Biology subjects

Yang, X.

Publications and source records attributed to Yang, X..

At least 199 records · Page 11Linked to original sources

bioRxiv2025

Brain endothelial PRMT5-ANGPTL4 axis regulates cerebellar inhibitory synaptogenesis and motor coordination

Brain vasculature is essential for central nervous system function, but its specific roles in synaptic development and motor function regulation are poorly understood. In this study, we identify a cerebrovascular signaling axis wherein endothelial PRMT5 critically regulates inhibitory synaptogenesis and motor coordination. Cerebrovascular-specific deletion of Prmt5 gene leads to excessive inhibitory synaptic input onto Purkinje cells and progressive motor deficits in mice. Mechanistically, PRMT5 deficiency epigenetically upregulates the expression of the secreted factor ANGPTL4 through reduced symmetric dimethylation of H3R8 and H4R3, along with increased H3K9 acetylation at the Angptl4 promoter, thereby driving excessive inhibitory synaptogenesis onto Purkinje cells. Importantly, Angptl4 deletion in brain endothelium normalizes inhibitory synaptic input onto Purkinje cells and restores motor coordination in PRMT5-deficient mice. Together, these findings define the endothelial PRMT5-ANGPTL4 axis as a key regulator of cerebellar inhibitory circuitry and motor function, highlighting cerebrovascular mechanisms as potential therapeutic targets for motor coordination disorders. Teaser Cerebrovascular PRMT5 prevents excessive inhibitory synaptogenesis via epigenetic repression of Angptl4.

neuroscience↗

bioRxiv2025

Hepatocyte FGFR2 Mediates the Antifibrotic Effects of FGF10 in Advanced Liver Fibrosis.

Fibroblast growth factor 10 (FGF10) supports epithelial repair, but its role in liver fibrogenesis remains uncertain. Here we found that hepatic FGF10 and its receptor FGFR2 decline with progression to advanced fibrosis in patient biopsies and in carbon tetrachloride (CCl4)- and diet-induced mouse models. Restoring hepatic FGF10 with recombinant human FGF10 or adeno-associated virus mediated liver-targeted expression attenuated and partially reversed established advanced bridging fibrosis, reduced inflammation, and decreased hepatocyte apoptosis, and these benefits required hepatocyte FGFR2 because hepatocyte-specific Fgfr2 deletion abolished protection. In primary hepatocytes, FGF10 activated FGFR2-FRS2, increased inhibitory GSK3{beta}(Ser9) phosphorylation, and suppressed NF-{kappa}B activation, reducing TGF-{beta}1 and other cytokines and thereby limiting paracrine hepatic stellate cells activation. The molecular and histologic benefits extended to metabolically primed steatohepatitis, supporting translatability beyond toxicant injury. Mechanistic signaling readouts confirm FGFR2 engagement, while the main text centers on the hepatocyte FGF10-FGFR2 axis as an intrinsic brake on fibrogenesis. Together, these data identify a hepatocyte-centric FGF10-FGFR2 axis as a negative regulator and potential regressor of fibrogenesis, highlighting it as a tractable therapeutic target in advanced liver fibrosis.

pharmacology and toxicology↗

bioRxiv2025

The cloacal outgrowth orchestrates co-development of the bladder and umbilical arteries

The urinary bladder is an innovation of eutherian mammals to manage liquid wastes. However, bladder development and its embryonic function remain poorly understood. Here, we report that in both humans and mice the bladder emerges from the cloaca as the rostral outgrowth, which is flanked by the umbilical arteries. Arrest of the outgrowth, as seen in Shh-null mouse embryos, causes bladder agenesis and unexpected severe defect of the umbilical arteries, suggesting that the outgrowth epithelial signal Shh coordinates development of both the bladder and umbilical arteries. Wnt2 signaling molecule is restricted to the rostral outgrowth mesenchyme; and its expression is dramatically down-regulated in Shh-null mutants. Furthermore, Wnt2-null mutants develop small bladder and single umbilical artery defects. Findings here uncover an unexpected co-development mechanism of the bladder and umbilical arteries orchestrated by the cloacal outgrowth signals. The co-development paradigm offers a conceptual framework to understand evolution and development of the bladder. Significance StatementThe urinary bladder is found in the eutherian mammals but largely absent from other vertebrates. Innovation of the bladder has been viewed as a necessity to manage liquid metabolic wastes on dry land. In this study, we report that the bladder emerges from the rostral outgrowth of the cloaca. We also report an unexpected developmental link between the bladder and umbilical arteries and, demonstrate that their co-development is orchestrated by the outgrowth epithelial signal Shh and mesenchymal signal Wnt2. Findings here highlight a possibility that, in addition to managing metabolic wastes, normal bladder development is required for growth of mammalian embryos by supporting development of the umbilical arteries.

developmental biology↗

bioRxiv2025

A disease mutation reveals a non-swapped pore and rewired gating cycle of the mechanically activated PIEZO1 channel

PIEZO1 is a versatile mechanotranduction channel critical for diverse cellular and physiological processes1-3. Gain-of-function mutations, such as R2482H, which slow channel inactivation, are linked to dehydrated hereditary stomatocytosis (DHS) and iron overload4-7. PIEZO1 forms a gigantic bowl-shaped trimer comprising flattenable peripheral blades and a central pore module with a switchable cap8-12. However, the structural dynamics governing its gating process and pathogenesis remain incompletely understood. Here, we determined distinct conformational states of the DHS-associated PIEZO1-R2482H mutant in both detergents and proteoliposomes. Remarkably, the R2482H mutation in the central pore induces a conformational change that propagates [~]25 nm to the most distal blade, enabling resolution of all 38 transmembrane helices. PIEZO1-R2482H is inherently flatter than PIEZO1, lowering the gating force and energy barrier and thus explaining its heightened mechanosensitivity. While the flattened PIEZO1 has its cap in the up-state adopting an inactivated state, the flattened PIEZO1-R2482H has its cap in the down-state and twisted by [~]75{degrees}, drastically switching the pore module from a swapped to a non-swapped organization. Such twisted conformation of PIEZO1-R2482H might hinder the transition to the inactivated state, instead proceeding through a deactivation-transition state characterized by curved blades and a partially twisted cap, thereby accounting for the slowed inactivation and deactivation kinetics. Taken together, we reveal the full transmembrane topology and a non-swapped pore conformation induced by R2482H, and propose the structural dynamic model that delineates the distinct gating cycle of activation, inactivation, and deactivation for PIEZO1 and PIEZO1-R2482H.

biophysics↗

bioRxiv2025

Modular mRNA lipid nanoparticle platform rescues diverse genetic male infertility

Genetic male infertility arises from diverse mutations that disrupt spermatogenesis, yet therapeutic strategies capable of restoring germ-cell function remain limited. Messenger RNA (mRNA) therapeutics offer a programmable and non-integrating approach, but their functional compatibility in structured and immune-privileged tissues remains unclear. Here we identify a temporal mismatch between mRNA delivery kinetics and the progression of spermatogenesis as a key barrier to functional rescue. We show that delivery efficiency does not predict outcome: formulations with high peak expression but short duration fail to support spermiogenesis, whereas sustained expression aligned with developmental timing enables recovery. Guided by this principle, we establish a testis-targeted mRNA delivery platform that restores spermatogenesis across multiple genetic models and generates functional sperm capable of supporting embryo development and multigenerational inheritance. The approach is modular, extends to human seminiferous tubules ex vivo, and is supported by comprehensive safety analyses. Together, these findings define a design constraint for RNA therapeutics in structured tissues: functional rescue requires temporal alignment between delivery kinetics and the therapeutic window defined by tissue-specific biological programs.

bioengineering↗

bioRxiv2025

An AI-driven approach for nanobody affinity maturation

B7-H3 (CD276), an immunoregulatory checkpoint molecule overexpressed in numerous cancers, is a promising therapeutic target. Nanobodies possess unique advantages for targeting B7-H3, such as small size, high stability, and the ability to bind cryptic epitopes. However, the rational affinity maturation of these nanobodies is challenging, especially in the absence of detailed structural data on antigen-antibody interactions. Here, we present a computational strategy that leverages artificial intelligence (AI) and molecular modeling, including homology modeling, molecular docking, and free-energy calculations--to systematically predict affinity-enhancing mutations for humanized anti-B7-H3 nanobodies. This AI-driven framework provides a powerful and cost-effective pipeline for accelerating the development of high-affinity nanobody therapeutics prior to experimental validation.

immunology↗

bioRxiv2025

Simple Methods to Acutely Measure Multiple Timing Metrics among Sexual Repertoire of Male Drosophila

Male Drosophila courtship behavior is a key model for studying temporal decision-making. While courtship index (CI) is widely used to quantify mating activity, other timing metrics like courtship latency, copulation latency (CL), and mating duration (MD) remain understudied. Traditional methods for quantifying these behaviors are often labor-intensive and prone to human error. In this study, we present a protocol combining a modular chamber system and automated software (DrosoMating) to quantify 6 key timing metrics during male courtship. Our image-based video analysis enables precise identification of courtship and copulation events, as well as quantification of their timing and duration under controlled conditions. Validation shows <0.05% error rates and 98-99% agreement with manual scoring for CL, CI, and MD. The protocol supports genetic and neural circuit manipulations, detecting subtle genetic, social, and environmental behavioral variations. By minimizing manual effort and standardizing data collection, this approach facilitates scalable, reproducible studies on adaptive trade-offs, learning, and neural mechanisms in mating behavior. This method streamlines timing analysis in male courtship, offering reproducible metrics for behavioral genetics. HighlightA high-throughput software pipeline for automated temporal profiling of Drosophila melanogaster mating behavior after brief user-guided calibration. An accompanying, open-hardware platform that can be assembled at minimal cost while maintaining experimental rigor. The system attains near-manual accuracy and outputs Temporal Measurement Parameters data that are readily adaptable to--and quantifiable within--diverse behavioral paradigms.

developmental biology↗

bioRxiv2025

RNA-Focused DNA-Encoded Library Construction, Screening, and Integration of Docking Identify Bioactive Ligands of Pathogenic r(G4C2)exp RNA

Disease-associated RNAs are increasingly recognized as promising therapeutic targets for small-molecule intervention. While DNA-encoded libraries (DELs) have long been established for protein ligand discovery, recent studies have demonstrated their feasibility for identifying RNA-binding small molecules. To further advance RNA-targeted ligand discovery, a diverse, solid-phase DEL enriched in privileged RNA-binding scaffolds was constructed and applied to identify ligands of r(G4C2)exp, a toxic RNA repeat expansion implicated in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). DEL selection outcomes were analyzed through large-scale molecular docking integrated with physicochemical and structure-activity relationship (SAR) analyses. Strong correlations were observed between docking predictions and experimental enrichment trends, supporting lead identification. The lead compound was subsequently optimized based on its docked pose to an NMR structure, resulting in analogs with enhanced binding affinity and bioactivity. These findings demonstrate that RNA ligand identification can be effectively achieved by combining DNA-encoded library technology with computational approaches for rational design and analysis, and highlight a broadly adaptable platform for RNA-targeted small molecule discovery. TOC graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/686979v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@1c9f24borg.highwire.dtl.DTLVardef@1422413org.highwire.dtl.DTLVardef@14e73a2org.highwire.dtl.DTLVardef@189a0bc_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2025

The dominance of gene expression controlled by trans-eQTL hotspots contributes to phenotypic heterosis in maize

Heterosis, or hybrid vigor, is a key phenomenon in genetics research and agricultural production, and has been primarily attributed to non-additive genetic effects such as dominance -- a prevailing consensus shaped by decades of empirical research and theoretical debate. Although dominance may arguably arise from distal modifiers, their selective advantage is debated due to presumably small individual effects. To address this long-standing question, particularly how genetic dominance manifests at the transcriptomic level and contributes to phenotypic heterosis, we integrated transcriptomic and phenotypic data from a large population of maize hybrids and their inbred parents. We found that [~] 30% of the expressed seedling genes in a significant proportion of hybrids exhibited expression patterns deviating from the average of the two parents, indicative of non-additivity. Further analysis suggests that while hybrid gene expression per se is primarily regulated by cis-eQTLs, expression dominance (or non-additivity) is disproportionately controlled by trans-eQTLs. These trans-eQTLs cluster into hotspots that regulate the non-additivity of hundreds of target genes, mostly within co-expression networks, and are notably enriched for transcription factors (TFs). Focusing on one such hotspot, we functionally validated a classical maize gene ZmR1, a basic helix-loop-helix (bHLH) TF associated with multiple seedling trait heterosis, as a candidate regulator of expression dominance across hundreds of genes. Overexpression of ZmR1 enhances expression dominance of downstream genes and increases phenotypic heterosis in both seedling and adult traits. Further experiments confirmed its direct regulatory role in modulating genes involved in anthocyanin biosynthesis and lignin metabolism, driving transcriptome-level dominance. These results provide empirical support for the modifier hypothesis under an omnigenic model, suggesting that heterosis arises not from the modification of a single genes inheritance but through the coordinated regulation of hundreds of phenotype-associated genes, thereby helping to reconcile the long-standing debate over the genetic basis of dominance in heterosis.

genetics↗

bioRxiv2025

Preclinical evaluation of antisense oligonucleotide therapy in a mouse model of HNRNPH2-related neurodevelopmental disorder

Mutations in HNRNPH2 cause an X-linked disorder characterized by developmental delay, intellectual disability, motor and gait disturbances, and seizures. Murine models that reproduce key clinical features of HNRNPH2-related neurodevelopmental disorder suggest that it may result from a toxic gain of function of the mutant protein or a complex loss of normal HNRNPH2 function with impaired compensation by its homolog, HNRNPH1. In this study, we tested gapmer antisense oligonucleotides (ASOs) that target murine Hnrnph2 in a non-allele-specific manner. The lead ASO reduced Hnrnph2 mRNA and protein levels while inducing compensatory upregulation of Hnrnph1 in both WT and Hnrnph2 mutant mouse brains. A single intracerebroventricular injection of the Hnrnph2 ASO into neonatal mutant Hnrnph2 mice rescued molecular and audiogenic seizure phenotypes and improved motor and cognitive functions. ASO treatment at the juvenile stage also rescued audiogenic seizures and motor deficits. In contrast, Hnrnph2 ASO administration did not improve survival, body weight, or hydrocephalus. In human iPSC-derived neurons, a human-specific HNRNPH2 research ASO reduced HNRNPH2 and upregulated HNRNPH1 mRNA levels. Mechanistically, we demonstrate that HNRNPH1 expression is regulated by alternative splicing and that HNRNPH2 modulates this process. These findings provide preclinical proof of concept for HNRNPH2 ASO therapy and offer insights into its underlying molecular mechanism. One Sentence SummaryASO-mediated Hnrnph2 knockdown induces Hnrnph1 upregulation and rescues phenotypes in a mouse model of HNRNPH2-related neurodevelopmental disorder.

animal behavior and cognition↗

bioRxiv2025

Live-Cell Covalent Profiling Reveals Principles of RNA-Small Molecule Recognition across the Human Transcriptome

RNA folds are abundant in mammalian cells yet poorly characterized as small-molecule targets. We present a scalable, unbiased live-cell pipeline that maps where small molecules bind RNA across the human transcriptome and convert those binders into selective degraders. A 200-member fragment library bearing diazirine/alkyne handles yielded 23 RNA-binding candidates. Chem-CLIP-Map-Seq in MDA-MB-231 cells identified 723 RNA targets and their binding sites, revealing a strong bias toward 5' and 3' untranslated regions (UTRs) in mRNAs and enrichment at thermodynamically stable structures, with limited binding to non-coding RNAs. Expression level and local stability contributed to engagement. An integrated machine-learning model trained on multiple fingerprints distinguished binders from non-binders, and highlighted chemotypes and physicochemical features that favor RNA recognition. Four fragments were converted to RiboTACs; despite broad binding, cleavage was highly selective, with X1-RiboTAC degrading MPP7 and SSC4D mRNAs in an RNase L-dependent manner and reducing their protein levels. A competitive profiling workflow quantified in-cell target occupancy and guided optimization of the RNA-binding module to reprogram selectivity: an X1 derivative produced an MPP7-selective RiboTAC that lowered MPP7 mRNA levels and suppressed breast-cancer cell migration, while sparing SSC4D transcripts. This end-to-end framework, including transcriptome-wide mapping, data-driven rules, and tunable degradation, establishes practical principles for ligandable RNA sites in cells and enables rational design of RNA-targeted small molecules and degraders. TEASERLive-cell mapping reveals ligandable RNA sites and guides design of selective RNA degraders. TOC graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=61 SRC="FIGDIR/small/686775v1_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@a216c8org.highwire.dtl.DTLVardef@185c59eorg.highwire.dtl.DTLVardef@17131e2org.highwire.dtl.DTLVardef@81ffa2_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2025

Domain-specific mechanisms of YAP1 variants in ocular coloboma revealed by in-vitro and organoid studies

The conserved transcriptional co-activator YAP1 is a central regulator of organ development and tissue homeostasis, integrating mechanical and biochemical cues to control cell proliferation and survival. YAP1 variants underlie a spectrum of congenital disorders, including autosomal dominant coloboma that can occur alone or with syndromic features. Despite this clinical significance, the functional role of YAP1 in human eye development, as well as the impact of disease-associated missense variants, remains poorly understood. Here we show YAP1 expression at the optic fissure in human embryos, a key structure involved in coloboma pathogenesis. We also identify a novel YAP1 variant in a proband with syndromic coloboma and investigate five previously reported coloboma-associated YAP1 variants. Using in silico prediction, cell-based assays, and fluorescence cross-correlation spectroscopy (FCCS) to directly quantify YAP1-TEAD binding, we demonstrate that the position of YAP1 missense variants dictates their functional changes. TEAD-binding domain mutations most strongly disrupted transcriptional activity in a luciferase assay, whereas all tested variants impaired induction of endogenous YAP1-TEAD target genes. Furthermore, mimicking reduced YAP1-TEAD binding using verteporfin small molecule in retinal organoids led to reduced progenitor proliferation and survival. These findings establish defective YAP1-dependent transcription as a mechanism driving congenital eye malformations and provide a framework for interpreting the pathogenicity of human YAP1 variants. More broadly, this study highlights the need for functional analyses to connect genetic variation with disease.

developmental biology↗

bioRxiv2025

Recruitment of the outer-membrane lipoprotein DolP to thedivision site via cardiolipin-mediated diffusion-stateswitching

In diderm bacteria, the outer membrane (OM) must invaginate in step with septal peptidoglycan (PG) remodeling during cytokinesis. One OM lipoprotein, DolP, has been shown to localize at the cell division site and to facilitate the daughter cell separation. Yet how DolP is recruited to the division site remains unclear at the molecular level. Here, we show that DolP arrives concomitantly with the late divisome protein FtsN. Utilizing single-particle tracking Photoactivated Localization Microscopy (spt-PALM), we investigated the dynamics of individual DolP molecules in living Escherichia coli cells. Single-molecule analysis revealed two diffusion states: a diffusive state across the cell envelope and a previously underappreciated immobile state enriched at the septal and polar regions. Importantly, DolP does not comigrate with the processive FtsW-FtsI-FtsN complex. Mutations of the cardiolipin-binding surface caused loss of mid-cell enrichment and reduced the immobile fraction. Cardiolipins preference for high negative curvature enriches it at the division site, creating a trapping environment for DolP. Additionally, the active septal constriction but not the late divisome proteins recruit DolP. Together, these findings indicate that cardiolipin-mediated immobilization underlies DolPs septal enrichment.

microbiology↗

bioRxiv2025

Direct identification of de novo mobile element insertions from single molecule sequencing of human sperm

Mobile element insertions (MEIs) are a significant source of human genetic variation, yet the rates and properties of de novo MEIs are poorly characterized due to technical limitations in sequencing technology. Here, we directly sequenced individual gametes from sperm samples of 19 donors (aged 27-62) using highly accurate PacBio long-read sequencing to identify de novo retrotransposition events without familial inference. We developed a "self-alignment" strategy using personalized genome assemblies that enables high-precision, single-read detection of de novo MEIs. Using this method, we identified 43 de novo Alu insertions, revealing >9-fold variation in Alu retrotransposition rates between individuals (ranging from 0 to 0.148 insertions/gamete). We found a significant increase in Alu activity with paternal age, yielding a 4.67% increase in insertions per gamete per year of additional paternal age, representing a direct observation of age-associated increases in structural variant (SV) mutation rates. De novo Alu insertions predominantly represent evolutionarily young AluYa5 and AluYb8 subfamilies and bear characteristic molecular signatures of target-primed reverse transcription (TPRT). Our population-averaged rate of 4.52 insertions per 100 gametes aligns well with previous population genetic estimates, validating both direct observation and population approaches for estimating de novo MEI rates. These results establish direct gamete sequencing as a powerful method for characterizing germline mutation processes and reveal age as a significant determinant of de novo retrotransposition in the male germline.

genetics↗

bioRxiv2025

M-DeepAssembly2: A Web Server for Predicting Multiple Conformations of Multi-domain Proteins Using Deep Learning

The biological functions of proteins often depend on dynamic transitions between multiple conformational states, and for multi-domain proteins, inter-domain dynamics play a critical role in their biological functions. Despite significant advances in static structure prediction, accurately modeling these multiple conformations remains a major challenge. Here, we present M-DeepAssembly2, a web server designed for modeling multiple conformations of multi-domain proteins using deep learning-predicted inter-domain and intra-domain constraints. The server operates in three stages. Firstly, evolutionary, physicochemical, and geometric features are extracted from the input and fed into AlphaFlex, a flexible residue prediction network. This process generates multiple distance maps that effectively capture both inter-domain and intra-domain interactions. Secondly, supplementary distance maps that characterize inter-domain interactions are generated using the DeepAssembly method. Finally, all distance maps are integrated as constraints in a multi-objective protein optimization to generate multiple conformational states of the protein. Compared to its predecessor, M-DeepAssembly2 leverages a flexible residue prediction network that directionally decouples coevolutionary information to generate heterogeneous distance maps. Benchmark results demonstrate that this strategy enables accurate prediction of protein multiple conformations and significantly improves the precision of static multi-domain protein structure prediction.

bioinformatics↗

bioRxiv2025

NyxBind: enhancing DNN representations via contrastive learning for TFBS prediction

While pretrained genomic language models effectively capture general DNA sequence patterns through masked language modeling, they often struggle to discriminate subtle yet biologically critical differences among transcription factor binding site (TFBS) motifs. Recent studies suggest that contrastive learning can enhance the discriminative power of embeddings by explicitly modeling inter-instance similarities and differences. Building on this insight, we introduce NyxBind, the first TFBS prediction model that applies contrastive learning across multiple TFBS types to enhance regulatory sequence representations. NyxBind better captures discriminative sequence features, enabling more accurate and biologically meaningful TFBS prediction. Extensive evaluations show that NyxBind consistently outperforms alternative models across multiple TFBS classification benchmarks, demonstrating strong robustness and generalizability. Moreover, NyxBind supports both full-parameter and parameter-efficient fine-tuning while maintaining high performance, and supports accurate motif visualization, aligning closely with experimentally validated transcription factor binding profiles. The code are available at https://github.com/ai4nucleome/NyxBind.

bioinformatics↗

bioRxiv2025

Graph-based tracing of dynamically functioning gene circuits in cell fate decisions with isoform resolution

Transcription factor (TF) regulatory networks govern complex cellular processes, yet their inference from single-cell transcriptomes is hindered by TFs low abundance, tissue-specific functions, and isoform diversity. We evaluated the utility of transcription factor activity (TFA) to infer TF-TF interactions, demonstrating its superiority over expression-based methods in capturing tissue-specific, high-fidelity regulatory relationships. Using simulated and real-world transcriptomes across human hematopoiesis and early embryonic development, we validated TFAs ability to render regulatory relationships. Through utilizing the concept of propagated regulome, we uncovered TFAs utility in retrospectively resolving dynamic regulatory circuits. To address regulome limitations, we developed an embryonic stem cell-specific Consensus REgulome Database (CRED) with isoform resolution, revealing functional heterogeneity among TF isoforms. Applied to human preimplantation single-cell datasets, CRED outperformed existing regulomes, identifying isoform-specific regulators and tracing regulatory modules despite severe dropouts. Our work unleashes TFA as a robust tool for reconstructing TF networks and highlights the importance of transited, isoform-level, cell-type-specific analysis in unraveling transcriptional regulatory circuits. HighlightsO_LITFA (Transcription Factor Activity) renders regulatory relationships with high fidelity compared to expression. C_LIO_LIUsing transited regulons enables TFA to retrospectively resolve dynamic regulatory interactions in cell fate decisions. C_LIO_LIAn integrated ESC-specific transited regulome database reveals isoform-level heterogeneity. C_LIO_LITFA successfully identifies key TF regulatory modules during human preimplantation from datasets with severe dropout. C_LI

systems biology↗

bioRxiv2025

Tumor-intrinsic MHC-II activation in pancreatic ductal adenocarcinoma enhances immune response and treatment efficacy

Pancreatic ductal adenocarcinoma (PDAC) is characterized by an immunosuppressive tumor microenvironment (TME) and poor prognosis. While major histocompatibility complex class II (MHC-II) expression is traditionally associated with professional antigen-presenting cells, its role in PDAC malignant cells remains underexplored. Herein, we utilized single-cell RNA sequencing (scRNA-seq), spatial transcriptomics, bulk RNA sequencing, multiplex immunohistochemistry (mIHC) and ex vivo studies in culture with both human and murine models to investigate the prognostic relevance of MHC-II expression in malignant PDAC cells. Elevated MHC-II expression in malignant cells was strongly associated with increased infiltration of CD4+ T and CD8+ T cells in human PDAC, and pronounced co-localization with plasma cells, indicative of an antigen-activated immune microenvironment. In the KPC mouse model of PDAC, pharmacologic induction of MHC-II expression by cobimetinib treatment in malignant epithelial cells significantly enhanced the therapeutic response to immune checkpoint blockade (ICB). These findings highlight the role of malignant cell- intrinsic MHC-II expression in promoting antigen presentation and fostering an anti-tumor immune microenvironment. Our results position MHC-II as a promising prognostic biomarker and therapeutic target in PDAC, paving the way for novel immunomodulatory strategies. Summary of highlightsSingle-cell and spatial transcriptomic analyses reveal that elevated MHC-II expression in malignant PDAC cells correlates with increased infiltration of CD4 and CD8 T cells. Stimulating MHC-II expression in tumors effectively enhances immunotherapeutic responses to ICB in the PDAC KPC mouse model, including PDAC tumors previously resistant to therapeutic interventions. MHC-II serves as a prognostic biomarker and a promising target for immunotherapy in PDAC.

cancer biology↗