Biology subjects

Shrestha, S.

Publications and source records attributed to Shrestha, S..

At least 19 recordsLinked to original sources

bioRxiv2026

The Influenza Hemagglutinin Cytoplasmic Tail Domain Interacts with Phosphatidylinositol 4,5-bisphosphate

During the influenza viral life cycle, the viral glycoprotein hemagglutinin (HA) mediates binding, entry, and fusion. Densely packed clusters of HA trimers at the plasma membrane are required to produce infectious virions; however, the mechanism of HA clustering is still unknown. We have shown previously that HA co-clusters with and modulates phosphatidylinositol 4,5-bisphosphate (PIP2) in host cell plasma membranes (PM). Here, we further characterize the relationship between HA and PIP2 using molecular dynamics simulations (MD) and fluorescence photoactivation localization microscopy (FPALM) to elucidate a mechanism of HA-PIP2 interaction. We found that the interaction occurs largely between the PIP2 head group and the cytoplasmic tail domain (CTD) of HA. Mutations of the CTD were made to alter charge (HARE, HARREQ), palmitoylation sites (HAMAY), or a combination thereof (HAREMAY, RREQMAY). MD showed that HARREQ and RREQMAY had the strongest effect on HA-PIP2 interactions through a depletion in the radial distribution function of PIP2 around HA at distances [≤]2.5 nm. FPALM revealed that HA cluster density at the PM was significantly reduced by CTD mutations, with the largest reduction occurring in mutants where the CTD charge and acylation were both altered (HAREMAY). HAREMAY clusters were also found to have larger circularities and perimeters, implying a structural change to the clusters. Mutations in the HA transmembrane domain also caused modest changes to the cluster properties of HA and its co-clustering with PIP2. FPALM showed PIP2 co-clustering with HA was also affected by HA mutations with more free PIP2 localized under HAREMAY clusters. A chemical model of simultaneous HA-PIP2 and PH-PIP2 binding enables interpretation of HA-PIP2 interactions and reveals quantitative differences between PIP2 binding by HA CTD mutants. We conclude that the mechanism of HA-PIP2 interaction consists of at least electrostatic and hydrophobic components. Our insights into the mechanism of HA-PIP2 interaction, and the prevalence of putative PIP2-interacting domains in a number of viral spike proteins suggest it may be fruitful to identify methods of disrupting interactions between phosphoinositides and viral proteins.

biophysics↗

bioRxiv2026

Overexpression of miR-424(322)/-503 induces severe dilated cardiomyopathy by regulating the fatty acid oxidation gene expression program

Fatty acid oxidation (FAO) is a major energy source in the adult heart, and disruption of cardiac metabolism is closely associated with heart failure. Here, we investigated the effects of cardiac-specific overexpression of the H19X-encoded miR-424(322)/-503 cluster using an inducible transgenic mouse model. Sustained miR-424(322)/-503 overexpression caused rapid metabolic and functional deterioration, with early impairment of fatty acid oxidation. Short-term induction reduced FAO activity and downregulated genes involved in lipid metabolism, while glycolytic enzyme activity remained largely unchanged. Continued miR-424(322)/-503 expression subsequently led to severe dilated cardiomyopathy characterized by ventricular dilation, wall thinning, fibrosis, reduced contractility, and high mortality. Importantly, disease progression was dependent on the level and duration of miR-424(322)/-503 expression, as intermittent or lower-dose induction delayed cardiac dysfunction and prolonged survival. Withdrawal of miR-424(322)/-503 expression after the onset of dysfunction promoted substantial functional recovery. Together, these findings identify miR-424(322)/-503 as a potent regulator of cardiac metabolic reprogramming that disrupts fatty acid metabolism and drives progressive heart failure.

molecular biology↗

bioRxiv2026

IL-17A Restrains Antiviral Immunity to Promote Chikungunya Virus Infection and Pathogenesis in the Heart

Chikungunya virus (CHIKV) infection is increasingly linked to cardiovascular complications, but the mechanisms underlying CHIKV-induced cardiovascular disease (CVD) remain unclear, and targeted therapies are lacking. Although elevated interleukin-17A (IL-17A) levels have been reported in CHIKV patients and associated with cardiovascular pathology, its role in CHIKV-induced cardiac disease is poorly defined. To address this question, we employed our newly developed heterozygous interferon /{beta}/{gamma} receptor-deficient (Ifnag+/-) mice and primary human cardiac fibroblasts to investigate the contribution of IL-17A signaling to CHIKV-associated cardiac pathology. We found that CHIKV infection induced IL-17A production in the heart, and that mice deficient in Il17a (Il17a-/-) and in its receptor gene, Il-17ra (Il17ra-/-), exhibited marked resistance to CHIKV infection in both cardiac tissue and primary cardiac fibroblasts. Genetic deletion of IL-17A signaling significantly enhanced type I interferon responses and decreased viral burden in mouse hearts. Interestingly, blockade of IL-17RA with an FDA-approved monoclonal antibody for plaque psoriasis, Brodalumab, drastically increased type I interferon production and reduced viral replication in both human cardiac fibroblasts and human embryonic kidney 293 (HEK 293) cells. In addition, inhibition of IL-17A signaling suppressed the expression of pro-inflammatory mediators, including Il-1{beta}, Tnf-, and Cxcl2, reduced immune cell infiltration into cardiac tissue, and mitigated cardiac injury. Importantly, therapeutic blockade of IL-17A signaling after CHIKV infection reduced viral replication in both the heart and circulation. Collectively, these findings identify IL-17A signaling as a critical regulator of CHIKV replication and cardiac inflammation and highlight the IL-17A/IL-17RA axis as a promising therapeutic target for CHIKV-associated cardiovascular disease. ImportanceChikungunya virus (CHIKV) infection has been frequently associated with cardiovascular complications, yet the host pathways that promote viral infection and cardiac injury remain poorly understood. Here, we identify IL-17A signaling as a previously unrecognized regulator of CHIKV pathogenesis in the heart. Using a novel heterozygous interferon receptor-deficient mouse model and primary human cardiac fibroblasts, we demonstrate that IL-17A signaling facilitates CHIKV replication via suppressing antiviral type I interferon responses. Genetic deletion or pharmacological blockade with an FDA-approved monoclonal antibody of IL-17A signaling reduced viral burden, attenuated inflammatory cytokine production, limited immune cell infiltration, and protected against cardiac injury. Importantly, therapeutic inhibition of IL-17A signaling after infection remained effective in reducing viral replication in both cardiac tissue and circulation and mitigating cardiac damage. These findings reveal a critical role for the IL-17A/IL-17RA axis in linking antiviral immunity to CHIKV-induced cardiovascular disease and identify a potential translatable therapeutic target for CHIKV-caused cardiac complications.

immunology↗

bioRxiv2026

Minimal Data, Maximal Insight (MDMI): A Structure-guided Pipeline for Discovering Functional Alternatives in Peptide-Protein Interfaces

Discovering functional peptides across vast sequence space remains a formidable challenge, particularly when experimental training data is scarce. We present Minimal Data Maximal Insight (MDMI), a two-stage structure-guided computational pipeline that designs functional peptide variants using only a small, annotated dataset. Rather than relying on sequence information alone, MDMI integrates three-dimensional structural features derived from predicted peptide-protein complexes into a machine learning model that captures interface geometry and binding energetics. This structure-aware predictor, paired with a genetic algorithm for sequence exploration, reduced false positives from 70% to close to zero in an all-negative benchmark panel compared with a sequence-only model in computational benchmarking, and produced approximately four-fold more high-confidence in silico binders than state-of-the-art peptide/protein design baselines. Using the split-GFP system as a testbed, where fluorescence provides a direct functional readout of peptide-protein complementation, MDMI identified peptides with up to 38% sequence divergence from wild-type in Stage 1 while retaining measurable activity. In Stage 2, motif-guided recombination of successful Stage 1 variants produced highly divergent yet functional peptides bearing over 50% sequence difference from wild-type, revealing two distinct functional clusters in sequence space. As further validation, a top-performing candidate expressed as a full-length GFP fusion retained a GFP-like emission profile, supporting formation of a fluorescent GFP-like scaffold. These results demonstrate that structure-informed pipelines can uncover remote functional sequence space from minimal data, with broad implications for peptide and therapeutic analog discovery.

synthetic biology↗

bioRxiv2026

PanKbase Integrated Single-Cell Map: A Comprehensive Atlas of Human Pancreatic Islets

Single-cell RNA sequencing (scRNA-seq) of human pancreatic islet tissue is a powerful tool for investigating type 1 diabetes (T1D). However, individual datasets are limited in size and fragmented across donors, laboratories, and experimental conditions. To address this, we constructed a comprehensive, integrated scRNA-seq atlas of isolated human pancreatic islets by collating publicly available data generated from tissue provided by resources including the Human Pancreas Analysis Program, the Integrated Islet Distribution Program, and Prodo Labs. Systematic quality controls were implemented to select high-quality samples, reads, and cells. During integration, we accounted for important variables such as age, sex, body mass index, origin study, treatments, islet distribution resources, and sequencing chemistry. Our single-cell atlas comprises 191 high-quality samples from 140 donors (59 female, 81 male) across five phenotypic groups: no diabetes (controls, n=69), autoantibody positivity without diabetes (n=12), pre-diabetes (n=11), T1D (n=12), and type 2 diabetes (T2D) (n=36). In total, the atlas contains 448,935 cells, capturing 13 distinct populations, including alpha cells (43.3%) and beta cells (26.8%), as well as groups such as immune cells (0.6%). Publicly available at www.pankbase.org, this atlas provides a platform for hypothesis-driven investigation of diabetes pathophysiology and, given rigorous quality control, is well-suited for downstream machine-learning applications. Article HighlightsO_LICurrent scRNA-seq datasets of pancreatic islet tissue are limited in size and scattered across donors, laboratories, and experimental conditions, underscoring the need for a consolidated resource. C_LIO_LIWe harmonized datasets from multiple sources to build a comprehensive single-cell map of isolated human pancreatic islets. C_LIO_LIOur atlas captures 448,935 cells from 191 high-quality samples across 140 donors and multiple phenotypic groups, identifying 13 distinct cell populations. C_LIO_LIAvailable at www.pankbase.org, the atlas provides a scalable, rigorously curated platform to support hypothesis-driven diabetes research and can enable a broad range of downstream computational applications. C_LI

genomics↗

bioRxiv2026

Adipose tissue as a site of immune activation and dysfunction in individuals with obesity and asthma

Obesity increases local inflammatory responses in adipose tissue. Individuals with obesity have increased asthma incidence and severity and reduced responses to asthma therapeutics through unknown mechanisms. To identify mechanisms by which increased fat mass augments asthma pathogenesis, single cell RNA sequencing of the immune-rich stromovascular fraction of subcutaneous adipose tissue was conducted from well-characterized adults with obesity-associated asthma matched to adults without asthma. Individuals with asthma had increased abundance of perivascular macrophages and lymphoid-associated macrophages (LAMs) and reduced abundance of classical monocytes and CD4+ and CD8+ naive T cells. Pseudo-bulk differential expression (DE) identified upregulation of cellular metabolism, specifically oxidative phosphorylation, and decreased immune homeostatic pathways in asthma across immune cell subsets. Cell type specific DE analysis of effector cell subtypes identified significant induction of metallothionein gene expression in asthma, a signature of immune cell dysfunction characterized by both an activation and exhaustion phenotype. Gene co-expression analysis identified gene modules associated with asthma diagnosis, lung function, and biomarkers of type 2 inflammation were enriched in effector cells. These data identify adipose tissue dysfunction occurs in obesity-associated asthma and support adipose tissue as therapeutic target to address the enhanced asthma risk among those with obesity. Grant SupportNIH U01AI155299, P30DK020593, R01AI182159, K23HL159351, UL1RR024975-03, P30CA68485, P30EY08126, G20RR030956, 5UL1TR002243, KL2TR002245, P30AI110527, DK020593, American Heart Association 17SFRN33520017.

immunology↗

bioRxiv2026

Mitotic kinase regulation of DNA replication forks

While DNA replication forks initiate in S phase, they do not necessarily complete and terminate prior to cell entry into mitosis. How mitotic proteins regulate leftover replication forks is not well-understood. Using reconstituted DNA replication forks with purified proteins, we show that the budding yeast mitotic kinases Clb2-CDK (M-CDK) and Cdc5 (Plk1 homolog) phosphorylate and regulate several replication elongation proteins. Mrc1 phosphorylation by both kinases results in slower replication, and Pol phosphorylation by M-CDK results in less lagging strand initiation. We further show that a phospho-resistant mutant of Pol bypasses M-CDK inhibition of Pol activity in reconstituted replication reactions. Yeast cells expressing the phospho-resistant mutant exhibit faster cell cycle progression revealing a potential negative feedback mechanism between DNA replication forks and mitotic progression.

molecular biology↗

bioRxiv2026

Orai1 is required for Ca2+-dependent plasma membrane repair and mechanoadaptation

Ca2+-dependent repair of plasma membrane breaches is essential for animal cell viability. An initial passive influx of extracellular Ca2+ triggers the formation of a protein plug that rapidly seals breaches. However, the mechanism of extracellular Ca2+ requirement for subsequent repair remains undefined. EHD2 protein stabilizes the plasma membrane caveolae, which sustain membrane repair, and maintains high surface levels of the caveolae-resident Ca2+ channel Orai1. We establish the requirement of both Orai1 and EHD2 for repair of plasma membrane lesions induced by mechanical injury or by a model bacterial pore-forming toxin. We demonstrate rapid EHD2 recruitment and Orai1-mediated Ca2+ entry at plasma membrane sites of localized mechanical stimulus, the latter requiring EHD2 and CAV1. EHD2 and Orai1 are necessary for mechanosensitive YAP/TAZ-TEAD activation and positive feedback for CAV1 expression that promotes membrane repair. Our studies establish EHD2 and Orai1 as novel components of mammalian plasma membrane repair and mechanoadaptation.

cell biology↗

bioRxiv2026

Dengue serotype-1 virus like particles induce antibody responses following HeLa cell expression

Dengue disease remains a significant global health threat, with current vaccines exhibiting variable efficacy and safety concerns. Virus-like particles (VLPs) offer a promising alternative by mimicking native virus structures without infectious genomes. We engineered a mammalian expression plasmid encoding Dengue-1 prM and E proteins, optimized for secretion using Japanese Encephalitis virus signal sequences, and transiently expressed it in HeLa cells. Purified VLPs exhibited spherical morphology ([~]39 nm diameter) consistent with native virions, as confirmed by transmission electron microscopy. Immunization of mice with these VLPs elicited robust Dengue-1 specific IgG antibody responses. Our study demonstrates production of immunogenic Dengue-1 VLPs in HeLa cells, highlighting their potential as a vaccine candidate and a tool for serodiagnosis. Further characterization of VLP epitopes and protective efficacy is warranted to advance vaccine development. ImportanceDengue remains a significant global health challenge, with serotype 1 being one of the dominant strains causing recurrent outbreaks in Nepal. Existing vaccines demonstrate limited efficacy and pose significant safety concerns, particularly in seronegative populations. To address these limitations, this study explores virus-like particles (VLPs) as a safer alternative vaccine platform. VLPs elicit robust immunogenicity by mimicking the structure of native virus while completely lacking genetic components. This study combines DENV1 structural proteins with optimized expression systems to enhance immunogenicity. This work is particularly significant as the first dengue vaccine research conducted in Nepal, directly addressing antigenic mismatches between existing commercial vaccines and locally circulating viral strains. Furthermore, the study provides scalable platform for developing region-specific dengue vaccines for other serotypes and flaviviruses.

immunology↗

bioRxiv2026

Global population structure and phase variation of serotype 12F Streptococcus pneumoniae following the introduction of pneumococcal conjugate vaccine

BackgroundAfter the global deployment of pneumococcal conjugate vaccines (PCVs), serotype 12F has become the predominant serotype responsible for invasive pneumococcal disease (IPD) worldwide. As PCVs that include serotype 12F are gradually introduced, we aim to characterise the global population structure and genetic diversity of the 12F capsule locus using whole-genome sequencing. Capsule variants with vaccine evasion potential were further investigated by functional experiments. MethodsA global collection of pneumococcal serotype 12F genomes (n=806) from 37 countries across six continents were included in this study. To characterise the serotype 12F population, Global Pneumococcal Sequence Cluster (GPSC), in silico serotype, and antimicrobial resistance profile were inferred from whole-genome data for each isolate. The capsule biosynthesis (cps) locus was analysed for gene content variations that could alter polysaccharide capsule production or structure, thereby influencing recognition by vaccine-induced antibodies. These isolates were further investigated by assessing their capsule production using immunofluorescence assays and its susceptibility to vaccine-elicited antibody killing by opsonophagocytosis assays. FindingsThe global increase in serotype 12F was driven by both distinct pneumococcal lineages across different continents, and a globally-disseminated and multidrug-resistant lineage GPSC26. We identified six capsule variants in nine isolates that had disruptive mutations in cps genes including wze, wcil, wciJ and fnlA. Most (6/9) of the disruptive mutations were a result of strand-slippage mutations. A convergent strand-slippage mutation disrupting the glycosyltransferase gene wciJ was identified in four isolates from distinct lineages and countries. Despite the truncation, three of four isolates with available Quellung typing results still identified them as 12F, indicating the production of the capsule. We then created a genetically engineered lab strain with wciJ knockout and complemented with wciJ containing the strand-slipppage mutation. The knockout strain did not produce any capsule. In contrast, the lab strain with wciJ containing the strand-slippage mutation produced a mixed population of encapsulated and non-encapsulated pneumococci, even within the same chain of pneumococcal cells. This observation indicated encapsulated subpopulation possesses a functional WciJ and rapidly reversible strand-slippage mutation during replication. Opsonophagocytosis assays indicated that the clinical 12F strain with strand-slippage mutation in wciJ exhibited reduced susceptibility to vaccine-elicited serum killing, compared to a genetically closely related 12F clinical strain with an intact wciJ. However, substantial inter-individual antisera variation limits definitive interpretation. InterpretationOur work revealed the global rise of serotype 12F pneumococci has been driven by both regional-specific lineages, and a globally-disseminated and multidrug-resistant lineage GPSC26. We demonstrated that strand-slippage mutation is one of the major drivers of serotype 12F capsule variants and represents a novel mechanism enabling reversible on-off switching of capsule production. The ability to switch off capsule expression in a subpopulation may enable evasion of antibody-mediated killing but increase susceptibility to innate immune clearance. FundingBill & Melinda Gates Foundation, Wellcome Sanger Institute, and the US Centers for Disease Control and Prevention.

genomics↗

bioRxiv2026

Microvascular architecture and dynamics of the choroid plexus brain barrier

The choroid plexus is a specialized blood-cerebrospinal fluid barrier that supports cerebrospinal fluid production, immune surveillance, and molecular exchange between the circulation and the central nervous system, yet its vascular bed remains poorly understood. Here, we combine whole-tissue clearing and light-sheet imaging, single-nucleus transcriptomic reanalysis, and live calcium imaging of intact murine explants to define the structural and functional organization of choroid plexus endothelial networks across development. Three-dimensional imaging and reconstruction highlights a dense, epithelial-ensheathed vascular plexus that is continuous with the broader cerebrovasculature and organized into anatomically distinct inflow and ventricular margin regions. Transcriptomic analysis identified developmentally stratified endothelial subtypes, with embryonic populations enriched for proliferative, motor, and membrane remodeling programs and adult and aged populations enriched for adhesion, extracellular matrix, transport, and mechanosensory pathways. Endothelial subsets across stages expressed genes linked to flow sensing and calcium-dependent mechanotransduction, including Piezo1, Piezo2, and Trpv4. Consistent with these signatures, intact explants exhibited spontaneous, spatially graded calcium oscillations, and pharmacologic activation by Piezo1, triggered robust network-wide calcium responses in embryonic and adult tissue with distinct temporal dynamics. Piezo1 activation also promoted stabilization of PECAM1-associated endothelial adhesion under ex vivo flow conditions. Together, these findings establish the choroid plexus endothelium as a structurally specialized, developmentally dynamic, and mechanosensitive vascular network and provide a framework for investigating endothelial contributions to blood-cerebrospinal fluid barrier function in health and disease.

neuroscience↗

bioRxiv2026

Defining the order of assembly of the Clostridioides difficile divisome complex

Cell division is the ancient pathway by which bacteria synthesize a septum of peptidoglycan, dividing the cell into two. Where all walled bacteria were previously thought to use FtsW-FtsI orthologs to synthesize septal peptidoglycan during division, we recently discovered that the major pathogen Clostridioides difficile is missing FtsW-FtsI and instead relies on the activity of the bifunctional Class A PBP called PBP1 to synthesize the septal peptidoglycan. Furthermore, C. difficile either does not encode or require the majority of canonical divisome proteins described in model bacteria aside from the divisome protein orthologs FtsZ, SepF, and ZapA. Indeed, unlike model systems, SepF and ZapA are essential in C. difficile, suggesting that they have evolved to have a critical function in cell division without the redundant mechanisms present in model organisms. Thus, C. difficile uses a fundamentally different division mechanism compared to previously studied bacteria. To understand how this unusual complex is assembled in C. difficile, we combine CRISPR interference (CRISPRi)-based knock-downs with fluorescent fusions to determine that the hierarchical order of assembly occurs in three phases: (i) FtsZ/ZapA, (ii) SepF, and (iii) PBP1. We further investigate the order of assembly of several non-essential mid-cell localizing proteins and discover that MldA, MldC, DivIVA, FtsK, and PBP3 depend on FtsZ, SepF, and PBP1 for localization, whereas MldB localizes independently of SepF and PBP1. Our work provides a model for divisome assembly in C. difficile and validates genetic and cytological tools that can be used to mechanistically dissect this pathway in the future. IMPORTANCEBacterial cell division has been extensively studied in model systems, but little is known about how this essential process occurs in the clinically important pathogen Clostridioides difficile. Studies in model systems have shown that cell division is carried out by a large multi-protein complex called the "divisome." While components of the divisome are widely conserved and can be traced back to the last bacterial common ancestor billions of years ago, C. difficile uses a unique mechanism of division that is independent of the majority of these canonical divisome genes. In the current study, we characterize the core, essential divisome comprised of FtsZ, ZapA, SepF, and PBP1, and build a model for the order of assembly of this unusual divisome complex.

microbiology↗

bioRxiv2026

Mutant ASXL1 Drives Transcriptional Activation and Repression in Human Hematopoiesis

Mutations in the epigenetic regulator ASXL1 are common in myeloid malignancies and portend a near-universally poor prognosis. While multiple mechanisms for mutant ASXL1-dependent oncogenesis have been proposed, none have been functionally validated in the context of the human hematopoietic stem cell, where these mutations almost certainly arise. Here, we extensively characterized a CRISPR-engineered human hematopoietic stem and progenitor cell model of ASXL1 mutations. In this context, mutant ASXL1 expression decreases differentiation, increases clonogenicity in serial replating experiments, and improves engraftment in immunodeficient mice. We also show that endogenous truncating mutations in ASXL1 drive protein stabilization and confirm that mutant ASXL1 is resistant to proteasomal degradation. At the transcriptional level, these phenotypes are driven by significant repression of the stress-response genes and by increased expression of bromodomain and extra-terminal family protein targets. Using protein-interaction screens, genomic and functional approaches, we link the positive transcriptional changes in ASXL1-mutant cells to BRD4-dependent RNA polymerase II pause release and identify a mechanism for transcriptional repression via a previously uncharacterized interaction with the transcription factor MECOM. Finally, we demonstrate that ASXL1-mutant AML exhibits increased MECOM activity consistent with our gene-editing models. Collectively, these studies highlight a highly reproducible model of mutant ASXL1 in the appropriate cell context. Further, they are the first to functionally describe the mutant ASXL1 interactome in the context of the human HSC, identifying new dependencies with therapeutic potential.

cancer biology↗

bioRxiv2026

MYST acetyltransferases are a targetable therapeutic vulnerability in SETBP1-mutant leukemia

Mutations in SET binding protein 1 (SETBP1) are associated with an adverse prognosis in myeloid malignancies. These mutations stabilize SETBP1 protein, driving increased expression of a progenitor-associated gene expression program through incompletely described mechanisms. A proteomic screen revealed interactions between SETBP1 and members of the MYST acetyltransferase complexes, including the catalytic subunits--KAT6A and KAT7. Mutant SETBP1 increases the localization of MYST complexes at known SETBP1 target genes, including the Hoxa cluster, where it drives increased histone acetylation and gene expression. Treatment of SETBP1D868N-expressing myeloid progenitors with MYST inhibitors reduced target gene expression. To establish the efficacy of MYST inhibition in vivo, we treated mice harboring a syngeneic SETBP1-mutant leukemia with the clinical-grade MYST inhibitor--PF-9363. This resulted in complete hematologic control and increased survival. MYST inhibition was also highly effective against a SETBP1-mutant PDX model. These studies identify MYST acetyltransferases as promising therapeutic targets in SETBP1-mutant malignancies. Statement of SignificanceSETBP1 mutations are markers of high-risk myeloid malignancies, but we lack any targeted therapies to improve outcomes. In this study, we identify MYST acetyltransferases as key drivers of mutant SETBP1-driven transcription. MYST inhibitors are highly effective against SETBP1-mutant leukemia and represent a promising avenue for clinical translation.

cancer biology↗

bioRxiv2025

A reusable model of pangenome selection informs optimal surveillance strategies over vaccine introductions

BackgroundThe human pathogen Streptococcus pneumoniae is a major cause of disease, including pneumonia and meningitis. The introduction of Pneumococcal Conjugate Vaccines (PCVs) initially reduced the burden of disease through a reduction of colonisation by vaccine-targeted serotypes. However, since PCVs only target a proportion of pneumococcal serotypes, they shift intraspecific competition, eventually allowing non-targeted types to replace vaccine types. Understanding the host and pathogen factors causing replacement is important for future vaccine development. Mechanistic understanding of vaccine replacement dynamics is crucial for forecasting and optimisation of genomic surveillance strategies to evaluate realised vaccine effectiveness. MethodsWe developed a mathematical model of the genomic and demographic factors which explain vaccine replacement, used this model to replicate serotype-frequency changes, and investigated cost-effective genomic surveillance strategies. We extended a forward-time model based on the Wright-Fisher model, developing a user-friendly model framework that describes the post-vaccine dynamics of S. pneumoniae populations. Our model describes vaccine replacement as a function of vaccine impact, immigration of new strains, and negative frequency-dependent selection (NFDS) on the accessory genome content. ResultsWe used our model to study vaccine replacement in newly sequenced genomic surveillance data from Nepal, and existing data from the US, and the UK, with distinct surveillance strategies. We showed that the model with NFDS better replicates replacement dynamics than a null model without NFDS, and that NFDS likely only acts on part of the S. pneumoniae accessory genome. We found consistent estimates for vaccination effectiveness across the different study locations and country-specific genes under NFDS, highlighting the importance of conducting genomic surveillance in each country of interest. By simulating data from the model, we showed that an optimal surveillance strategy prioritises per-sampling sample size over sampling frequency for small sampling budgets. ConclusionsOur model can be used to predict vaccine replacement dynamics after PCV introduction, and can be easily reapplied to analyse new data from vaccine introductions or new regions. Our model is available in the R package STUBENTIGER (Studying Balancing Evolution (NFDS) To Investigate Genome Replacement) on GitHub https://github.com/bacpop/Stubentiger.

genomics↗

bioRxiv2025

Warmer temperature accelerates senescence by modifying the aging-dependent changes in the mosquito transcriptome, altering immunity, metabolism, and DNA repair

BackgroundGlobal environmental temperatures are rising, which is increasing the body temperature of mosquitoes. This increase in body temperature is accelerating senescence, thereby weakening immune responses and reproductive processes, and shortening lifespan. To determine how warmer temperature and aging, individually and interactively, shape the transcriptome of the African malaria mosquito, Anopheles gambiae, we conducted RNA-sequencing and network-analysis in naive and immune-induced mosquitoes that had been reared at 27C, 30C or 32C and were 1, 5, 10 or 15 days into adulthood. Results: We demonstrate that immune induction, warmer temperature, and aging alter the transcriptome. Notably, the transcriptome of 1-day-old mosquitoes is pronouncedly different from older mosquitoes, and importantly, warmer temperature modifies the aging-dependent changes to accelerate senescence. For example, warmer temperature amplifies the aging-dependent decrease in immune gene expression but dampens both the aging-dependent decrease in metabolic gene expression and the aging-dependent increase in DNA repair gene expression. ConclusionsAltogether, warmer temperature accelerates senescence, shaping the transcriptome in ways that alter the mosquitos ability to fight infection and survive in a warming environment.

zoology↗

bioRxiv2025

Sustained Yap/Taz activation promotes aberrant alveolar epithelial cell differentiation and drives persistent fibrotic remodeling

YAP/TAZ signaling is required for initiation of lung alveolar repair, yet previous studies in idiopathic pulmonary fibrosis (IPF) predicted increased YAP/TAZ signaling in alveolar epithelial cells (AECs). We investigated whether persistent YAP/TAZ AEC signaling contributes to failed epithelial repair and persistent fibrotic remodeling. In IPF lungs, we identified increased YAP+/TAZ+ AECs and increased expression of YAP/TAZ transcriptional targets compared to donor control lungs. In human lung organoids, pharmacological YAP/TAZ activation resulted in phenotype shifts of AECs into aberrant transitional states. In mice with Yap/Taz activation (YTactive) resulting from deletion of Hippo-kinases Stk3/4 in alveolar-type 2 (AT2) cells, resulted in persistent fibrotic remodeling at 28- and 56-days post-bleomycin injury. Gene promoter activity associated with transitional cell markers (Krt19, Hopx, and Runx2) was increased in YTactive AT2 cells. Immunofluorescent staining showed a loss of AT2 associated Cebpa and increased Krt19 in YTactive lineage traced AT2 cells 28 days post-injury. Inhibition of Yap/Taz using Verteporfin resulted in improved lung repair in YTactive mouse lungs, including increased Cebpa and decreased Krt19+ transitional cells. These findings demonstrate sustained Yap/Taz activation drives abnormal alveolar repair and persistent fibrotic remodeling. Blocking aberrant persistent Yap/Taz activity promotes adaptive repair and has potential as a therapeutic strategy for PF.

molecular biology↗

bioRxiv2025

Disulfide cross-linked redox-sensitive peptide condensates are efficient cell delivery vehicles of molecular cargo.

Biomolecular condensates (BCs) are phase-separated viscoelastic hubs within demixed solutions enriched in proteins and nucleic acids. Such condensates, also called membraneless organelles, are increasingly observed in cells and serve as transient hubs for spatial organization and compartmentalization of biomolecules. Along with the transiency of formation and dissolution, their ability to sequester molecules has inspired us to develop BCs as potential vehicles to transport and deliver molecular cargo. We recently reported the design of disulfide bond cross-linked phase-separating peptide (PSP) condensates that spontaneously dissolve in reducing conditions (JACS, 2024, 146, 255299). Based on the premise that the highly reducing cytoplasm could dissolve PSP condensates and release partitioned cargo, here, we demonstrate the ability of PSP condensates to deliver molecular cargo to the cytoplasm of HeLa cells efficiently. We show that PSP condensates deliver a variety of cargos that differ in their sizes and chemistries, including small molecules, peptides, GFP protein (31 kDa), DNA (1.7 kbp), and mRNA. The transfection efficiencies of PSP condensates for delivering DNA and mRNA were also significantly greater than those of a commercial transfection agent. With room to tailor the condensate properties based on cargo and cell types, these results showcase the potential of disulfide-cross-linked PSPs as effective and customizable cellular delivery vehicles, filling a critical demand gap for such delivery systems.

biophysics↗