri-trophic experimental system with Vicia faba (plant), pea aphids (prey), and seven-spot ladybirds (predator), implementing three aphid behavioral treatments: droppers, non-droppers, and a mix of both droppers and non-droppers. To minimize genetic differences, we used clonal aphid populations across all treatments. We then tracked predator-prey population dynamics and species persistence over 25 days. Our results showed that aphid dropping behavior reduced food-chain persistence, with extinction risk significantly higher in dropper treatments than in the mix or non-dropper treatments. Ladybirds persisted across treatments, although they showed a steeper decline in abundance in the dropper treatment. In the mixed behavioral treatment, they had an intermediate persistence, suggesting a buffering effect of behavioral variation. Trophic food-chain state transitions also differed by treatment, with tri-trophic states most stable in the non-dropper, and least frequent in the dropper treatment. Furthermore, our results showed a trend of dropper treatments becoming more stable and robust towards the end of the experiment. These results demonstrate that prey behavior influences the persistence and dynamics of food-chains, with important implications for behavior-driven community dynamics.

ecology↗

bioRxiv2025

Neural Network-Enhanced Investigation of Ferroptosis and Druggability in Early-Onset Alzheimers Disease

BackgroundAlzheimers disease (AD) is a complex neurodegenerative disorder which is multifactorial in nature. Some of its characteristics are slow cognitive decline, memory problems and behavioral changes. AD patient brains show a progressive synaptic toxicity, autophagy, neuroinflammation, excess generation of reactive oxygen species (ROS), neuronal death and oxidative stress, which occurs due to disrupted metal homeostasis along with tau and amyloid-{beta} protein deposition. Notably, lipid peroxidation, iron buildup and elevated oxidative stress in AD brains suggest a possible molecular connection between ferroptosis and AD neurodegeneration. MethodsThis study explores the genetic and bioinformatics perspective on the relationship between ferroptosis and AD aiming to identify potential therapeutic biomarkers using Neural network (NN) and Machine learning models. Six ferroptosis related genes were found to be differentially expressed in AD. Further machine learning analysis shortlisted four key biomarker genes. An NN-based diagnostic prediction model was developed and validated using AUC-ROC anaysis, which gave high diagnostic values (AUC-0.92) in the analysis. ResultsThe findings highlight a strong correlation between ferroptosis and altered metabolic functions in AD. miRNA-gene interaction analysis revealed that two biomarker genes, CYBB and ACSL4 can be regulated by several regulatory miRNAs i.e., hsa-miR-146-5p, hsa-miR-106b-5p, hsa-miR-223-3p, hsa-miR-155-5p, hsa-miR-34a-5p, hsa-miR-125b-5p and hsa-miR-27a-3p suggesting their potential as early diagnostic biomarkers. Immune microenvironment analysis revealed strong neuroinflammatory responses were in AD with increased infiltration of macrophages (M0, M1 and M2), monocytes and multiple T cell subsets. This heightened immune activity may be driven by ferroptosis-induced oxidative stress, contributing to neuronal death. Furthermore, druggability of these targets was evaluated and several drugs were identified that may be potentially repurposed for therapeutic intervention in AD pathogenesis. ConclusionThis study presents a diagnostic predictive model integrating gene expression, miRNA regulation and immune infiltration analysis, offering a novel perspective on early AD detection. The identified ferroptosis-related biomarkers and regulatory miRNAs could serve as valuable tools for clinical diagnosis and targeted therapeutic intervention, advancing personalized treatment strategies for Alzheimers disease.

genomics↗

bioRxiv2025

Phosphorylation-inducing chimera rewires oncogenic kinase to trigger apoptosis

The active sites electric field is integral to enzymatic catalysis (e.g., substrate recognition) and nature employs charge-altering post-translational modifications (e.g., phosphorylation) to perturb this electric field and regulate enzymes. A chromosomal translocation converts Abelson kinase (ABL) to BCR-ABL, whose hyperactivity drives several cancers. Here, we developed a small molecule, BRD8833, that induces BCR-ABL phosphorylation, which perturbs its active sites electric field with loss of hyperactivity. Unlike "occupancy-driven" inhibitors that require stoichiometric concentrations, BRD8833 operates through an event-driven, substoichiometric mechanism by inducing proximity between two BCR-ABL molecules to trigger the inhibitory phosphorylation and selective apoptosis of BCR-ABL-dependent cancer cells. Furthermore, BRD8833 is effective against other oncogenic ABL fusions or clinically observed resistance mutations, including those to occupancy-driven drugs with the same binding site as BRD8833, suggesting differences in their resistance mechanisms. These studies lay the foundation for electric-field and "event-driven" modalities to control hyperactive enzymes with orthogonal resistance mechanisms to occupancy-driven drugs. GRAPHICAL ABSTRACT (TOC) O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=197 SRC="FIGDIR/small/659082v2_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1607a40org.highwire.dtl.DTLVardef@fa10deorg.highwire.dtl.DTLVardef@1698232org.highwire.dtl.DTLVardef@1e2e0f1_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2025

Desiccation-induced DNA damage facilitates drug resistance in Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) is an obligate human pathogen that depends on its ability to spread from host-to-host to survive as a species. Yet, knowledge of transmission-specific traits remains lacking. Here, we report the discovery of a specific adaptive response to desiccation, a stress intrinsically linked to the generation of the aerosol droplets within which Mtb transmits. We show that desiccation inflicts oxidative damage and activates Mtbs DNA repair responses but that this repair is imperfect and results in mutations. We further show that activation of these DNA repair responses is accompanied by increased expression of the transcription-coupled repair factor, mfd, but that this expression serves to buffer the fitness cost of specific resistance-conferring mutations in rpoB, the target of the frontline drug rifampin, rather than to facilitate transcription-coupled DNA repair. Silencing mfd during aerosolization impairs survival of strains harboring the rifampin resistance allele S450L. This function is further supported by whole genome sequence data from over 50,000 clinically circulating strains. These studies indicate that Mtb has evolved transmission-specific stress responses that have enabled it to leverage desiccation-induced DNA damage as a potential source of genetic diversification and drug resistance.

microbiology↗

bioRxiv2025

Graphical and Interactive Spatial Proteomics Image Analysis Workflow

Spatial proteomics provides a spatially resolved view of protein expression and localization within cells and tissues by mapping the location and abundance of proteins. There is a need for fully-integrated end-to-end imaging workflows for spatial proteomic analysis that are flexible, high-throughput, and support graphical and interactive visualizations. We present a modular and interactive spatial proteomic image analysis workflow with individual steps containerized that empowers biomedical researchers to reproducibly execute and customize complex analyses. Our workflow consists of cell segmentation, unsupervised clustering, validation of clusters on the image, and cell type clustering results visualization. Users can utilize a form-based graphical interface to execute and customize multi-step workflows with a single click or interactively adjust image processing steps within the workflow, apply workflows to various datasets, and modify input parameters as needed. We illustrated the functionality of our workflow using a cancer imaging dataset consisting of a tissue microarray (TMA) stained by high-plex immunohistochemistry. This TMA contained a variety of cancer and tissue cell types to assess the broad applicability of this workflow to different biopsy and tissue types.

bioinformatics↗

bioRxiv2025

A First-in-Class Therapy Targeting Transcription-Replication Conflicts Drives Synthetic Lethality in Homologous Recombination Deficient Cancers

Hidden beneath the layer of tumor malignancy lies unchecked proliferation driven by hyperactive transcription, which fosters the formation of RNA:DNA hybrids. These structural intermediates precipitate collisions between the transcription and replication machineries. Central to mitigating this genomic threat is interleukin enhancer-binding factor 2 (ILF2), which orchestrates the recruitment of RNA:DNA helicases to resolve R-loops and, in doing so, reveals cancer cells dependency on stress-mitigating mechanisms to avert genome catastrophe. Our discovery of Molephantin and its lead derivative, NYH0002 is the outcome of delineating ILF2s function. NYH002 exerts direct binding to the ILF2 complex and disrupts RNA:DNA helicase activity to elicit genome-wide DNA breakages. Tumors with elevated cyclin E and E2F1 expression are sensitive to NYH002, while those deficient in homologous recombination repair exhibit lethality. Collectively, these findings position ILF2 as a key regulator of R-loop homeostasis and reveal a therapeutic vulnerability that NYH002 exploits to suppress tumor malignancy.

cancer biology↗

bioRxiv2025

Evolutionary dynamics of gene and isoform regulation underlying rapid and parallel adaptive radiations

Adaptive radiation is a major driver of biodiversity. In some of the largest radiations, the pace of morphological change far outstrips the protein-coding mutation rate, suggesting that gene regulation may be important for diversification. Yet, the specific gene regulatory mechanisms shaping adaptive radiation remain poorly understood. Analysing of 200 transcriptomes from three independent but phylogenetically nested African cichlid adaptive radiations we show that alternative splicing (AS) evolved faster than gene expression (GE), playing a pivotal role in shaping novel trophic adaptations in the youngest and most species rich radiations in Lakes Victoria and Malawi. This divergence was largely driven by ancestral alternative isoforms, which, though present at low levels in related lineages that did not form radiations, increased in frequency during adaptive radiation. In addition, novel isoforms of craniofacial genes - some evolving within just a few thousand years contributed further to adaptation. The rapid turnover of AS is consistent with periods of relaxed selection followed by directional selection on alternative isoforms and splice sites, a dynamic that may have preserved a rich cache of isoform variation in these radiations and enabled ecological diversification as adaptive zones became available. We argue that the interplay between splicing and different forms of selection facilitates the generation and maintenance of protein-coding diversity, promoting evolutionary innovation into many ecologically different species at extremely short timescales. Significance statementAdaptive radiation--when species rapidly diversify to fill new ecological niches--is a key driver of biodiversity but its underlying molecular mechanisms remain unclear. Using comparative transcriptome sequencing across multiple African cichlid fish adaptive radiations, we found that rapid changes in alternative splicing--the process generating different protein-coding isoforms from the same gene--contributed more to early ecological divergence than shifts in gene expression levels. Most adaptive isoforms were present at low levels in the ancestor of these fishes but some new isoforms evolved remarkably fast, shaping diverse ecologies. We show that alternative splice variation, often thought to be biological "noise," can be a powerful and labile source of rapid evolutionary innovation during early stages of adaptive radiation.

evolutionary biology↗

bioRxiv2025

Cellular Senescence Mediates Doxorubicin Chemotherapy-Induced Aortic Stiffening: Role of Glycation Stress

BackgroundMechanisms underlying Doxorubicin (Doxo) chemotherapy-induced aortic stiffening are incompletely understood. ObjectivesDetermine the role of cellular senescence and the senescence-associated secretory phenotype (SASP) in mediating Doxo-induced aortic stiffening and the influence of senolytic therapy. MethodsAortic stiffness (aortic pulse-wave velocity [PWV]), and associated mechanisms were assessed in young adult p16-3MR mice, a model that allows for genetic-based clearance of senescent cells with ganciclovir [GCV]. Young (4-6 month) mice were injected with Doxo and subsequently treated with GCV or the senolytic ABT263. We evaluated the influence of SASP-associated circulating factors in plasma (the circulating SASP milieu) in mediating aortic stiffening ex vivo (aortic elastic modulus) and examined the contribution of glycation stress. ResultsDoxo increased aortic PWV (425D{+/-}D6 vs. control, 353D{+/-}D5Dcm/sec; P<0.05), an effect prevented by both GCV (348D{+/-}D4Dcm/sec) and ABT263 (342D{+/-}D7Dcm/sec; P<0.05 for both vs. Doxo). Plasma from Doxo-treated mice induced aortic stiffening ex vivo (P<0.05 vs. plasma from control mice), whereas plasma from Doxo-GCV and Doxo-ABT263 groups did not. Glycation stress was implicated in SASP-mediated aortic stiffening with Doxo, as inhibition of receptor mediated glycation stress signaling attenuated plasma-induced aortic stiffening. ConclusionCellular senescence and the circulating SASP milieu contribute to Doxo-induced aortic stiffening. Senolytics hold promise for preserving aortic stiffening following Doxo exposure. Translational perspectiveChemotherapy-induced cardiovascular toxicity is a concern for cancer survivors. This study identifies cellular senescence and the senescence-associated secretory phenotype (SASP) as underlying mechanisms of doxorubicin chemotherapy-induced aortic stiffening - an antecedent to overt cardiovascular disease (CVD). We also provide complementary lines of evidence that glycation stress mediates the mechanistic link between doxorubicin, cellular senescence, the SASP and aortic stiffening. Lastly, we demonstrate the efficacy of senolytic therapy for targeting cellular senescence, the SASP and glycation stress to prevent doxorubicin-induced aortic stiffening. These results offer a novel and clinically actionable approach to preserving vascular health in cancer survivors and mitigating CVD risk.

physiology↗

bioRxiv2025

EZH2 alters aggressive variant prostate cancer subtype evolution, but is not required for neuroendocrine prostate cancer

Advanced prostate cancer (PrCa) remains a leading cause of cancer-related death among men because nearly all patients progress on standard therapy that targets androgen receptor (AR) signaling. An important mechanism driving therapeutic resistance is lineage plasticity, the ability of PrCa cells to reprogram into lineage variants like neuroendocrine (NE) PrCa that no longer depend on AR signaling. The histone methyltransferase EZH2 has been implicated in PrCa lineage plasticity and EZH2 inhibitors are being evaluated clinically for the treatment of PrCa. Here we test how Ezh2 impacts the evolution of PrCa lineage plasticity using genetically engineered mice and human patient data. Reduced Ezh2 expression diversifies the evolution of PrCa lineage variants, including a newly discovered variant with high expression of neurofilament genes, in both mice and humans. Lineage variant diversification is associated with activation of Klf transcription factor programs and AR cistrome reprogramming that is normally repressed by Ezh2. These findings advance the understanding of PrCa lineage plasticity and have implications for using EZH2 as a target for PrCa therapy.

cancer biology↗

bioRxiv2025

Ultrasmall chemogenetic tags with group-transfer ligands

Chemogenetic tags are valuable tools for studying functions of a given protein-of-interest (POI) lacking small-molecule ligands, but most tags are too large for several POIs. Here, we report two ultrasmall chemogenetic tags (mgTag and cTag) of 36 and 50 amino acids (aa) that, to the best of our knowledge, are the smallest reported. These tags exhibit transferase-type reactivity with their ligands to append any moiety of interest to the tag. cTag utilizes an engineered C1 domain-bearing cysteine that undergoes group-transfer reaction with its ligand. Likewise, mgTag utilizes an engineered zinc-finger domain-bearing cysteine that undergoes group-transfer reaction with its molecular-glue ligand in the presence of cereblon (CRBN). We applied these tags in the context of cell signaling and proximity induction. While the fusion of HaloTag (297 aa) to the KRASG12D (188 aa) disrupted its ability to activate the growth-signaling pathway, fusion of mgTag or cTag did not. Group-transfer of BRD4 binder to tags appended to Abelson kinase (ABL) induced proximity between ABL and BRD4, resulting in the latters phosphorylation. Deletion of the transferase-type reactivity reduced phosphorylation levels, suggesting that proximity-inducing chimeras with group-transfer design may be more efficacious in certain scenarios. We envision these ultrasmall tags to have wide-ranging applications, including in basic science, biotechnology, and medicine. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=168 SRC="FIGDIR/small/653252v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1f99888org.highwire.dtl.DTLVardef@18d3e0corg.highwire.dtl.DTLVardef@1e4a044org.highwire.dtl.DTLVardef@d610ee_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2025

HIF1α and HIF2α mRNA Expression in Oral Squamous Cell Carcinoma does not correlate with protein expression

BackgroundOral cancer is one of the common cancers among men, its presentation is usually late. Tumor growth leads to hypoxia, and HIF1 and HIF2 are important regulators of hypoxia induced pathways, inducing angiogenesis and tumor growth. This study compares the expression levels of HIF1 and HIF2 mRNA in OSCC tissues to adjacent normal mucosa. MethodsRNA was isolated from 85 OSCC tissues and adjacent normal mucosa followed by real-time (RT) PCR to quantify HIF1 and HIF2 expression using GAPDH and {beta}-actin respectively as reference genes. ResultsHIF1 overexpression was detected in 76.47% of case and 71.76% of controls (p=0.4906), whereas HIF2 overexpression occurred in 91.76% of cases 94.11% of controls (p=0.35). HIF1 and HIF2 expression level were not significantly different in OSCC tissues compared to adjacent normal mucosa. ConclusionHIF1 and HIF2 play important role in hypoxia induced pathways and tumor progression, yet this study found no significant variation in expression between OSCC tissues and surrounding normal tissues. This suggest that their expression may be due to physiological adaptability to hypoxia or stress, or may be due to use of adjacent tissue as control and loss of protein expression could be due to post translational modifications.

cancer biology↗

bioRxiv2025

Development and validation of HIV SMRTcap for the characterization of HIV-1 reservoirs across tissues and subtypes

Human Immunodeficiency Virus type 1 (HIV-1) is responsible for the global HIV/AIDS epidemic and the establishment of an integrated HIV-1 reservoir remains the primary obstacle to cure. Upon therapy interruption, reactivation of the persistent HIV-1 reservoir propagates viral rebound and mediates continued immunological decline. While furthering understanding of the HIV-1 reservoir is essential for HIV-1 cure, commonly used sequencing strategies are often limited by the reliance on short-read sequencing across separate assays to determine integration sites and proviral integrity - something that does not always adequately resolve complex human genomic repeats or low complexity regions. Simultaneous identification of proviral integration sites and proviral integrity at the single molecule level would enable HIV-1 reservoir characterization with minimal imputation or bioinformatic reconstruction. Here we present HIV Single Molecule Real Time Capture (HIV SMRTcap), a novel molecular and computational pipeline that directly and simultaneously identifies HIV-1 integration sites, defines proviral integrity, and characterizes clonal expansion of HIV-1 provirus-containing cells with single molecule resolution. In combination with long-read, single-molecule, real-time (SMRT) sequencing and custom analytic pipelines, HIV SMRTcap enables a highly comprehensive characterization of HIV-1 reservoirs. Moreover, we demonstrate here that HIV SMRTcap performs robustly across the major global subtypes (HIV-1 subtype A, B, C, D and A/D recombinant viruses), and can use both cell- and tissue-derived inputs, including samples from antiretroviral therapy (ART) treated individuals with undetectable viral loads. Our results demonstrate that HIV SMRTcap serves as a comprehensive, robust method for unbiased HIV-1 reservoir characterization. Used alone, or in combination with single-cell based methods, HIV SMRTcap will enable novel exploration of viral reservoirs across subtypes and in tissue-specific compartments, providing critical information needed to inform HIV-1 cure.

genomics↗