Biology subjects

Hu, J.

Publications and source records attributed to Hu, J..

At least 181 records · Page 10Linked to original sources

bioRxiv2025

Viral Infection Detector: Ensemble Learning for Predicting Viral Infection in Single-cell Transcriptomics of Virus-Induced Cancers

Oncoviruses cause around 15-20% of all human cancers. As 3 end-based single-cell RNA-sequencing (scRNA-seq) methods become more popular, the relationship between viral infection and tumor progression at the transcriptomic level has been increasingly studied at single-cell resolution. However, the identification of infected cells is challenging as viral reads from infected cells may not be captured, leading to high false negative rates. To recover these infected cells that have been missed, we developed VID, a stacked ensemble machine-learning model that predicts virus-infected cells in scRNA-seq datasets. Using VID, we uncovered biologically meaningful differences between infected and uninfected cells across different oncovirus-mediated cancers and different cell types. Our VID tool provides a user-friendly way to study changes induced by viral infections in scRNA-seq datasets.

bioinformatics↗

bioRxiv2025

Evidence for Functional Regulation of the KLHL3/WNK Pathway by O-GlcNAcylation

The 42-member Kelch-like (KLHL) protein family are adaptors for ubiquitin E3 ligase complexes, governing the stability of a wide range of substrates. KLHL proteins are critical for maintaining proteostasis in a variety of tissues and are mutated in human diseases, including cancer, neurodegeneration, and familial hyperkalemic hypertension. However, the regulation of KLHL proteins remains incompletely understood. Previously, we reported that two KLHL family members, KEAP1 and gigaxonin, are regulated by O-linked {beta}-N-acetylglucosamine (O-GlcNAc), an intracellular form of glycosylation. Interestingly, some ubiquitination targets of KEAP1 and gigaxonin are themselves also O-GlcNAcylated, suggesting that multi-level control by this post-translational modification may influence many KLHL pathways. To test this hypothesis, we examined KLHL3, which ubiquitinates with-no-lysine (WNK) kinases to modulate downstream ion channel activity. Our biochemical and glycoproteomic data demonstrate that human KLHL3 and all four WNK kinases (WNK1-4) are O-GlcNAcylated. Moreover, our results suggest that O-GlcNAcylation affects WNK4 function in both osmolarity control and ferroptosis, with potential implications ranging from blood pressure regulation to neuronal health and survival. This work demonstrates the functional regulation of the KLHL3/WNK axis by O-GlcNAcylation and supports a broader model of O-GlcNAc serving as a general regulator of KLHL signaling and proteostasis.

cell biology↗

bioRxiv2025

Clonal interference, genetic variation and the speed of evolution in structured populations

When it comes to understanding the role that population structure plays in shaping rates of evolution, it is commonly accepted that interference between evolutionary innovations is more prevalent in structured populations compared to well-mixed, and that population structure reduces the rate of evolution, while simultaneously promoting maintenance of genetic variation. Prior models usually represent population structure using two or more connected demes or lattices with periodic boundary conditions. Fundamentally, the observed spatial evolutionary slow-down is rooted in the fact that these types of structures increase the time it takes for a selective sweep and therefore, increase the probability that multiple beneficial mutations will coexist and interfere. Here we show that systematically introducing more heterogeneity in population structure can reshape these prior conclusions and lead to a much wider range of observed evolutionary outcome, including increased rates of evolution. At a big picture level, our results showcase that the evolutionary effects of population structure crucially depend on the properties of the topology considered. Understanding these spatial properties is therefore requisite for making meaningful evolutionary comparisons across different topologies, for forming sensible null expectations about experimental or observational data or for developing an intuition for when well-mixed approximations, or approximations done using spatial models with a high degree of symmetry, might not apply.

evolutionary biology↗

bioRxiv2025

Evolution of Eukaryotic Specific DNA Binding Sites in Asgard Archaeal RecA Recombinases

Homologous recombination (HR) relies on the RecA-family recombinases Rad51 and Dmc1, which both catalyze DNA strand pairing and exchange but promote distinct recombination outcomes. How these recombinases became functionally specialized remains unclear. Here, we combine phylogenetic analysis with functional assays to define the basis of this specialization. We identify conserved amino acid variation in the DNA binding Loop 2 region across the archaeal eukaryotic lineage and show that these residues tune recombinase interactions with double-stranded DNA. Guided by these evolutionary signatures, we engineer Rad51 variants with altered DNA binding properties and test their functions. Rad51 like variants promote break-induced replication (BIR), whereas substitutions characteristic of Dmc1 suppress BIR and instead favor crossover outcomes, as measured using mitotic crossover assays. These results indicate that evolutionary divergence in Loop 2 alters recombinase binding on reaction intermediates, influencing their stability and progression toward distinct repair outcomes. Thus, evolutionary tuning of recombinase DNA interactions shift the balance between recombination pathways, providing a mechanistic basis for the functional specialization of Rad51 and Dmc1 during mitotic and meiotic HR.

evolutionary biology↗

bioRxiv2025

A novel method for detection of Cas9 gRNAs using a fluorophore-labeled DNA oligo

Clustered regularly interspaced short palindromic repeats (CRISPR) associated protein 9 (Cas9) has become one of the most important gene editing tools. CRISPR guide RNA (gRNA) serves as an important component in guiding Cas proteins to a target site in gene editing processes. For effective gene editing or site specific gene integration, levels of gRNA present in a target cell are critical. Especially, for prime editing, even the ratio of different gRNAs present in target cells has a major effect on gene editing efficiency. Therefore, having a convenient, highly sensitive method for direct detection of gRNAs can improve editing design and optimize targeting efficiency. Here, we have developed a convenient, highly sensitive method for direct detection of gRNAs, which may be used to improve editing design and optimize targeting efficiency.

molecular biology↗

bioRxiv2025

Varigraph: an accurate and widely applicable pangenome graph-based variant genotyper for diploid and polyploid genomes

Graph pangenome references can address single-reference bias, thereby enhancing the accuracy of variant genotyping and empowering downstream applications in population genetics and quantitative genetics. However, existing pangenome-based genotyping methods struggle with large or complex pangenome graphs, particularly in polyploid genomes. Here, we introduce Varigraph, an algorithm that leverages the comparison of unique and repetitive k-mers between variant sites and short reads for genotyping both small and large variants. Varigraph outperforms current state-of-the-art linear and graph-based genotypers across non-human genomes while maintaining comparable accuracy in human genomes. By employing an efficient data structure, Varigraph achieves higher accuracy in repetitive regions while managing computational costs for large datasets. Notably, Varigraph is the first tool capable of effectively utilizing pangenome graphs for genotyping autopolyploids, enabling precise determination of allele dosage. This work provides a robust and accurate solution for genotyping non-human genomes and will facilitate genomic studies of polyploid crops.

genomics↗

bioRxiv2025

GPR182 is a lipoprotein receptor for dietary fat absorption

The lymphatic system plays a central role in lipid absorption, which transports chylomicrons from the small intestine to the circulation. However, the molecular mechanism by which chylomicrons get into the intestinal lymphatics is unknown. Here we demonstrated that GPR182, a receptor in lymphatic endothelial cells (LECs), mediates dietary fat absorption. GPR182 knockout mice are resistant to dietary-induced obesity. GPR182 ablation in mice leads to poor lipid absorption and thereby a delay in growth during development. GPR182 binds and endocytoses lipoproteins broadly. Mechanistically, loss of GPR182 prevents chylomicrons from entering the lacteal lumen of the small intestine. GPR182 blockage with a monoclonal antibody (mAb) protects mice from dietary induced obesity. Together, our study identifies GPR182 as a lipoprotein receptor that mediates dietary fat absorption.

physiology↗

bioRxiv2025

Mac/Lac-tosylceramide regulates intestinal homeostasis and secretory cell fate commitment by facilitating Notch signaling

Cell-to-cell communication via Delta-Notch signaling is widely used in various tissues and organs to regulate development and patterning; however, the mechanisms regulating Notch signaling for precise cell fate decisions remain poorly understood. Similar to mammals, the intestinal stem cells (ISCs) in the adult Drosophila midgut generate both absorptive and secretory cell progeny, guided by differential levels of Notch activation. Here we performed a forward genetic screen in Drosophila and identified glucosylceramide synthase (GlcT), a rate-limiting enzyme for glycosphingolipid (GSL) production, whose mutation causes the development of secretory cell tumors. Genetic analysis of the GSL synthesis pathway, combined with metabolite rescue experiments, revealed that the tumor formation is linked to a deficiency in Mactosylceramide/Lactosylceramide. This deficiency impaired the endocytic recycling of the Delta, subsequently reducing Notch signaling activation. Conditional knockout of UGCG, the mammalian ortholog of GlcT, in mouse small intestine caused an excessive differentiation of goblet cells, phenotypes similar to these caused by Notch inhibition. Our study suggests an evolutionarily conserved role for a specific GSL metabolite in modulating Notch signaling during stem cell fate decisions and provides a molecular connection between ceramide metabolism and Notch signaling in regulating tissue homeostasis and tumor formation.

developmental biology↗

bioRxiv2025

The hepatitis E virus capsid protein ORF2 counteracts cell-intrinsic antiviral responses to enable persistence in hepatocytes

Hepatitis E virus (HEV) is a significant human pathogen causing both acute and chronic infections worldwide. The cell-intrinsic antiviral response serves as the initial defense against viruses and has been shown to be activated upon HEV infection. HEV can replicate in the presence of this response, but the underlying mechanisms remain poorly understood. Here, we investigated the role of the HEV structural proteins ORF2 and ORF3 in immunocompetent cells. Mechanistically, we validated that ectopic ORF2, but not ORF3, interfered with antiviral and inflammatory signaling downstream of pattern recognition receptors, in part through interaction with the central adaptor protein TANK binding kinase 1. In the full-length viral context, ORF2 contributed to a reduced antiviral response and consequently, more efficient viral replication. In addition, we discovered a protective mechanism mediated by ORF2 that shielded viral replication from antiviral effectors. Using single-cell RNA-sequencing, we confirmed that the presence of ORF2 in infected cells dampened antiviral responses in both actively infected cells and bystanders. As a consequence, we found that early in the infection process, the progression of authentic HEV infection relied on the presence of ORF2, facilitating a balance between viral replication and the antiviral response within immunocompetent cells. Altogether, our findings shed new light on the multifaceted role of ORF2 in the HEV life cycle and improve our understanding of the determinants that may contribute to HEV persistence. Significance statementHepatitis E virus (HEV) is an important yet often underestimated pathogen. Depending on the genotype, HEV infections can progress to chronicity, but the underlying mechanisms remain poorly understood. To gain insight into potential determinants, we investigated how HEV evades the bodys first line of defense, the cell-intrinsic antiviral response. We discovered that the HEV capsid protein ORF2 is crucial in limiting this response by interfering with antiviral signaling pathways and shielding viral replication from immune effectors. This balance between viral replication and the antiviral response contributes to persistent HEV infection in immunocompetent cells. Our findings reveal a new role for the HEV capsid protein in the viral life cycle and highlight it as an important target for novel therapeutic approaches.

microbiology↗

bioRxiv2025

A Sensitive Soma-localized Red Fluorescent Calcium Indicator for Multi-Modality Imaging of Neuronal Populations In Vivo

Recent advancements in genetically encoded calcium indicators, particularly those based on green fluorescent proteins, have optimized their performance for monitoring neuronal activities in a variety of model organisms. However, progress in developing red-shifted GECIs, despite their advantages over green indicators, has been slower, resulting in fewer options for end-users. In this study, we explored topological inversion and soma-targeting strategies, which are complementary to conventional mutagenesis, to re-engineer a red genetically encoded calcium indicator, FRCaMP, for enhanced in vivo performance. The resulting sensors, FRCaMPi and soma-targeted FRCaMPi (SomaFRCaMPi), exhibit up to 2-fold higher dynamic range and peak {Delta}F/F0 per single AP compared to widely used jRGECO1a in neurons in culture and in vivo. Compared to jRGECO1a and FRCaMPi, SomaFRCaMPi reduces erroneous correlation of neuronal activity in the brains of mice and zebrafish by two- to four-fold due to diminished neuropil contamination without compromising the signal-to-noise ratio. Under wide-field imaging in primary somatosensory and visual cortex in mice with high labeling density (80-90%), SomaFRCaMPi exhibits up to 40% higher SNR and decreased artifactual correlation across neurons. Altogether, SomaFRCaMPi improves the accuracy and scale of neuronal activity imaging at single-neuron resolution in densely labeled brain tissues due to a 2-3-fold enhanced automated neuronal segmentation, 50% higher fraction of responsive cells, up to 2-fold higher SNR compared to jRGECO1a. Our findings highlight the potential of SomaFRCaMPi, comparable to the most sensitive soma-targeted GCaMP, for precise spatial recording of neuronal populations using popular imaging modalities in model organisms such as zebrafish and mice.

neuroscience↗

bioRxiv2025

Intense oocyte competition builds the female reproductive reserve in mice

During the process of ovariogenesis, more than two-thirds of germ cells are sacrificed to enhance the quality of the remaining oocytes. But the detail process of selection is not completely understood in mammals. Here, by creating a high resolution 4- dimensional ovariogenesis imaging system, we recorded the progress of oocyte fate determination in live mouse ovaries, and discovered an oocyte competition mechanism that determines the survival of oocytes. We found that dominant oocytes catch and absorb the cell debris from sacrificed oocytes to enrich their cytoplasm for survival. Single-cell sequencing demonstrated that this competition follows a classic autophagy- driven model. Blocking competition by suppressing autophagy resulted in an enlarged pool of survived oocytes. However, these oocytes lacking competition were unable to fully develop and contribute to fertility. Our study suggests that mammals have evolved a unique intensive competition system to enhance oocyte quality, necessary to support their long reproductive lifespan.

developmental biology↗

bioRxiv2025

DNA origami directed integration of colloidal nanophotonic materials with silicon photonics

Incorporation of colloidal quantum emitters into silicon-based photonic devices would enable major advances in quantum optics. However, deterministic placement of individual sub-10 nm colloidal particles onto micron-sized photonic structures with nanometer-scale precision remains an outstanding challenge. Here, we introduce Cavity-Shape Modulated Origami Placement (CSMOP) that leverages the structural programmability of DNA origami to precisely deposit colloidal nanomaterials within lithographically-defined resist cavities. CSMOP enables clean and accurate patterning of origami templates onto photonic chips with high yields. Soft-silicification-passivation stabilizes deposited origamis, while preserving their binding sites to attach and align colloidal quantum rods (QRs) to control their nanoscale positions and emission polarization. We demonstrate QR integration with photonic device structures including waveguides, micro-ring resonators, and bullseye photonic cavities. CSMOP therefore offers a general platform for the integration of colloidal quantum materials into photonic circuits, with broad potential to empower quantum science and technology.

bioengineering↗

bioRxiv2025

Multi-scale simulations of membrane adhesion mediated by CD47-SIRPα complexes

Adhesion of biological cells is essential for various processes, including tissue formation, immune responses, and signaling. It involves multiple length scales, ranging from nanometers to micrometers, which are characteristic of (a) the intercellular receptor-ligand binding that mediates the cell adhesion, (b) the spatial distribution of the receptor and lignad proteins in the membranes of adhering cells, (c) adhesion-induced deformations and thermal undulations of the membranes, (d) the overall size of the interface between adhering cells. Therefore, computer simulations of cell membrane adhesion require multi-scale modeling and suitable approximations that capture the essential physics of the system under study. Here, we introduce such a multi-scale approach to study membrane adhesion mediated by the CD47-SIRP binding, which is an immunologically relevant process. The synergetic use of coarse-grained molecular dynamics simulations and mesoscale kinetic Monte Carlo simulations allows us to explore both equilibrium properties and dynamical behavior of adhering membranes on the relevant length scales between 1 nm and 1 {micro}m on time scales ranging from 0.1 ns all the way up to about 20 s. The multi-scale simulations not only reproduce available experimental data but also give quantitative predictions on binding-induced conformational changes of SIRP and membrane-mediated cooperativity of the CD47-SIRP binding as well as fluctuation-induced interactions between the CD47-SIRP complexes. Our approach is applicable to various membrane proteins and provides invaluable data for comparison with experimental findings.

biophysics↗

bioRxiv2025

Metatranscriptomics sheds light on 'who is doing what' in the coral Porites lutea

Global decline of coral reefs due to climate change necessitates nature-based protection strategies for these crucial ecosystems. Developing such strategies requires a thorough understanding of the complex roles and interactions occurring within the coral holobiont. Using RNAseq, we investigated the active microbiome of healthy stony coral Porites lutea, focusing on the coral tissue, the green endolithic algal layer (Ostreobium layer), and the deeper coral skeleton. We identified distinct, metabolically active communities within these compartments and highlight substantial metabolic redundancy across carbon, nitrogen, and sulphur pathways. Our study provides first transcriptomic evidence of Ostreobiums ability to transfer fixed carbon to other holobiont members and the coral host. Additionally, we highlight critical roles of diverse coral holobiont members in nutrient cycling and maintaining homeostasis through scavenging of reactive oxygen and nitrogen species. This study provides novel molecular-level understanding of the functional roles played by diverse coral holobiont members in their respective compartments and underscores that corals harbour distinct microbiomes with wide-ranging functions.

microbiology↗

bioRxiv2025

Neuronal LAG3 facilitates pathogenic α-synuclein neuron-to-neuron propagation

Lymphocyte activation gene 3 (LAG3) is a key receptor involved in the propagation of pathological proteins in Parkinsons disease (PD). This study investigates the role of neuronal LAG3 in mediating the binding, uptake, and propagation of -synuclein (Syn) preformed fibrils (PFFs). Using neuronal LAG3 conditional knockout mice and human induced pluripotent stem cells-derived dopaminergic (DA) neurons, we demonstrate that LAG3 expression is critical for pathogenic Syn propagation. Our results show that the absence of neuronal LAG3 significantly reduces Syn pathology, alleviates motor dysfunction, and inhibits neurodegeneration in vivo. Electrophysiological recordings revealed that Syn PFFs induce pronounced neuronal hyperactivity in wild-type (WT) neurons, increasing firing rates in cell-attached and whole-cell configurations, and reducing miniature excitatory postsynaptic currents. In contrast, neurons lacking LAG3 resisted these electrophysiological effects. Moreover, treatment with an anti-human LAG3 antibody in human DA neurons inhibited Syn PFFs binding and uptake, preventing pathology propagation. These findings confirm the essential function of neuronal LAG3 in mediating Syn propagation and associated disruptions, identifying LAG3 as a potential therapeutic target for PD and related -synucleinopathies.

neuroscience↗

bioRxiv2025

High-pressure Golgi neuronal staining for X-ray whole-brain imaging

Whole-brain imaging has revolutionized neuroscience research, providing comprehensive insights into neural networks across the entire brain. This powerful approach has greatly advanced our understanding of brain functions and the mechanisms underlying various diseases. One primary challenge in whole-brain imaging technology is to achieve high-resolution observation of neural networks at large scales. Although Golgi method allows labeling random neurons in their entirety in the brain, visualizing individual dendritic trees, and tracing long-distance axonal projections, the lengthy processing time pose a limit on its use, i.e., staining a mouse whole-brain sample of just 300 mm3 takes over two weeks. Here, we developed a rapid staining technique for whole-brain neurons using high-pressure assisted Golgi (HP Golgi). This method significantly reduced the staining time for mouse whole-brain neurons from 16 days to only 4 days. We demonstrated the broad applicability of the HP Golgi method across various model organisms, achieving whole-brain neuronal staining in zebrafish, mice, and rats. Further, we successfully performed rapid staining of hippocampal neurons in an intact pig brain, which is difficult to achieve with the classic Golgi-Cox method. We also demonstrated that the combination of the HP Golgi method with synchrotron-based X-ray microscopy for high-resolution imaging of whole-brain neurons in mice. This HP Golgi method enables fast and high-resolution neuronal imaging in large model organisms, showcasing its broad applicability for diverse applications.

biophysics↗

bioRxiv2024

Cellular cartography reveals mouse prostate organization and determinants of castration resistance

Inadequate response to androgen deprivation therapy (ADT) frequently arises in prostate cancer, driven by cellular mechanisms that remain poorly understood. Here, we integrated single-cell RNA sequencing, single-cell multiomics, and spatial transcriptomics to define the transcriptional, epigenetic, and spatial basis of cell identity and castration response in the mouse prostate. Leveraging these data along with a meta-analysis of human prostates and prostate cancer, we identified cellular orthologs and key determinants of ADT response and resistance. Our findings reveal that mouse prostates harbor lobe-specific luminal epithelial cell types distinguished by unique gene regulatory modules and anatomically defined androgen-responsive transcriptional programs, indicative of divergent developmental origins. Androgen-insensitive, stem-like epithelial populations - resembling human club and hillock cells - are notably enriched in the urethra and ventral prostate but are rare in other lobes. Within the ventral prostate, we also uncovered two additional androgen-responsive luminal epithelial cell types, marked by Pbsn or Spink1 expression, which align with human luminal subsets and may define the origin of distinct prostate cancer subtypes. Castration profoundly reshaped luminal epithelial transcriptomes, with castration-resistant luminal epithelial cells activating stress-responsive and stemness programs. These transcriptional signatures are enriched in tumor cells from ADT-treated and castration-resistant prostate cancer patients, underscoring their likely role in driving treatment resistance. Collectively, our comprehensive cellular atlas of the mouse prostate illuminates the importance of lobe-specific contexts for prostate cancer modeling and reveals potential therapeutic targets to counter castration resistance. Significance StatementAndrogen deprivation therapy is a mainstay in prostate cancer treatment, yet many patients eventually develop castration-resistant disease--a lethal progression driven by poorly understood cellular mechanisms. Our study provides a comprehensive cellular map of the prostate, identifying key determinants of normal organization and castration-induced remodeling. By pinpointing the cell types and molecular programs that confer ADT responsiveness or resistance, our findings offer new directions for prostate cancer modeling and pave the way toward novel therapeutic strategies aimed at enhancing ADT efficacy and preventing the emergence of castration-resistant prostate cancer.

cancer biology↗

bioRxiv2024

Dissecting the evolving cellular landscape of a remyelinating microenvironment

Demyelination, or the loss of myelin in the central nervous system (CNS) is a hallmark of multiple sclerosis (MS) and occurs in various forms of CNS injury and neurodegenerative diseases. The regeneration of myelin, or remyelination, occurs spontaneously following demyelination. The lysophosphatidylcholine (LPC)-induced focal demyelination model enables investigations into the mechanisms of remyelination, providing insight into the molecular basis underlying an evolving remyelinating microenvironment over a tractable time course. Here, we present a detailed analysis using high-resolution single nucleus RNA sequencing to investigate gene expression dynamics across multiple cell populations involved in the remyelination process. We examine three specific time points following focal demyelinating injury in mice, and by delineating activation states within the heterogeneous cell populations of demyelinated lesions, we highlight changes in gene expression within subclusters of each cell population from the early stages of injury response to the initiation and maintenance of remyelination. Our findings reveal how shifts in microglial, astrocytic and fibroblast activities within lesions are associated with efficient oligodendrocyte differentiation during remyelination.

neuroscience↗