Biology subjects

Biswas, S.

Publications and source records attributed to Biswas, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

Investigating the Consequences of Non-active site Mutations on the Structure, Function and Dynamics of the Molten Globule Enzyme Monomeric Chorismate Mutase

Intrinsically disordered enzymes serve as useful models to understand their catalytic function against the backdrop of an unstructured protein. The characteristic flexibility in conformation seen in IDPs is a rare occurrence among enzymes and one such enzyme is the engineered protein: monomeric Chorismate Mutase (mCM). mCM surprisingly retains similar enzyme activity as its parent dimeric protein Chorismate Mutase from Methanococcus jannaschii (MjCM) despite losing the ordered globular structure. In this work using a previously demonstrated transition state analogue (TSA), we analyze the structural transitions in mCM during catalysis. Additionally, consequences of three non-active site single point mutations were investigated using CD; Trp-Dansyl FRET measurements using fluorescence lifetime; and time-resolved fluorescence anisotropy measurements; to map the local (near Trp) and global structural transitions in mCM during catalysis. Mutant2 (W24K + C69A); and Mutant3 (W24K + C69A + A6C); revealed a 97 and 89% drop-in activity compared to mCM; quite unlike Mutant1 (W24K, 19% drop). Mutant1 as opposed to Mutant3 was most sensitive to binding of TSA as quantified by structural displacement measured using FRET. This was consistent with an overall globular structure compaction induced by TSA binding in Mutant1 as reflected by a dip in rotational correlation time of Cys-conjugated dansyl probe from 10.3 to 8.4 ns. Our results highlight the critical role of Cys69 residue, that is ~19 [A] away from mCM active site, in influencing the hydrophobic collapse upon substrate binding and subsequent catalytic activity.

biochemistry↗

bioRxiv2026

β-motifs and molecular flux promote amyloid nucleation at condensate interfaces

Biomolecular condensates are increasingly implicated as intermediates in the formation of pathological amyloid assemblies, yet the mechanisms by which sequence-encoded structural motifs and non-equilibrium molecular transport cooperate at condensate interfaces remain incompletely understood. Here, we introduce Flux-Driven Molecular Dynamics (FD-MD), a simulation framework that combines sequence-encoded {beta}-prone interactions with sustained molecular influx to examine fibril formation at condensate interfaces. Within this framework, we establish three main results. First, a scaling analysis of orientational entropy suggests that condensate interfaces can enhance nucleation relative to the bulk by as much as two orders of magnitude, by reducing the entropic cost of coalignment of rigid {beta}-prone segments. Second, varying segment rigidity and molecular supply rate organizes a non-equilibrium phase diagram with four interfacial growth morphologies, ranging from uniform wetting to fibrillar protrusions and inter-condensate bridging networks. Third, directional fibril elongation displays an inverse relationship with drift velocity, consistent with a mechanism in which higher transport rates to the interface favor planar saturation over directed tip incorporation. Together, these results support a picture in which condensate interfaces can act as kinetically favorable nucleation environments, sequence-encoded rigidity helps determine whether interfaces remain liquid-like or become fibrillar, and molecular flux emerges as an additional control axis in the model for condensate aging trajectories.

biophysics↗

bioRxiv2026

Osmotically Induced Shape Changes in Membrane Vesicles

We develop a self-consistent free-energy framework in which membrane shape and osmotic pressure are determined simultaneously in a finite reservoir by minimizing bending elasticity and solute entropy. Solute conservation makes osmotic pressure a thermodynamic variable rather than an externally prescribed parameter, producing a nonlinear coupling between membrane mechanics and solvent entropy. This coupling modifies the classical stability condition for spherical vesicles: instability emerges from global free-energy competition rather than the linear Helfrich stability criterion. The resulting critical pressures differ by orders of magnitude from Helfrich predictions and agree with simulations for small and large unilamellar vesicles. The framework is relevant to cellular environments involving biomolecular condensate confinement as well as synthetic vesicles and the development of osmotic-pressure-driven encapsulation platforms.

biophysics↗

bioRxiv2026

Beyond the skin barrier: optical clearing enables non-invasive cortex-wide optical coherence angiography in mice in-vivo

Non-invasive, high-resolution visualization of mouse brain vasculature remains challenging due to significant light scattering and absorption by mammalian tissues, hence many optical imaging protocols require scalp and/or skull excision. Here we present a fully reversible tartrazine-based optical clearing strategy that enables cortex-wide optical coherence tomography angiography (OCTA) through intact scalp and skull. We characterized tartrazine properties in the near infrared (NIR)-II band of the 1.3 {micro}m swept-source OCTA system, confirming minimal absorption across 1.25-1.35 {micro}m wavelength range and an effectively constant refractive index, suggesting negligible OCTA distortions. Spatially selective agent application showed that intracranial vessels emerge selectively within the treated region of interest (ROI), whereas untreated regions retain strong interference by the scalp vascular features. Depth-encoded projections and cross-sectional OCTA demonstrated an increased signal recovery at depth and an extended vessel-detection range after clearing. Vessel-map changes were quantified using intersection-over-union and Dice coefficients, yielding high similarity outside the ROI and reduced similarity within the ROI, consistent with a transition from scalp to brain vasculature. Reproducibility was confirmed in three independent 11-week-old mice and validated against scalp-removed reference OCTA. Screening tartrazine in the 0.3-0.8 Molar concentration range (7-min application) identified 0.6 M as optimal for whole-cortex scanning, balancing clearing efficacy and solution handling. Finally, the protocol generalized across mice aged 5-18 weeks. This approach provides a practical route to non-invasive structural cerebrovascular mapping with OCTA.

bioengineering↗

bioRxiv2026

Co-infection with Leptomonas seymouri enhances macrophage survival and promotes intracellular parasite persistence during Leishmania donovani infection

BackgroundLeishmania donovani (LD) is an obligate intracellular parasite that survives and replicates within macrophages. Leptomonas seymouri (LS), a traditionally monoxenous trypanosomatid, has been repeatedly co-isolated with LD from kala-azar cases in India, often together with Leptomonas seymouri narna-like virus 1 (Lepsey NLV1). Whether LS can survive and replicate within mammalian macrophages, and how co-infection influences parasite and virus dynamics, remain unresolved. Methods and FindingsIntracellular survival, replication, and revival of LS alone and during co-infection with LD were systematically evaluated using murine (RAW 264.7) and human (THP-1) macrophages. Quantitative ITS1 PCR demonstrated significant increases in intracellular parasite DNA over 48-168 hours post-infection (h.p.i), indicating active replication rather than persistence. Reduced extracellular count suggested restricted cell lysis and enhanced macrophage survival in co-infection compared to mono-infections. Giemsa staining confirmed intracellular localization of LS. Amastigotes from infected macrophages revived as motile promastigotes upon transformation, whereas extracellular parasites failed to revive beyond 48 hours, confirming macrophages as the exclusive niche for prolonged viability. Co-infection dampened macrophage IL-12 (p40) production, augmenting macrophage survival. MTT assay confirmed the same. In THP-1, co-infection resulted in a marked increase in intracellular LS count and viral load than LS mono-infection at 96 and 168 h.p.i. relative to 48 h.p.i., suggesting that LD co-infection favors higher intracellular virus titres. ConclusionsOur findings demonstrated that LS could replicate within mammalian macrophages and persist during mono-or LD co-infection. The identification of a stable LD-LS-virus interaction highlights a previously underappreciated "triple-pathogen" biology with potential implications for LD pathogenesis.

microbiology↗

bioRxiv2026

Displayed and Encoded Antigens on Adenovirus Vectors Optimize Humoral and Cellular Immune Responses in Rhesus Macaques

Adenovirus vector-based vaccines were deployed widely during the COVID-19 pandemic. In this study, we explored the potential of displaying an antigen on the surface of the adenovirus capsid as well as encoding an antigen as a transgene in the adenovirus vector to optimize both humoral and cellular immune responses. We show that displaying SARS-CoV-2 Spike receptor biding domain (RBD) on the Ad5 capsid while simultaneously encoding Spike as a transgene induced robust antibody and T cell responses in rhesus macaques. These data demonstrate that adenoviruses can be utilized simultaneously as both nanoparticle scaffolds and viral vectors.

immunology↗

bioRxiv2026

PreS1-decorated recombinant adenovirus encoding HBV antigens generates neutralizing humoral and cellular immunity

Background & AimsAchieving functional cure for chronic hepatitis B (CHB) will likely require a combinatorial approach targeting multiple aspects of the complex hepatitis B virus (HBV) life cycle and host adaptive immune responses. Here, we developed a therapeutic vaccination strategy targeting the PreS1 region of L-HBsAg, required for cellular entry of both hepatitis B and D viruses. An established potent T-cell inducing platform, recombinant adenovirus (Ad), was used as a nanoparticle scaffold for PreS1 attachment, to generate antibodies that neutralize virus entry and to establish T-cell mediated immune control. Approach & ResultsScreening a cohort of 61 patients diagnosed with CHB revealed minimal evidence of natural anti-PreS1 responses. Thus, Ad particles encoding multiple HBV antigens were decorated with PreS1 peptide using DogTag/DogCatcher protein superglue. Mice vaccinated with PreS1-decorated Ad induced robust anti-PreS1 antibody responses that neutralized HBV and HDV infection. In contrast, an undecorated Ad encoding L-HBsAg failed to neutralize HBV, demonstrating that PreS1 decoration was required for potent HBV neutralization. Strong CD8+ and CD4+ T-cell responses were induced against HBV antigens encoded in the Ad genome. ConclusionsPreS1-decorated Ad combines immunological HBV and HDV entry inhibition with potent anti-HBV T-cell induction in a single platform, providing a promising addition to current therapeutic strategies against CHB, with particular utility in HBV/HDV co-infection.

bioengineering↗

bioRxiv2026

Designing epitope-based vaccines against Nipah virus Glyco and Fusion proteins using integrated immunoinformatics and structural modeling techniques

Nipah virus (NiV) is a re-emerging zoonotic virus belonging to the Paramyxoviridae family that results in significant neurological damage and raises fatality rates. NiV is classified as a stage III pathogen with a high likelihood of transmission to humans, causing outbreaks intermittently and without any predictable pattern. In Bangladesh, outbreaks of NiV have happened every year since 2001. Even though the disease is severe, no antiviral medications exist for NiV infections. As a consequence, developing a vaccine is crucial. The present research aims to predict an effective epitope-based vaccine by applying immunoinformatic techniques against the fusion and glycoprotein of the Nipah virus. Fusion and glycoproteins were obtained from the UniProt protein database and screened for the T and B cell epitopes using the IEDB and ABCpred servers. Moreover, the constructed 3D structure of the NiV vaccine was occupied with Toll-like receptor 4 for molecular docking and dynamic (MD) simulation studies. Finally, the vaccine design was validated for expression in the PET28a (+) vector of Escherichia coli, and immunological simulations were also conducted. After determination of the allergenicity, antigenicity, and toxicity, the non-allergenic, nontoxic, antigenic, and immunogenic epitopes were used to construct the vaccine with adjuvants and an appropriate linker, including AAY, GPGPG, and KK. The constructed NiV vaccine was confirmed based on its physicochemical properties and docking scores for further analysis. The MD simulations indicated a stabilized structure and increased duration of epitope visibility, indicating strong immune responses. Furthermore, codon optimization and in silico cloning were used to verify bacterial expression of the developed NiV vaccine in E. coli. The results showed that NiV can increase immune responses against the Nipah virus. Furthermore, in vitro and in vivo research and clinical studies are recommended to establish the findings of this study.

immunology↗

bioRxiv2026

Humanized Kcnv2 E151X Mouse Captures Hallmarks of KCNV2-Associated Retinal Dystrophy

BackgroundKCNV2-associated retinopathy is a rare inherited retinal dystrophy caused by variants in the KCNV2 gene, leading to disrupted photoreceptor function and slowly progressive vision loss. Patients have characteristic electroretinography abnormalities, including reduced cone responses, delayed and reduced rod responses to low light flashes and paradoxally large rod-driven responses to bright flashes of light. To model this condition, we generated the Kcnv2 E151X mouse line and assessed its structural and functional retinal features. MethodsWe have employed CRISPR/Cas 9 gene editing technology to generate a mouse line with an early stop mutation in position E151- orthologous to the commonly encountered E143X mutation in humans - and performed a combination of Iimmunohistochemistry and Western Blot to confirm the absence of the full-length KCNV2-encoded protein, Kv8.2. To assess how closely it models the human disease, we have characterised the KCNV2 mutant mouse line at histological and functional levels employing immunohistochemistry and electroretinography, respectiveley. ResultsKcnv2 mutant mice showed markedly reduced photopic responses and reproduced the supernormal rod phenotype as described in affected individuals. In the morphological context, mutant retinas demonstrated strong and early glial fibrillary acidic protein upregulation together with reduced counts of cone arrestin positive cells as well as photoreceptors in general. Power calculations based on the data obtained herein suggest therapeutic trials are feasible with small sample sizes. ConclusionsThe Kcnv2 mutant mouse line replicates key functional and structural hallmarks of KCNV2- associated retinopathy. This model provides a relevant platform for mechanistic studies and preclinical evaluation of gene based or pharmacological therapies targeting cone and rod photoreceptor dysfunction.

neuroscience↗

bioRxiv2025

Aβ42 facilitates the activation of Discoidin Domain Receptor 2 and Its nuclear enrichment in Alzheimer's Disease Model

Discoidin domain receptor 2 (DDR2), a receptor tyrosine kinase (RTK) family member, plays a pivotal role in collagen-mediated signaling, influencing several key cellular processes. In Alzheimers disease (AD), DDR2 expression and activity are significantly upregulated. While collagens, the endogenous ligands of DDR2, either remain unchanged or are downregulated in AD-like conditions, Amyloid-beta 42 (A{beta}42) has been found to activate DDR2 in the absence of collagen non-canonically. Additionally, DDR2 naturally translocates to the nucleus in the neuroblastoma cell line SH-SY5Y, and the pool of DDR2 increases further in the nucleus in the presence of A{beta}42, suggesting its novel pathological role in AD. The E113K mutant of DDR2, where the collagen-binding site has been compromised, fails to get activated by collagen, whereas A{beta}42 could still activate it. Furthermore, the DDR2 inhibitor WRG-28 effectively inhibits A{beta}42 induced phosphorylation of the protein. We have also modeled in silico the ectodomain of DDR2, extending through the transmembrane helix, and subsequently docked A{beta}42 pentamer to elucidate the mechanistic basis of kinase activation by amyloid A{beta}. Additionally, we analyzed WRG-28 binding by constructing the ternary complex of DDR2, A{beta}42, and WRG-28. Our structural analyses offer key insights into the molecular mechanisms governing DDR2 activation by A{beta}42 and its inhibition by WRG-28 in the AD cell model, providing a foundation for targeted therapeutic strategies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/694181v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@1fd0a08org.highwire.dtl.DTLVardef@19db92borg.highwire.dtl.DTLVardef@1e7b7e7org.highwire.dtl.DTLVardef@1dad3fc_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2025

Proteus mirabilis entails a switch from commensal to pathogen - Genomic insights from a blaTEM-1B-harboring novel isolate from India.

BackgroundProteus mirabilis is an opportunistic pathogen that can shift from gut commensalism to causing severe urinary tract and bloodstream infections. The genomic basis of P. mirabilis transition from commensalism to severe infection is not well understood, especially in Indian clinical isolates. This study aimed to explore the genomic attributes of a prevalent P. mirabilis strain in India. MethodsWhole-genome sequencing (WGS) was performed on a clinical P. mirabilis isolate by using NGS. AMR genes and virulence factors were identified using ResFinder/CARD and VFDB, respectively. Sequence typing was performed with MLST. Protein-protein interactions and noncoding RNAs were analyzed using STRING, CytoHubba, and BAKTA. ResultsWGS confirmed the multidrug-resistant nature of the isolate, revealing a broad repertoire of AMR genes, including blaTEM-1B, conferring resistance to beta-lactam antibiotics. Several resistance determinants were co-localised with insertion sequences on a single contig, indicating a high potential for horizontal gene transfer. The virulome profile highlighted multiple fimbrial genes, hemolysins, and toxin-encoding loci, together supporting strong adhesion, biofilm formation, and host tissue damage. Comparative analysis revealed unique genomic signatures not present in reference isolates, including additional virulence determinants and mobile genetic elements. MLST profile of the isolate is closely related to ST-675 & ST-797, while virulence typing (vST-1157) indicated a novel profile distinct from reference ST-675 strains. Insilico protein-protein interaction of this isolate revealed a highly interconnected virulence network, with urease, fimbrial adhesins, and hemolysins emerging as central functional modules driving pathogenicity. In addition, a noncoding RNA was identified within the genomic context of the ure operon (Contig 10), suggesting a possible regulatory role in urease expression. PPI network analysis highlighted urease, fimbrial proteins, and hemolysins as central hubs contributing to pathogenicity. ConclusionsThis study presents the first genomic characterization of a multidrug-resistant P. mirabilis clinical isolate from India, designated as P. mirabilis strain Indica. The convergence of AMR genes, virulence factors, MGEs and a novel noncoding RNAs linked to urease expression highlights the emergence of a high-risk variant in India. These findings have significant implications for infection control and emphasize the need for genomic surveillance to guide therapeutic strategies and develop new interventions globally.

microbiology↗

bioRxiv2025

Lipid Nanoparticles Enable mRNA Delivery to Diverse Cell Types of the Inner Retina

Lipid nanoparticles (LNPs) have emerged as a promising platform for retinal genetic therapy, offering a non-viral alternative to adeno-associated viruses (AAVs). While LNPs can transfect outer retinal cells, their tropism for inner retinal cell types remains insufficiently characterized. Here, we systematically assessed cellular tropism of conventional LNPs encapsulating chemically modified mRNA encoding mCherry in murine retinal explants and dissociated retinal cells. We compared quasi-subretinal and quasi-intravitreal administrations and evaluated how retinal degeneration and inner limiting membrane (ILM) integrity influence LNP-mediated transfections. We observed that LNPs efficiently transfected Muller glia under all experimental conditions. In addition, LNPs transfected several other retinal cell types, including neurons in dissociated cells and explants, and vascular cells exclusively in explants. Subretinal delivery resulted in significantly higher transfection rates than intravitreal administration, and overall efficiency was higher in degenerate as compared to non-degenerate healthy retinas. In healthy retinas, removal of ILM-associated barriers significantly increased transfection efficiency following intravitreal administration. Together, these findings demonstrate that conventional LNPs can transfect a broader range of retinal cell types than previously recognized and highlight LNPs as a versatile tool for mRNA delivery to the retina, with applications in gene supplementation, gene editing, and regenerative therapies for inner retinal disorders.

neuroscience↗

bioRxiv2025

Engineered bacterial M1GS ribozyme efficiently cleaves the most abundant ribosomal RNA in a human cancer cell line

M1GS, an engineered Ribonuclease P (RNase P), can be customized to specifically target and cleave any RNA. It involves tethering an Escherichia coli M1 RNA to a short guide sequence (GS) complementary to the RNA targeted for cleavage. We have developed a Python script-based bioinformatic tool built based on the requirements of M1GS, coupled to secondary structure predictions, to identify M1GS target sites for any given RNA. Primarily, we demonstrate the utility of the tool for predicting M1GS target sites across multiple RNAs, including human 28S ribosomal RNA (rRNA), and then we demonstrate efficient M1GS-mediated downregulation of 28S rRNA in a human cancer cell line. Efficient cleavage of rRNA, the most abundant RNA molecule, suggests a high turnover number for the M1GS ribozyme. Lastly, we discuss the utility of M1GS ribozyme-mediated rRNA downregulation as a potential anticancer modality in cancers where rRNAs are upregulated.

molecular biology↗

bioRxiv2025

PEG400 regulates Falcipain 2 activity through an unprecedented allosteric mechanism

The malarial parasite Plasmodium falciparum cleaves host hemoglobin by cascade of proteolytic enzymes. The cysteine protease, Falcipain-2 (FP2) plays an essential role in the process and important for parasite survival, making it a potential drug target. However, similarities with host cysteine cathepsins hamper selective inhibition, thus necessitates detailed structural and functional characterizations of FP2. The present study uncovers a novel regulatory role of polyethylene glycol 400 (PEG400) on FP2 activity. PEG400 inhibits FP2 activity on small peptide substrate and azo-casein, while enhancing hemoglobin degradation, rendering a dual effect on FP2 catalysis. A mixed-type of inhibition has been observed for PEG400 against small peptide substrate of FP2, consistent with binding of PEG400 to catalytic cleft, confirmed by fluorescence quenching and docking studies. Unlike typical nonspecific PEG-protein interactions, PEG400 adopts a fit within catalytic region of FP2 and partially overlaps with leupeptin binding sites, albeit with lower affinity. Computational analysis further identifies a novel allosteric binding pocket of PEG400, supported by in-silico mutagenesis and molecular dynamics simulation. This pocket exhibits minimal conservation in human cathepsins, suggesting selective potential. In contrast to this inhibitory role, biochemical assay reveals that PEG400 promotes haemoglobin proteolysis. Spectroscopic analyse suggests PEG400 alter hemoglobin structural dynamics to favour proteolysis. ENM based normal mode analysis reveals upon haemoglobin binding, PEG400 restricts FP2 hinge-bending motion, improves FP2-hemoglobin proximity, and simultaneously PEG400 is dislodged from the active site, thereby promoting proteolysis. The combined experimental and computational findings reveal a novel mechanism of FP2 regulation, opening new therapeutic avenues.

biochemistry↗

bioRxiv2025

Designer Aromatic Cations for Photo-Induced Protein Ligation, Imaging, and Intracellular Labelling at Extended Wavelengths

Photo-induced protein labelling strategies have become essential tools in chemical biology, but most strategies require high energy wavelengths of light as input to drive reactivity. Recently, we reported a biocompatible method for engaging photo-induced electron transfer to drive protein labelling using biaryl pyridinium salts and, here, we report the design of a series of aromatic cation salts that trigger this process using longer wavelengths of light while maintaining a sterically minimal profile. We achieved this through the systematic study of structure-reactivity relationships of various donor-acceptor pyridinium salts possessing extended conjugation, and these studies revealed the need of a constrained trans-stilbene relationship between the probes donor and acceptor substituents in order to achieve protein labelling. Probes with chromene-based donor groups in particular showed either robust protein labelling, significant fluorescence quantum yields, or state-dependent photophysical properties; in turn enabling the same probes to be used for both photo-induced protein labelling and wash-free live-cell imaging. We also demonstrate that these enhanced probes possess robust reactivity in complex biological environments through green light-triggered intracellular labelling in live HeLa cells, resulting in the identification of 659 enriched proteins. This series of experiments not only demonstrates the ability of this latest generation of probes to engage in photo-induced labelling using lower energy light in complex proteomes, but also reveals new capabilities for photophysical state-dependent reactivity and measurements. Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/681063v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@1be9a0eorg.highwire.dtl.DTLVardef@741221org.highwire.dtl.DTLVardef@525596org.highwire.dtl.DTLVardef@170ed11_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2025

Dengue NS1 Antibodies drive Immune Complex Formation, Hyperglycaemia and systemic pathology in a murine NS1 plasmid challenge model

Dengue virus (DV) NS1, a secreted virotoxin and key pathogenic factor, can trigger immune responses with poorly understood long-term effects. This study assessed immunopathology in mice administered with DV NS1 plasmid DNA via intraperitoneal (IP), intramuscular (IM), or intravenous (IV) route for DV serotypes 1-4. IP delivery caused the most pronounced effects, including elevated AST/ALT and GRP78 levels, hyperglycemia, and altered organ weights, with DV4 NS1 showing the strongest hepatic damage. Despite serum NS1 antigen being undetectable, mice developed strong NS1-specific antibodies (Abs) and immune complexes. Liver histology revealed degeneration and immune cell depletion. DV NS1 plasmid DNA was detected in liver tissue, but not RNA. DV could infect and replicate in murine pancreatic beta cells. In liver cells, DV increased GAPDH expression, while NS1-Ab-positive serums reduced it. Findings indicated that NS1-specific Abs, not the antigen, drove immune-metabolic dysfunctions, emphasizing the need to evaluate Ab-mediated effects in dengue pathogenesis. Significance StatementDengue virus (DV) remains a leading tropical pathogen, yet the long-term effects of its secreted virotoxin NS1 are incompletely understood. Using a murine model, we demonstrated that NS1 plasmid DNA administration drove systemic and organ-specific pathology. Serotype-specific differences in immune responses, biochemical alterations, and histopathological changes underscored NS1s complex role in disease severity. Elevated immune complexes and liver enzyme profiles highlighted mechanisms of immune modulation and hepatic injury, key features of dengue, while increased serum glucose and GRP78 levels pointed to early markers of diabetes onset. These findings provide foundational evidence that NS1- and NS1 antibody-mediated pathways link dengue pathogenesis with metabolic dysfunction, offering critical insights into host-pathogen interactions and comorbidity development.

microbiology↗

bioRxiv2025

Hematopoietic stem cell conditioned media induces excessive mitochondrial fission via Drp-1 to target colorectal cancer

Mitochondria, often referred to as the "powerhouses of the cell," are particularly crucial in cancer cells due to their high energy demands. Mitochondrial fusion-fission dynamics play a critical role in regulating signaling pathways and metabolic activities in colorectal cancer (CRC) cells. Increased mitochondrial fission drives metabolic reprogramming, enabling CRCs to proliferate, metastasize, and resist chemotherapy. Paradoxically, excessive fission induces mitochondria-mediated apoptosis. Our previous studies have shown that hematopoietic stem cell-derived conditioned media (CM) modulate the apoptosis pathway and mitochondrial bioenergetics of cancer stem cells by altering the cancer microenvironment. In this study, We found that HSCs-CM facilitates excessive fission in colorectal cancer cells by modulating Drp-1 and concurrently activating the PINK1-Parkin mitophagy pathway and intrinsic apoptosis, leading to loss of viability of these cells. Moreover, proteomics data showed that HSCs-CM dysregulated the electron transport chain complexes, with an exceptionally high degree of dysregulation of complexes III and IV. Metabolomics revealed dysregulation of critical metabolites, RNA sequencing revealed dysregulation of transcripts, and proteomics revealed dysregulation of proteins, involved in mitochondrial bioenergetics and the autophagy pathway in CRCs treated with CM. Taken together, our studies reveal the therapeutic potential of HSC-conditioned media for treating colorectal cancer. HighlightsO_LIHematopoietic stem cell-derived conditioned media (HSC-CM) induces excessive mitochondrial fission in colorectal cancer (CRC) cells by upregulating Drp-1, thereby activating the intrinsic apoptotic pathway. C_LIO_LIExcessive fission and bioenergetic dysfunction caused by HSC-CM result in loss of mitochondrial membrane potential and elevated reactive oxygen species production. C_LIO_LIHSC-CM severely disrupts the electron transport chain in CRC cells, precipitating an energy crisis and engaging the PINK1-mediated mitophagy pathway. C_LI SummaryHematopoietic stem cell-derived conditioned media (HSC-CM) compromises mitochondrial dynamics by inducing excessive fission in colorectal cancer (CRC) cells through upregulation of Drp-1 and its associated protein complex. The resulting hyperfission leads to severe mitochondrial dysfunction, characterised by a loss of mitochondrial membrane potential, decreased ATP production, and disrupted electron transport chain complexes. This bioenergetic crisis and overproduction of reactive oxygen species ultimately trigger intrinsic apoptosis and engage the mitophagy pathway, leading to loss of CRC cell viability.

cancer biology↗

bioRxiv2025

Identification of Prospective PETases Across Prokaryotes Using an in silico Approach

BackgroundPlastics count as one of the most potent threats to the habitats and survival of global flora and fauna. Reports keep accumulating globally about the ever-exploding load of plastic wastes, but the need and economics of multiple industrial and household processes compels the production of more plastic materials. It has always been imperative to look for natural sources of degradation of plastic. The identification of plastic-degrading microbes, therefore remains a major focus of the microbial fraternity. While the discoveries of Ideonella sakaiensis or later, Rhizobacter gummiphilus were more out of providence, the structure determination of the enzyme responsible for PET degradation does provide a fillip to efforts towards identification of more such prokaryotic entities. ResultsIn this work, a comprehensive profiling of prokaryotic sequences has been undertaken to look for the presence of similar plastic-degradation abilities across the kingdom. The identification of twenty-seven such hits across different bacterial species led us to believe in the natural diversity of plastic-degradation enzymes. Moreover, there seems to be conservation of the structural motif that renders such ability as has been observed from the constructed models and analysis of their interfaces. Docking of BHET, one of the key products of PET, against these 27 entities showed considerable interactions with the above and pointed towards the possible roles of these bacteria as natural plastic degradation models. Eight of these proteins have very close similarity in binding interactions and surface properties to the PETase from I. sakaiensis and were shortlisted as prospective candidates. ConclusionsOf these eight, three PETases from Halopseudomonas pertucinogena Halopseudomonas bauzanensis and Ketobacter sp. revealed significant similarity in structure and conformational stability to the PETase from I. sakaiensis as was evident from the analysis of their molecular dynamics parameters. Principal Component Analysis and the free energy landscape during binding to BHET also validated the hypothesis and these three PETases could be immediately explored for possible plastic degradation activity.

bioinformatics↗