Biology subjects

Hu, G.

Publications and source records attributed to Hu, G..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Androgen Signaling in Type 2 Innate Lymphoid Cells Drives Sex Differences in Helicobacter-Induced Gastric Inflammation and Atrophy

Background & AimsGastric cancer is the fifth most common cancer worldwide. Men are disproportionately affected by gastric cancer, which ranks as the fourth most common cancer in men compared to eighth in women worldwide. Chronic inflammation driven by Helicobacter pylori infection remains the leading gastric cancer risk factor. Emerging evidence suggests that sex hormones modulate immune responses, contributing to sex differences in infection outcomes and cancer susceptibility. This study investigates how androgens influence the gastric inflammatory response to Helicobacter infection and contribute to sex disparities in disease progression. MethodsMale and female C57BL/6 mice were colonized with Helicobacter felis to investigate sex differences in gastric inflammation. Androgen levels were manipulated by bilateral castration in males and dihydrotestosterone (DHT) treatment in females. Single-cell RNA sequencing was used to identify androgen-responsive leukocyte populations and to establish cell communication networks between leukocyte clusters. The functional roles of these cells were further defined using ILC2- and T cell-deficient mouse models. ResultsInfected female mice developed significantly more severe gastric inflammation, atrophy, and metaplasia infection compared to males. Androgen depletion by castration increased gastric inflammation and accelerated preneoplastic lesion development, while these pathological features were reduced by DHT treatment. Androgen-responsive type 2 innate lymphoid cells (ILC2s) were key initiators of gastric inflammation and ILC2 depletion abolished the sex differences in H. felis pathogenesis. ConclusionsThis study reveals that androgens suppress Helicobacter-induced gastric inflammation by modulating ILC2 activation. We found that androgens are protective, as androgen depletion exacerbated gastric inflammation and accelerated preneoplastic lesion development. These findings provide mechanistic insight into the age-related increase in male gastric cancer incidence, coinciding with declining androgen levels. Our results suggest that circulating androgen concentrations may serve as a prognostic biomarker for gastric cancer risk in men. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=120 HEIGHT=200 SRC="FIGDIR/small/643321v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@6cf3c3org.highwire.dtl.DTLVardef@f797d6org.highwire.dtl.DTLVardef@1118c21org.highwire.dtl.DTLVardef@106e2ab_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Atomistic Profiling of KRAS Interactions with Monobodies and Affimer Proteins Through Ensemble-Based Mutational Scanning Unveils Conserved Residue Networks Linking Cryptic Pockets and Regulating Mechanisms of Binding, Specificity and Allostery

KRAS, a historically "undruggable" oncogenic driver, has eluded targeted therapies due to its lack of accessible binding pockets in its active state. This study investigates the conformational dynamics, binding mechanisms, and allosteric communication networks of KRAS in complexes with monobodies (12D1, 12D5) and affimer proteins (K6, K3, K69) to characterize the binding and allosteric mechanisms and hotspots of KRAS binding. Through molecular dynamics simulations, mutational scanning, binding free energy analysis and network-based analyses, we identified conserved allosteric hotspots that serve as critical nodes for long-range communication in KRAS. Key residues in {beta}-strand 4 (F78, L80, F82), -helix 3 (I93, H95, Y96), {beta}-strand 5 (V114, N116), and -helix 5 (Y157, L159, R164) consistently emerged as hotspots across diverse binding partners, forming contiguous networks linking functional regions of KRAS. Notably, {beta}-strand 4 acts as a central hub for propagating conformational changes, while the cryptic allosteric pocket centered around H95/Y96 positions targeted by clinically approved inhibitors was identified as a universal hotspot for both binding and allostery. The study also reveals the interplay between structural rigidity and functional flexibility, where stabilization of one region induces compensatory flexibility in others, reflecting KRASs adaptability to perturbations. We found that monobodies stabilize the switch II region of KRAS, disrupting coupling between switch I and II regions and leading to enhanced mobility in switch I of KRAS. Similarly, affimer K3 leverages the 3-helix as a hinge point to amplify its effects on KRAS dynamics. Mutational scanning and binding free energy analysis highlighted the energetic drivers of KRAS interactions. revealing key hotspot residues, including H95 and Y96 in the 3 helix, as major contributors to binding affinity and selectivity. Network analysis identified {beta}-strand 4 as a central hub for propagating conformational changes, linking distant functional sites. The predicted allosteric hotspots strongly aligned with experimental data, validating the robustness of the computational approach. Despite distinct binding interfaces, shared hotspots highlight a conserved allosteric infrastructure, reinforcing their universal importance in KRAS signaling. The results of this study can inform rational design of small-molecule inhibitors that mimic the effects of monobodies and affimer proteins, challenging the "undruggable" reputation of KRAS.

biophysics↗

bioRxiv2025

Spatial transcriptomics elucidates the central role of cerebral vasculatures in the progression of Japanese encephalitis

The cellular and molecular mechanisms underlying the neuropathology of Japanese encephalitis virus (JEV) remain obscure. Herein, we designed Stereo-seq chips to simultaneously capture the in situ transcriptomes of both the host and JEV, constructing a comprehensive spatiotemporal pathological landscape for Japanese encephalitis (JE). This study reveals the central role of the vascular system in JE pathogenesis, particularly the meninges, which displayed the strongest signal of inflammation and cell death in the JEV-infected brain. The activation of the Ackr1+ endothelial cells, disruption of the blood-brain barrier, the migration of JEV-infected monocytes, secretion of immune factors by the infected cells, and the occurrence of pyroptosis and necroptosis form a positive feedback loop, resulting in an increasing influx of immune cells and tissue damage. JEV infection leads to neurological impairments, which may be attributed to the interaction between viral proteins and host cellular factors such as Stat1, Stat3, Nfkb1, and Sp1. As the vascular system serves as a central receiver and amplifier of inflammatory signals, regulating inflammation within the vascular system is essential in mitigating JE progression.

immunology↗

bioRxiv2025

Molecular Mechanisms Limiting the Therapeutic Window of AAV Gene Therapy in Mouse Models of Blue Cone Monochromacy

Blue cone monochromacy (BCM) is an X-linked retinal disorder caused by mutations in the OPN1LW/OPN1MW gene locus, resulting in impaired cone function and structural degeneration. We conducted a comparative analysis of AAV-mediated gene therapy in Opn1lw/Opn1mw double knockout (DKO) and Opn1mwC198R/Opn1sw-/- (C198R) BCM mouse models and evaluated the therapeutic window, efficacy, and longevity. Our results demonstrate that the AAV8-Y733F capsid achieved superior cone rescue compared to AAV5. While both DKO and C198R models showed similar therapeutic windows and rescue longevity, treatment efficacy decreased markedly in older mutant mice. Structural analysis revealed that aged cones in both models displayed degenerative changes, including mislocalized mitochondria and compromised connecting cilia. At the molecular level, we observed reduced AAV-mediated transgene expression in DKO and C198R older cones, which may result from decreased transduction efficiency, decreased circular episome stability, genome-wide transcription/translation downregulation, targeted mRNA/protein degradation, or overall cone degeneration. Notably, the cone-specific promoters for Pde6c and Cngb3 maintained robust activity in degenerating cones. These findings suggest that combining an efficient AAV serotype with an optimized cone promoter could be a viable approach to extend the therapeutic window and enhance treatment longevity for BCM patients.

molecular biology↗

bioRxiv2025

PROBind: A Web Server for Prediction, Analysis and Visualization of Protein-Protein and Protein-Nucleic Acid Binding Residues

Protein-protein and protein-nucleic acids interactions are fundamental to numerous cellular functions, yet only a small fraction have been experimentally characterized. Although modern computational methods have been developed for predicting interacting residues in proteins, they are challenging to use due to individual installation and execution requirements, lack of a standardized input or output format, and absence of support for result analysis. Moreover, methods trained using structures of complexes or intrinsically disordered regions, may not perform well on other types. To overcome these challenges, we develop PROBind, a web server for predicting, analyzing, and interactively visualizing protein, DNA and RNA binding residues from both protein sequences and structures. PROBind integrates 12 predictors trained on structural or disordered proteins, and supports the upload of results from external predictors. By normalizing and averaging predictions from multiple predictors targeting the same ligand type, PROBind generates meta-predictions that balance discrepancies among different methods. Furthermore, it provides interactive graphical tools for result analysis and contextualization. Overall, PROBind accommodates diverse ligand types and supports predictions and analysis based on both structure and sequence data, overcoming the limitations of existing tools. PROBind is freely accessible at https://www.csuligroup.com/PROBind.

bioinformatics↗

bioRxiv2025

Enterococcus faecalis-derived lactic acid facilitates persistent and polymicrobial wound infections by suppressing macrophage activation

Macrophage activation is essential for innate immunity, and its suppression enables pathogen persistence. We show that Enterococcus faecalis suppresses macrophage activation through lactic acid-mediated extracellular acidification. Mutants lacking lactate dehydrogenase (ldh), and thus unable to acidify the environment, fail to inhibit NF-{kappa}B. E. faecalis-derived lactic acid acts via MCT-1 and GPR81 through two distinct but complementary mechanisms that culminate in the reduction of NF-{kappa}B activity. Lactic acid acts through MCT-1 to inhibit ERK and STAT3 phosphorylation, leading to reduced STAT3 binding to the Myd88 promoter, and reduced MyD88 protein levels. Lactic acid signaling to GPR81 mediates the phosphorylation of the transcriptional factor YAP, ultimately attenuating NF-{kappa}B signaling. In a murine wound infection model, this lactic acid-driven immunosuppressive niche enables prolonged E. faecalis persistence and enhances the fitness of co-infecting bacteria such as Escherichia coli. These findings reveal how bacterial lactic acid subverts innate immunity to support chronic and polymicrobial infections. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/635924v2_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@2e1565org.highwire.dtl.DTLVardef@17eaf85org.highwire.dtl.DTLVardef@356139org.highwire.dtl.DTLVardef@821581_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- E. faecalis lactate dehydrogenase mutants fail to acidify the environment and cannot suppress NF-{kappa}B signaling. - Lactic acid is necessary and sufficient to drive NF-{kappa}B suppression in macrophages. - Lactic acid NF-{kappa}B suppression is relayed through two complementary routes: MCT- 1 transport and the lactate sensor GPR81. - MCT-1-dependent signaling blunts ERK/STAT3 phosphorylation, lowering MyD88 levels; whereas GPR81 drives YAP phosphorylation, reducing NF-{kappa}B activation. - E. faecalis-derived lactic acid drives immunosuppression, potentiates persistence, and promotes multi-species wound infection in vivo.

microbiology↗

bioRxiv2025

Accurate Characterization of the Allosteric Energy Landscapes, Binding Hotspots and Long-Range Communications for KRAS Complexes with Effector Proteins : Integrative Approach Using Microsecond Molecular Dynamics, Deep Mutational Scanning of Binding Energetics and Allosteric Network Modeling

KRAS is a pivotal oncoprotein that regulates cell proliferation and survival through interactions with downstream effectors such as RAF1. Oncogenic mutations in KRAS, including G12V, G13D, and Q61R, drive constitutive activation and hyperactivation of signaling pathways, contributing to cancer progression. Despite significant advances in understanding KRAS biology, the structural and dynamic mechanisms of KRAS binding and allostery by which oncogenic mutations enhance KRAS-RAF1 binding and signaling remain incompletely understood. In this study, we employ microsecond molecular dynamics simulations, Markov State Modeling, mutational scanning and binding free energy calculations together with dynamic network modeling to elucidate the effect of KRAS mutations and characterize the thermodynamic and allosteric drivers and hotspots of KRAS binding and oncogenic activation. Our simulations revealed that oncogenic mutations stabilize the open active conformation of KRAS by differentially modulating the flexibility of the switch I and switch II regions, thereby enhancing RAF1 binding affinity. The G12V mutation rigidifies both switch I and switch II, locking KRAS in a stable, active state. In contrast, the G13D mutation moderately reduces switch I flexibility while increasing switch II dynamics, restoring a balance between stability and flexibility. The Q61R mutation induces a more complex conformational landscape, characterized by the increased switch II flexibility and expansion of functional macrostates, which promotes prolonged RAF1 binding and signaling. Mutational scanning of KRAS-RAF1 complexes identified key binding affinity hotspots, including Y40, E37, D38, and D33, and together with the MM-GBSA analysis revealed the hotspots leverage synergistic electrostatic and hydrophobic binding interactions in stabilizing the KRAS-RAF1 complexes. Network-based analysis of allosteric communication identifies critical KRAS residues (e.g., L6, E37, D57, R97) that mediate long-range interactions between the KRAS core and the RAF1 binding interface. The central {beta}-sheet of KRAS emerges as a hub for transmitting conformational changes, linking distant functional sites and facilitating allosteric regulation. Strikingly, the predicted allosteric hotspots align with experimentally identified allosteric binding hotspots that define the energy landscape of KRAS allostery. This study highlights the power of integrating computational modeling with experimental data to unravel the complex dynamics of KRAS and its mutants. The identification of binding hotspots and allosteric communication routes offers new opportunities for developing targeted therapies to disrupt KRAS-RAF1 interactions and inhibit oncogenic signaling. Our results underscore the potential of computational approaches to guide the design of allosteric inhibitors and mutant-specific therapies for KRAS-driven cancers.

biophysics↗

bioRxiv2025

Decoding Mechanisms of PTEN Missense Mutations in Cancer and Autism Spectrum Disorder using Interpretable Machine Learning Approaches

Missense mutations in oncogenic proteins that are concurrently associated with neurodevelopmental disorders have garnered significant attention. Phosphatase and tensin homolog (PTEN) serves as a paradigmatic model for mapping its mutational landscape and identifying genotypic predictors of distinct phenotypic outcomes, including cancer and autism spectrum disorder (ASD). Despite extensive research into the genotype-phenotype correlations of PTEN mutations, the mechanisms underlying the dual association of specific PTEN mutations with both cancer and ASD (PTEN-cancer/ASD mutations) remain elusive. This study introduces an integrative approach that combines machine learning (ML) with structural dynamics to elucidate the molecular effects of PTEN-cancer/ASD mutations. Analysis of biophysical and network biology-based signatures reveals a complex energetic and functional landscape. Subsequently, an ML model and corresponding integrated score were developed to classify and predict PTEN-cancer/ASD mutations, underscoring the significance of protein dynamics in predicting cellular phenotypes. Further molecular dynamics simulations demonstrated that PTEN-cancer/ASD mutations induce dynamic alterations characterized by open conformational changes restricted to the P loop and coupled with inter-domain allosteric regulation. This research aims to enhance the genotypic and phenotypic understanding of PTEN-cancer/ASD mutations through an interpretable ML model integrated with structural dynamics analysis. By identifying shared mechanisms between cancer and ASD, the findings pave the way for the development of novel therapeutic strategies.

bioinformatics↗

bioRxiv2025

protPheMut: An Interpretable Machine Learning Tool for Classification of Cancer and Neurodevelopmental Disorders in Human Missense Variants

MotivationRecent advances in human genomics have revealed that missense mutations in a single protein can lead to distinctly different phenotypes. In particular, some mutations in oncoproteins like Ras, MEK, PI3K, PTEN, and SHP2 are linked various cancers and Neurodevelopmental Disorders (NDDs). While numerous tools exist for predicting the pathogenicity of missense mutations, linking these variants to certain phenotypes remains a major challenge, particularly in the context of personalized medicine. ResultsTo fill this gap, we developed protPheMut (Protein Phenotypic Mutations Analyzer), leveraging multiple interpretable machine learning methods and integrate diverse biophysics and network dynamics-based signatures, for the prediction of mutations of the same protein can promote cancer, or NDDs. We illustrate the utility of protPheMut in phenotypes (cancer/NDDs) prediction by the mutation analysis of two protein cases, that are PI3K and PTEN. Compared to seven other predictive tools, protPheMut demonstrated exceptional accuracy in forecasting phenotypic effects, achieving an AUROC of 0.8501 for PI3K mutations related to cancer and Cowden syndrome. For multi-phenotypes prediction of PTEN mutations related to cancer, PHTS, and HCPS, protPheMut achieved an AUC of 0.9349 through micro-averaging. Using SHAP model explanations, we gained insights into the mechanisms driving phenotype formation. A userfriendly website deployment is also provided. AvailabilitySource code and data are available at https://github.com/Spencer-JRWang/protPheMut. We also provide a user-friendly website at http://netprotlab.com/protPheMut. Supplementary informationSupplementary data are available at Bioinformatics online. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=64 SRC="FIGDIR/small/631365v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@df55e3org.highwire.dtl.DTLVardef@7fe3f9org.highwire.dtl.DTLVardef@501cccorg.highwire.dtl.DTLVardef@192c98e_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Butyrolactol A is a phospholipid flippase inhibitor that potentiates the bioactivity of caspofungin against resistant fungi

Fungal infections cause millions of deaths annually and are challenging to treat due to limited therapeutic options and rising resistance. Cryptococci are intrinsically resistant to the latest generation of antifungals, echinocandins, while Candida auris, a notorious global threat, is also increasingly resistant. We performed a natural product extract screen to rescue caspofungin fungicidal activity against Cryptococcus neoformans H99 and identified butyrolactol A, which restores echinocandin efficacy against resistant fungal pathogens, including multidrug-resistant C. auris. Mode of action studies revealed that butyrolactol A inhibits the phospholipid flippase Apt1-Cdc50, blocking phospholipid transport. Cryo-electron microscopy analysis of the Apt1*butyrolactol A complex revealed that the flippase is trapped in a dead-end state. Apt1 inhibition disrupts membrane asymmetry, vesicular trafficking, and cytoskeletal organization, thereby enhancing echinocandin uptake and potency. This study identifies lipid flippases as promising antifungal targets and demonstrates the potential of revisiting natural products to expand the antifungal arsenal and combat resistance.

biochemistry↗

bioRxiv2024

The Septin Cytoskeleton is a Novel Regulator of Intestinal Epithelial Barrier Integrity and Mucosal Inflammation

Background and AimsIntestinal epithelial barrier-integrity is essential for human health, and its disruption induces and exacerbates intestinal inflammatory disorders. While the cytoskeleton is critical for maintaining gut barrier-integrity, the role of the septins- the newest family of cytoskeletal proteins- is unknown. To address this knowledge gap, we evaluate the role of SEPT9- a critical component of the septin-cytoskeleton- in intestinal epithelial cell (IEC) barrier permeability and inflammation. MethodsWe developed SEPT9-NeonGreen knockin mice, inducible intestinal epithelial cell (IEC)-specific SEPT9 knockout (KO) mice, and SEPT9-KO human IEC lines. SEPT9 localization was analyzed using super-resolution microscopy. Barrier-integrity was assessed via transepithelial electrical resistance, FITC-dextran flux, and visualization of tight junction (TJ) and adherens junction (AJ) proteins. Dextran sodium sulfate-induced experimental colitis was evaluated in control and KO mice through measuring cytokine expression, immune cell infiltration, and IEC death. SEPT9 expression was examined in intestinal tissue of IBD patients. ResultsSEPT9 overlapped with TJs and AJs at IEC apical junctions. SEPT9 loss resulted in a leaky epithelial barrier due to mislocalization of junctional proteins. SEPT9 interacted with non-muscle myosin IIC (NMIIC) at the IEC apical-junctional actomyosin belt, and its ablation displaced NMIIC from IEC junctions. Loss of NMIIC also caused barrier disruption. SEPT9 KO mice exhibited increased susceptibility to experimental-colitis. SEPT9 expression was significantly reduced in intestinal mucosa of IBD patients. ConclusionSEPT9 regulates intestinal barrier integrity, supporting TJ and AJ assembly through NMIIC recruitment to the actomyosin belt. SEPT9 safeguards the intestinal mucosa during acute inflammation, and its reduced expression in IBD suggests a loss of this protective function. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=126 SRC="FIGDIR/small/629767v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@5141dcorg.highwire.dtl.DTLVardef@bae524org.highwire.dtl.DTLVardef@19c8eaorg.highwire.dtl.DTLVardef@d5b218_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2024

Oligomerization of PRMT1 Regulated RNA-Binding Protein Cascade Promotes Pancreatic Ductal Adenocarcinoma

PRMT1 is the predominant type I protein arginine methyltransferase responsible for generating monomethylarginine and asymmetric dimethylarginine (MMA and ADMA) in various protein substrates. It regulates numerous biological processes, including RNA metabolism, mRNA splicing, DNA damage repair, and chromatin dynamics. The crystal structure of rat PRMT1 has been previously reported as a homodimer; however, higher-order oligomeric species of human PRMT1 have been observed in vivo, and their structural basis remains elusive. In this study, we present the cryo-EM structure of human PRMT1 in its oligomeric form, revealing novel interfaces crucial for PRMT1 self-assembly and normal function. PRMT1 shows a strong preference for intrinsically disordered RGG/RG motifs, which are commonly found in RNA-binding proteins (RBPs), highlighting its critical role in regulating RNA metabolism. In vitro studies indicate that disrupting PRMT1 oligomerization impairs its binding affinity for RGG motif substrates, thereby reducing arginine methylation levels on these substrates. This finding emphasizes the essential role of the oligomeric state of PRMT1 in its function with RGG motif-containing RBPs. In vivo, the loss of PRMT1 oligomerization leads to decreased global ADMA levels in pancreatic ductal adenocarcinoma (PDAC) cells and inhibits PDAC tumor growth. Collectively, we propose that PRMT1 oligomerization, rather than mere dimerization, is sufficient for PRMT1-driven PDAC tumor growth, offering novel insights into the potential therapeutic targeting of PRMT1 oligomeric forms in PDAC. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/627716v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1ed6232org.highwire.dtl.DTLVardef@1c9e688org.highwire.dtl.DTLVardef@3e2c43org.highwire.dtl.DTLVardef@1cbf33e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2024

Generation of a novel constitutive smooth muscle cell-specific Myh11-driven Cre mouse model

Dysfunction in either embryonic or postnatal vascular smooth muscle cells (SMCs) significantly contributes to the progression of various cardiovascular diseases. Therefore, elucidating the molecular mechanisms governing VSMC development and homeostasis is crucial. MYH11 is the most reliable lineage gene for SMCs and has been utilized to develop tamoxifen-inducible Cre driver lines for achieving SMC-specific gene manipulation by crossing with mice carrying the loxP-flanked gene, particularly in adult mice. For studies involving SMCs during embryogenesis, the commonly used constitutive Cre driver is controlled by the Tagln (Sm22) promoter. However, this Cre driver exhibits activity in multiple non-SMC populations, including cardiomyocytes and skeletal muscle precursors, introducing confounding effects. Additionally, most existing SMC-specific Cre drivers are generated using a transgenic approach, raising concerns about random site integration and variable gene copy numbers. To address these limitations, we report a novel Cre mouse model generated by knock-in (KI) of a nuclear-localized Cre recombinase into the Myh11 gene locus using homologous recombination. We confirmed that the Cre activity precisely recapitulates endogenous Myh11 expression by crossing with Rosa26 mTmG or tdTomato reporter mice. Moreover, Myh11-driven Cre can efficiently delete the floxed allele of the transcription factor Tead1 specifically in SMCs. The Tead1 SMC-specific knockout mice did not exhibit an overt phenotype, thereby circumventing the embryonic lethal phenotype mediated by Tagln-driven Cre, as we previously reported. These findings establish this novel Cre driver line as a robust tool for tracing the Myh11-positive SMC lineage and manipulating gene function specifically in SMCs during embryonic development in mice.

genetics↗

bioRxiv2024

Tear secretion via a paracellular pathway in lacrimal gland is regulated by myosin-mediated modulation of tight junction permeability

"Dry eye", characterized by symptoms of ocular discomfort and visual disturbances due to decreased tear secretion, affects 16 million Americans. Yet, there is currently no cure for dry eye as the mechanistic details of water secretion in the tear-producing lacrimal gland have not been fully elucidated. While a transcellular water secretion pathway via water channels like AQP5 has been reported, the existence and function of a paracellular pathway via tight junctions between epithelial cells remains controversial. The actomyosin cytoskeleton localizes to the apical junctions of epithelial cells across organs and regulates tight junction integrity. Here, we report that non-muscle myosin IIC (NMIIC) is enriched at apical junctions of ductal epithelial cells in the lacrimal gland, leading us to hypothesize that NMIIC regulates tear secretion through modulation of tight junction permeability. Consistent with this hypothesis, we found that tear volume after carbachol stimulation was significantly increased in mice lacking NMIIC, and levels of the tight junction protein ZO-1 were significantly reduced. Furthermore, pharmacological activation of NMIIC by 4-Hydroxyacetophenone in wildtype mice significantly inhibited tear secretion. In summary, our findings reveal a paracellular water secretion pathway in the lacrimal gland, which is regulated by NMIIC-mediated modulation of ductal cell tight junctional permeability, and can be targeted by small molecules. Significance StatementWhile dry eye affects more than 16 million Americans, there is currently no cure as the mechanisms underlying tear secretion are incompletely understood. Here, we report first evidence for the existence and function of a paracellular water pathway, in which water flows between cells, in the lacrimal gland. We also show that this pathway is directly regulated by the modulation of tight junction permeability by non-muscle myosin IIC (NMIIC). This study thus identifies a new mechanism for exocrine secretion, which can be targeted towards developing treatments for dry eye/mouth syndrome.

physiology↗

bioRxiv2024

Multi-omics investigation of spontaneous T2DM macaque reveals gut microbiota promote T2DM by up-regulating the absorption of excess palmitic acid

Although gut microbiota and lipid metabolites have been suggested to be closely associated with type 2 diabetes mellitus (T2DM), the interactions between gut microbiota, lipid metabolites and the host in T2DM development remains unclear. Rhesus macaques may be the best animal model to investigate these relationships given their spontaneous development of T2DM. We identified eight spontaneous T2DM macaques and conducted a comprehensive study investigating the relationships using multi-omics sequencing technology. Our results from 16S rRNA, metagenome, metabolome and transcriptome analyses identified that gut microbiota imbalance, tryptophan metabolism and fatty acid {beta} oxidation disorders, long-chain fatty acid (LCFA) accumulation, and inflammation occurred in T2DM macaques. We verified the accumulation of palmitic acid (PA) and activation of inflammation in T2DM macaques. Importantly, mice transplanted with spontaneous T2DM macaque fecal microbiota and fed a high PA diet developed prediabetes within 120 days. We determined that gut microbiota mediated the absorption of excess PA in the ileum, resulting in the accumulation of PA in the serum consequently leading to T2DM in mice. In particular, we demonstrated that the specific microbiota composition was probably involved in the process. This study provides new insight into interactions between microbiota and metabolites and confirms causative effect of gut microbiota on T2DM development.

microbiology↗

bioRxiv2024

Polarized exocytosis and anionic phospholipid species implicated in the initiation of clathrin-mediated endocytosis

Understanding of the mechanisms that initiate clathrin-mediated endocytosis (CME) is incomplete. Recent studies in budding yeast identified the endocytic adaptor proteins Yap1801/Yap1802 (budding yeast AP180) as key CME factors that promote CME initiation in daughter cells during polarized growth, but how Yap1801/2 are recruited preferentially to the plasma membrane of daughter cells is not clear. The only known cargos for Yap1801/2 in yeast are the synaptobrevins Snc1 and Snc2, which serve as v-SNARES for exocytic vesicles reaching the plasma membrane and are crucial for polarized cell growth. In this study, we examine the spatiotemporal dynamics of functional, fluorescent protein-tagged Snc2 expressed from its endogenous locus and provide evidence that, along with anionic phospholipids, Snc2 specifically recruits Yap1802 to growing daughter cells. This protein-protein interaction creates a direct link between polarized secretion and CME and has further implications in CME initiation.

cell biology↗

bioRxiv2024

SoyOD: An Integrated Soybean Multi-omics Database for Mining Genes and Biological Research

Soybean is a globally important crop for food, feed, oil, and nitrogen fixation. A variety of multi-omics research has been carried out generating datasets ranging from genotype to phenotype. To utilise this data, a soybean multi-omics database that has broad data coverage and comprehensive data analysis tools would be of value for basic and applied research. We present the soybean omics database (SoyOD), which integrates significant new datasets with existing public datasets for the most comprehensive collection of soybean multi-omics information. Compared to the existing soybean database, SoyOD incorporates an extensive collection of novel data derived from the deep-sequencing of 984 germplasms, 162 novel transcriptome datasets from seeds at different developmental stages, 53 phenotypic datasets, and over 2500 phenotypic images. In addition, SoyOD integrates existing data resources, including 59 assembled genomes, genetic variation data from 3904 soybean accessions, 225 sets of phenotypic data, and 1097 transcriptomic sequences covering 507 different tissues and treatment conditions. SoyOD is a novel tool, as it can be used to mine and analyze candidate genes for important agronomic traits, as shown in a case study on plant height. Additionally, powerful analytical and easy-to-use toolkits enable users to easily access the available multi-omics datasets, and to rapidly search genotypic and phenotypic data in a particular germplasm. The novelty, comprehensiveness, and user-friendly features of SoyOD make it a valuable resource for soybean molecular breeding and biological research. SoyOD is publicly accessible at https://bis.zju.edu.cn/soyod.

plant biology↗

bioRxiv2024

Transmembrane channel-like 4 and 5 proteins at microvillar tips are potential ion channels and lipid scramblases

Microvilli--membrane bound actin protrusions on the surface of epithelial cells--are sites of critical processes including absorption, secretion, and adhesion. Increasing evidence suggests microvilli are mechanosensitive, but underlying molecules and mechanisms remain unknown. Here, we localize transmembrane channel-like proteins 4 and 5 (TMC4 and 5) and calcium and integrin binding protein 3 (CIB3) to microvillar tips in intestinal epithelial cells, near glycocalyx insertion sites. We find that TMC5 colocalizes with CIB3 in cultured cells and that a TMC5 fragment forms a complex with CIB3 in vitro. Homology and AlphaFold2 models reveal a putative ion permeation pathway in TMC4 and 5, and molecular dynamics simulations predict both proteins can conduct ions and perform lipid scrambling. These findings raise the possibility that TMC4 and 5 interact with CIB3 at microvillar tips to form a mechanosensitive complex, akin to TMC1 and 2, and CIB2 and 3, within the mechanotransduction channel complex at the tips of inner ear stereocilia.

cell biology↗