Biology subjects

Du, L.

Publications and source records attributed to Du, L..

At least 19 recordsLinked to original sources

bioRxiv2026

High-plex protein imaging across thick tissue by synchronized amplification

High-plex protein imaging is essential for decoding complex tissue organization, yet it remains largely confined to thin sections. Current approaches in millimeter-thick samples suffer from poor signal penetration and depth-dependent, uneven signal amplification. Even with specialized instrumentation to facilitate probe diffusion, iterative millimeter-thick tissue workflows can take more than four days per staining round, and reported eleven-marker experiments extend beyond a month. We introduce Amplification by Cyclic Extension (ACE), a DNA-barcoded, synchronized in situ amplification strategy that uses thermally controlled cyclic extension to coordinate DNA concatemer growth across tissue depths. By synchronously activating and coordinating reaction progression throughout the sample, ACE enables controlled, uniform signal amplification across tissue depths. Using ACE, we demonstrate 63-plex protein and RNA imaging in thin sections, 22-plex protein imaging in 200 m mouse brain sections, and 16-plex protein imaging in 1 mm-thick brain tissue encompassing over three million cells. For millimeter-thick tissues, ACE reduces per-round target-switching time from multiple days to less than eight hours, shifting volumetric high-plex protein imaging from months to days. Together, these results establish ACE as a synchronized amplification strategy that enables scalable high-plex protein imaging in thick tissues.

bioengineering↗

bioRxiv2026

Computer vision-aided locomotor behavioral analysis identifies therapeutic motor signatures in a mouse model of Huntington disease

Huntington disease (HD) is a neurodegenerative disorder characterized by progressive motor dysfunction. Traditional open-field tests quantify spontaneous locomotor parameters; however, fine mouse motor signatures, particularly disease stage-specific changes in HD motor symptoms and pharmacodynamic responses to therapeutic treatments. Here, we employed a computer vision-aided behavioral flow analysis designed to quantify fine, HD-relevant motor dysfunction in the zQ175DN HD mouse model, ranging from early HD-like motor signatures to well-defined motor deficits. Markerless pose estimation and Keypoint-MoSeq segmented standard top-view open-field recordings into recurrent behavioral syllables, which were then organized into higher-order clusters and transition networks. Disease stage-dependent changes in syllable occurrence, syllable duration, behavioral-state composition, and transition structure were identified. These analyses are not possible with traditional open-field assays. Syllable-duration features provided the strongest genotype discrimination, and HD-like motor features were also characterized by hub remodeling and transition-network disorganization. These features were integrated into an HD motor dysfunction (HDMD) score based on age- or HD progress-matched wild-type (WT) -standardized absolute deviations. The HDMD score distinguished HD mice from WT across multiple symptomatic stages and correlated with HD pathology and disease severity. Effect-size and power analyses suggested improved efficiency for detecting potential therapeutic effects. This framework requires only standard top-view recordings and may also support retrospective analysis of existing open-field video datasets. Overall, the HDMD framework provides a practical strategy for identifying fine motor changes in HD mice, aiding study design and preclinical efficacy assessment in HD drug development.

neuroscience↗

bioRxiv2026

Resolving Orsay Virus δ Protein Architecture Using Molecular Rulers in Single-Molecule Force Spectroscopy

Understanding the mechanical stability and architecture of viral proteins can provide valuable information about their biological function, but it remains a significant biophysical challenge. This study employs single-molecule force spectroscopy (SMFS) to investigate the multi-domain architecture of the Orsay virus {delta} protein, which lacks repeat structures and exhibits weak unfolding peaks. We engineered a construct using titin (I27)4 domains as an internal molecular ruler, enabling us to bracket the {delta} protein peaks to determine domain length and identify unfolding forces with an atomic force microscope (AFM). To address limitations of one-dimensional (1D) force distributions in resolving overlapping structural states, we created a two-dimensional (2D) mechano-structural signature map. By plotting kinetic stability (unfolding force F) against physical structural footprint (domain length L), we distinguished distinct unfolding domains, successfully separating degenerate 1D data into two statistically distinct populations corresponding to the {delta} proteins internal domain (I) and C-terminal domain (C). This label-free method provides the first mechanical evidence of the {delta} proteins multi-domain architecture. It establishes a robust, multi-dimensional framework for decoding the mechanics of complex biomolecular assemblies in their native state.

biophysics↗

bioRxiv2026

Endothelial Baf60c in BPD-Associated Pulmonary Hypertension

BACKGROUNDBronchopulmonary dysplasia-associated pulmonary hypertension (BPD-PH) complicates prematurity and carries substantial morbidity in extremely preterm infants. Pulmonary microvascular endothelial cell (PMVEC) dysfunction promotes capillary rarefaction and vascular remodeling, but epigenetic mechanisms after neonatal hyperoxia are poorly defined. Baf60c (SMARCD3), a SWI/SNF subunit supporting vascular homeostasis, and Smarcc2 (BAF170), a PBAF scaffold subunit linked to proliferative signaling, have not been studied together in BPD-PH. METHODSNeonatal C57BL/6 mice were exposed to 85% oxygen for 14 days. Right ventricular systolic pressure (RVSP), right ventricular hypertrophy, lung weight index, and pulmonary histopathology were assessed; PMVEC proliferation, migration, and invasion were measured. Transcriptome sequencing with GO/KEGG analyses, siRNA knockdown, LY294002 inhibition, coimmunoprecipitation, and Western blotting mapped the Baf60c-Smarcc2-PI3K-Akt-mTOR axis. A Tie1-driven, lung-tropic adeno-associated virus delivered by superficial facial vein injection at postnatal day 1 enabled PMVEC-specific Baf60c overexpression. RESULTSHyperoxia increased RVSP, right ventricular hypertrophy, and lung weight index, impaired alveolarization, reduced capillary density, and promoted arteriolar remodeling. PMVEC function was impaired, with PI3K-Akt pathway enrichment and suppressed signaling. Hyperoxia decreased Baf60c and increased Smarcc2. Baf60c knockdown upregulated Smarcc2, suppressed PI3K-Akt-mTOR, and phenocopied hyperoxia; Smarcc2 knockdown had opposite effects. Baf60c bound Smarcc2 but not PI3K. PMVEC-specific Baf60c overexpression attenuated pulmonary hypertension and right ventricular hypertrophy and partially improved alveolar and microvascular injury. CONCLUSIONSHyperoxia-induced BPD-PH is associated with reduced Baf60c, increased Smarcc2, and suppressed PI3K-Akt-mTOR signaling in PMVECs. Baf60c may indirectly regulate this pathway through Smarcc2. Endothelial Baf60c is a potential therapeutic target in BPD-PH. Novelty and RelevanceO_ST_ABSWhat Is New?C_ST_ABSA neonatal hyperoxia BPD-PH model was characterized with adverse hemodynamic and pulmonary vascular endpoints together with PMVEC functional impairment. Hyperoxia downregulates Baf60c, upregulates Smarcc2, and suppresses PI3K-Akt-mTOR signaling in PMVECs. Baf60c interacts with Smarcc2 without direct PI3K binding. PMVEC-specific Baf60c overexpression via superficial facial vein AAV delivery ameliorates pulmonary hypertension. What Is Relevant?BPD-PH is a developmental pulmonary hypertension phenotype in which pulmonary vascular resistance and right ventricular load are measurable with RVSP and Fulton index, consistent with Group 3 PH biology. SWI/SNF-mediated chromatin remodeling in pulmonary endothelium may link neonatal oxygen injury to impaired angiogenic endothelial function. Clinical/Pathophysiological Implications?Endothelial Baf60c deficiency may contribute to pulmonary vascular rarefaction after neonatal hyperoxia. Restoring endothelial Baf60c may improve pulmonary hemodynamics in BPD-PH, although structural lung injury may be only partially reversible.

physiology↗

bioRxiv2026

Disentangling polymer confinement from specific-folding interactions reveals the drivers of E. coli chromosome organization

The three-dimensional organization of the bacterial chromosome is critical for gene regulation. Chromosome conformation capture (Hi-C) has enabled genome-wide mapping of chromosomal folding, yet ensemble-averaged contact maps entangle biologically specific-folding interactions (SFIs) with nonspecific polymer compaction and mask single-cell heterogeneity. Here, we developed a polymer-based simulation framework in E. coli to address these limitations. We found that a null model of 300,000 random polymer configurations recapitulated the global chromosomal organization observed in Hi-C data, establishing that most Hi-C signals reflect generic polymer behavior in a confined volume. Contrasting null-model predictions with experimental Hi-C data isolated a small subset of SFIs (< 7%) and generated a specific-fold ensemble of 20,000 single-cell conformations that reproduced chromosomal interaction domains and single-cell heterogeneity. SFIs were enriched in the ter region, reduced nucleoid accessibility, colocalized with cryptic prophages, and depleted in positively supercoiled regions. H-NS and MatP emerged as major chromosome-wide and local determinants of SFIs, respectively. Furthermore, high-SFI regions correlated with stress-adaptive genes, whereas low-SFI regions harbored housekeeping genes. Together, our results established that a small number of biologically encoded SFIs superimposed on a polymer background shape the E. coli chromosome and gene expression, providing a quantitative framework for dissecting chromosome architecture and function.

genomics↗

bioRxiv2026

SigBridgeR: An Integrative Framework and Toolkit for Comprehensive Screening and Benchmarking of Phenotype-Associated Cell Subpopulations in Single-Cell Transcriptomics

Single-cell RNA sequencing has revolutionized our understanding of cellular heterogeneity, yet linking specific cell subpopulations to clinically relevant phenotypes remains a persistent challenge. Although multiple computational methods have been developed to bridge this gap, they are typically implemented as standalone packages with heterogeneous preprocessing pipelines, incompatible parameter conventions, and divergent output formats, thereby hindering rigorous cross-method benchmarking and reproducible multi-method workflows. Here, we present SigBridgeR, an extensible R framework and comprehensive toolkit that currently unifies eight state-of-the-art phenotype-associated cell screening algorithms within consistent workflows. We conducted a systematic benchmarking study across four cancer types HER2-positive breast cancer, triple-negative breast cancer, lung adenocarcinoma, and ovarian cancer using both binary phenotypes and patient survival endpoints. Our evaluation incorporated positive and negative control assessments based on differentially expressed genes and randomly selected marker panels, alongside quantitative accuracy comparisons using ground-truth cell labels. Building upon these insights, SigBridgeR provides standardized preprocessing for scRNA-seq and bulk transcriptomic data, unified algorithmic interfaces through a registry-based architecture, ensemble analysis via weighted voting, and comprehensive visualization utilities for multi-method comparison. By lowering technical barriers and promoting methodological standardization, SigBridgeR facilitates reliable discovery of phenotype-relevant cell subpopulations and enhances the translational potential of single-cell omics research.

bioinformatics↗

bioRxiv2026

High-resolution single-cell atlas of the human B cell compartment and immune microenvironment across tissues

The immune system is a dynamic network of diverse cell types distributed across tissues. While mouse studies have highlighted the importance of tissue-localized B-cell responses in infection and tissue repair, research on human B cells has remained largely confined to peripheral blood. Here, we specifically investigated the human B-cell compartment and its interactions with the immune microenvironment across 10 tissues, using single-cell RNA sequencing and paired B-cell receptor sequencing. We mapped the full spectrum of B-cell populations and revealed diverse differentiation trajectories spanning naive, memory, and plasma cell types. Germinal center B cells and plasma cells showed tissue-specific adaptations in transcriptional states, isotype usage, and functional profiles. Plasma cell isotypes influenced both effector functions and predicted interactions with T cells. Finally, we defined tissue-specific residency gene modules that outperformed existing memory B-cell signatures. Together, this dataset serves as a foundation for systematically studying tissue-localized B cells and reveals how local microenvironments shape humoral immunity.

immunology↗

bioRxiv2026

Spatial Multi-Omics Reveals Extracellular Matrix Remodeling and VSMC Phenotypic Switching in Moyamoya Disease

BackgroundMoyamoya disease (MMD) is a progressive cerebrovascular disorder characterized by steno-occlusive lesions and intimal hyperplasia. Although vascular smooth muscle cell (VSMC) phenotypic switching is implicated in its pathogenesis, the precise spatial interplay between extracellular matrix (ECM) remodeling and local metabolic alterations within the distinct vascular microenvironments remains unknown. MethodsSuperficial temporal artery (STA) samples from patients with MMD and controls were analyzed by histology, immunofluorescence, spatial transcriptomics, spatial proteomics, and spatial metabolomics. Single cell RNA sequencing was used to profile the cellular landscape of STA tissues. To functionally validate the identified pathway, human brain vascular smooth muscle cells (HBVSMCs) were stimulated with fibronectin 1 (FN1), and patient derived induced pluripotent stem cell smooth muscle cells (iPSC-SMCs) were generated for migration and protein expression assays following ITGA5 silencing or focal adhesion kinase (FAK) inhibition. ResultsMMD STA samples exhibited marked intimal hyperplasia with medial thinning and intimal accumulation of -SMA positive cells. Spatial transcriptomic and proteomic analyses identified an intimal remodeling program characterized by increased FN1, EFEMP1, fibronectin, ITGA5, and FAK, together with reduced MYH11. FN1 stimulation promoted smooth muscle cell migration, ECM associated protein expression, and FAK phosphorylation, whereas ITGA5 knockdown or FAK inhibition attenuated these effects. Patient derived MMD iPSC-SMCs showed similar abnormalities, including enhanced migration, increased FAK activation, reduced contractile markers, and increased ECM associated proteins. Spatial metabolomics and integrated multi-omics analyses further revealed that these changes were coupled to a metabolically depleted intimal niche enriched for reduced acyl-CoA related metabolites. ConclusionsIntegrated spatial multi-omics identifies coupled ECM remodeling and metabolic alteration in the hyperplastic intima of MMD. Within this context, the FN1-ITGA5-FAK axis emerges as a plausible mediator of smooth muscle remodeling that warrants further validation. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=165 SRC="FIGDIR/small/721225v1_ufig1.gif" ALT="Figure 1"> View larger version (90K): org.highwire.dtl.DTLVardef@3f6e19org.highwire.dtl.DTLVardef@554d49org.highwire.dtl.DTLVardef@451dd9org.highwire.dtl.DTLVardef@1aac1e9_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Characterizing the metabolomes of microglia, astrocytes, and neurons in aging and Alzheimer's brains

Neurons and glia are distinct in their morphology, development, and function, possessing unique transcriptomes and proteomes, but little is known about their metabolomes. The challenge of brain cell metabolic profiling is to obtain a large number of cells for reliable analysis. Here, we purified microglia, astrocytes, and neurons from mouse brains, identifying >70 metabolites through targeted metabolomics and 9,854 metabolite features via untargeted metabolomics. We systematically characterized cell type-enriched metabolites and metabolic pathways, revealing an enrichment of glutathione (GSH) and polyamine metabolism in microglia. This enrichment was validated in vivo and showed significant decreases with aging and in an Alzheimers disease (AD) model. Notably, GSH and polyamine metabolism correlated strongly with chemokine-related gene expression. Disrupting the GSH pathway in microglia resulted in downregulation of chemokine-related genes, aberrant morphogenesis, and {beta}-amyloid deposition. Our results provide a valuable resource (https://metabolismocean.org/braincell) for metabolic studies related to aging, AD, and other neurological diseases.

neuroscience↗

bioRxiv2026

Leptin Receptor+ cells create a perisinusoidal niche for thrombopoiesis in the bone marrow by synthesizing CXCL14

Leptin Receptor-expressing (LepR+) stromal cells in the bone marrow are a critical source of growth factors for the maintenance of hematopoietic stem cells (HSCs) and most restricted hematopoietic progenitors. An important unresolved question is whether they also regulate terminal differentiation in some hematopoietic cells. We found that LepR+ cells promote thrombopoiesis by synthesizing the chemokine CXCL14, which is expressed in the bone marrow by a subset of LepR+ cells. Cxcl14-expressing LepR+ cells extend fine processes that wrap around perisinusoidal megakaryocytes. Deletion of Cxcl14 from LepR+ cells did not significantly alter HSC function or most aspects of bone marrow hematopoiesis, including megakaryocyte generation; however, it significantly reduced the numbers of proplatelet-forming megakaryocytes in the bone marrow and platelets in the blood. CXCL14 promoted platelet formation by remodeling lipid metabolism in megakaryocytes, increasing fatty acid transporter expression and enabling megakaryocytes to take up more polyunsaturated fatty acids from circulation. A high fat diet rescued the formation of proplatelet-forming megakaryocytes and platelets in Lepr-cre; Cxcl14 fl/fl mice. CXCL14 protein was sufficient to promote platelet formation by megakaryocytes in vitro and in vivo. LepR+ cells thus create a perisinusoidal niche for thrombopoiesis by producing CXCL14, which regulates lipid metabolism and terminal differentiation in megakaryocytes. Key pointsO_LILeptin Receptor+ stromal cells regulate terminal differentiation in megakaryocytes in addition to maintaining stem and progenitor cells C_LIO_LICXCL14 from Leptin Receptor+ cells promotes the formation of platelets by remodeling lipid metabolism in megakaryocytes in the bone marrow C_LI

cell biology↗

bioRxiv2026

BaCas12a3 represents a new subtype of type V CRISPR effector with collateral activity toward tRNA

The CRISPR-Cas12 family encompasses diverse RNA-guided nucleases with both DNA-targeting and RNA-targeting subtypes. They can trigger antiviral activities mainly through either direct elimination invading nucleic acids, or activating broad collateral cleavage to induce abortive infection. Here, we report a novel type V CRISPR effector BaCas12a3 that causes growth inhibition through a unique tRNA-cleavage mechanism. Plasmid interference assays indicated that BaCas12a3 inhibits host growth arrest without invoking the DNA damage response, suggesting that the immune responses may not involve double strand breaks of DNA. Indeed, biochemical characterization of the BaCas12a3-crRNA ribonucleoprotein (RNP) unraveled that the effector is an RNA-activating nuclease that cleaves 3' terminus of tRNAs. Cryo-EM structures of BaCas12a3 reveal a conserved bilobed architecture featuring a unique nucleic acid-loading (NL) domain adjacent to the RuvC catalytic center. Structural and mutagenesis analyses show that the NL domain, together with a zinc ribbon domain, form a gated substrate groove. Target RNA binding induces conformational changes that open this groove and expose the RuvC active site, enabling specific tRNA cleavage while preventing other non-specific degradation. Our findings identified the NL domain aside the RuvC active site responsible for the tRNA recognition in BaCas12a3, expanding the functional diversity of CRISPR immunity.

molecular biology↗

bioRxiv2025

A high-resolution dimeric structure reveals a critical role of alpha-helix 9 in apoptotic Bax pore assembly in the mitochondrial membrane

Bax functions as a proapoptotic protein perforating the mitochondrial membrane to release mitochondrial proteins and DNA that kill the cell. Structures of full-length Bax monomer in solution and core domain dimers in solution and bound to lipid/detergent bicelles reveal how Bax proteins must be activated, unfold, refold and dimerize prior to forming oligomeric pores in the membrane. In particular, amphipathic core dimer forms part of the pore wall between the nonpolar lipid bilayer and the aqueous conduit. However, atomic resolution structures of other parts of the pore were scarce. Here, we elucidate a structure of Bax C-terminal -helix 9 (9) in lysolipid micelles using NMR. According to this high-resolution structure the 9 regions form an amphipathic helical dimer with an extended nonpolar interface and several uncharged polar residues on the surface. Structure-guided mutagenesis and functional assessment demonstrate that the nonpolar interactions are important for Bax dimerization in and perforation of the mitochondrial membrane. Surprisingly the polar residues are also important because they form bifurcated hydrogen bonds between helical turns to stabilize each helix and thereby the dimer. Molecular dynamics simulations of an oligomer constructed with wall-forming core and transmembrane 9 dimers linked by flexible 6-7-8 bridges in a mitochondrial lipid bilayer generate an atomic resolution model for a stable Bax pore capable to release cytochrome C. Thus, we made an important step toward elucidating molecular mechanisms of apoptotic Bax perforation.

biochemistry↗

bioRxiv2025

Ferritin Nanoparticle-Based Indirect ELISA for Immunodominant Region Screening and B-cell Epitope Validation of African Swine Fever Virus

African swine fever virus (ASFV) poses a severe threat to the global pig industry, creating an urgent need for effective vaccines and reliable diagnostic tools. While numerous ASFV B-cell epitopes have been reported based on murine-derived monoclonal antibodies (mAbs), such epitopes may not fully represent the antibody targets recognized in naturally infected pigs. Therefore, we need an approach to screen immunodominant regions by evaluating the reactivity of pre-defined epitopes with ASFV-infected pigs sera. In this study, we developed an indirect enzyme-linked immunosorbent assay (iELISA) based on recombinant epitope-ferritin nanoparticles (EFNs), as a platform for immunodominant region screening and epitope validation. This assay, hereafter referred to as EFNs-iELISA, shows an approximately twofold improvement in sensitivity compared with synthetic peptide-based ELISA for linear B-cell epitopes, and improved discrimination over GST-His-peptide and BSA-conjugated peptide formats. Using EFNs-iELISA, we evaluated 22 known murine-derived B-cell epitopes and 17 predicted epitopes from ASFV proteins CD2v, P30, P54, and P72 with ASFV-positive and negative porcine sera. According to the calculated cut-off threshold, tested epitopes were classified into different groups with positive or negative reactivity. Our analysis confirmed that among 22 known murine-derived ASFV B-cell epitopes, 16 (72%) also showed positive reactions with porcine-derived sera. Whats more, we also screened 7 new porcine-specific immunoreactive regions on these proteins. The EFNs-iELISA method provides a practical tool for screening immunodominant regions of ASFV, with potential applications in vaccine and diagnostic research.

immunology↗

bioRxiv2025

Semi-rational Engineering of Terminal Deoxynucleotidyl Transferase for High-efficiency Enzymatic de Novo DNA Synthesis

The 21st century has witnessed rapid advancements in synthetic biology, with DNA synthesis emerging as a foundational technology. Conventional phosphoramidite-based methods face significant limitations, including short DNA elongation lengths (<300 nt), hazardous chemical waste, and low stepwise incorporation efficiency. Enzymatic DNA synthesis using terminal deoxynucleotidyl transferase (TdT) offers a promising alternative, enabling kilobase-scale assembly with greater efficiency and minimal environmental impact. Here, we identified Bos taurus TdT (BtTdT) through UniProt database mining as a catalytically active scaffold for natural and 3'-modified dNTPs. Comprehensive characterization of BtTdTs enzymatic properties--including pH, temperature, metal ion dependence, and substrate specificity--revealed its optimal conditions. Truncation of the BRCT domain generated variants with enhanced activity compared to wild-type BtTdT. Guided by AlphaFold3-predicted structural models, we engineered a quintuple mutant (M5: Bt15AAR336L/K338G/L397M/E456S/D395G) optimized for 3'-ONH2-dNTP incorporation. M5 exhibited 30-fold activity enhancement relative to the triple mutant M3 (Bt15AAR336L/K338G/L397M) and achieved stepwise incorporation efficiency exceeding 98% in de novo synthesis of 10-nt ssDNA, demonstrating its potential for scalable enzymatic DNA synthesis. This work establishes a rational framework for TdT engineering through rational domain truncation and computational design, showing potential toward industrial-scale enzymatic DNA manufacturing.

synthetic biology↗

bioRxiv2025

Structural and Functional Insights into HBx-Smc6 Targeting for HBV Inhibition

The hepatitis B virus (HBV) X protein (HBx) is a multifunctional regulator essential for HBV replication and HBV-associated hepatocarcinogenesis. Despite its central role, the molecular basis of HBx function has remained elusive. Here, we present the first cryo-electron microscopy structure of the human HBx-CRL4-Smc5/6 complex at 3.1 [A] resolution. In this reconstituted ten-subunit assembly, HBx adopts a Zn2 {square}-stabilized Y-shaped architecture that simultaneously engages the DDB1 and the Smc6 subunit. A composite helix-turn-helix (HTH) pocket in HBx accommodates a conserved "Leucine Key" motif (LRCKL) on Smc6, forming a critical interface essential for complex stability and function. Molecular docking and biochemical validation reveal that the compound Tranilast binds this HTH pocket, disrupts the HBx-Smc6 interaction, and suppresses HBV replication. These findings define the structural mechanism by which HBx counteracts host restriction and establish the HBx-Smc6 interface as a previously unrecognized and druggable target for antiviral intervention.

cell biology↗

bioRxiv2025

A Chromatin-Structure-Guided Framework for Predictive and Interpretable Regulatory Genomics

Chromatin organization shapes gene regulation by linking distal elements across megabase scales, yet most predictive genomics models still treat the genome as linear, without incorporating three-dimensional structure. Hi-C provides genome-wide chromatin conformation information, but its contact maps are population-averaged, distance-biased, and noisy, obscuring the biologically specific contacts. We present CHROME, a framework built on a self-avoiding polymer ensemble null model that identifies physically specific, non-random Hi-C contacts. By integrating these contacts into graph representations, CHROME enables efficient information transfer across spatially connected loci. It integrates sequence, chromatin accessibility, or pre-trained embeddings into a graph attention architecture to predict cell line-specific ChIP-seq profiles, consistently outperforming local encoder baselines and generalizing to an unseen cell line. The resulting graph embeddings also enhance prediction on tissue-specific eQTL and ClinVar variant pathogenicity, outperforming local sequence-based embeddings. Beyond predictive performance, CHROME provides interpretability through attention-derived neighbor-to-center contributions that reveal how spatially connected loci influence local regulatory activity over multi-megabase distances. Together, these results show that incorporating physically validated chromatin interactions enables more accurate and interpretable modeling of gene regulation and variant effects.

genomics↗

bioRxiv2025

ChromPolymerDB: A High-Resolution Database of Single-Cell 3D Chromatin Structures for Functional Genomics

The three-dimensional (3D) organization of chromatin plays a critical role in regulating gene expression and genomic processes like DNA replication, repair, and genome stability. Although these processes occur at the individual-cell level, most chromatin structure data are derived from population-averaged assays, such as Hi-C, obscuring the heterogeneity of single-cell conformations. To address this limitation, we developed a polymer physics-based modelling framework, the Sequential Bayesian Inference Framework (sBIF), that deconvolutes bulk Hi-C data to reconstruct single-cell 3D chromatin conformations. To support a broader use of sBIF, we created ChromPolymerDB, a publicly accessible, high-resolution database of single-cell chromatin structures inferred by sBIF. The database contains [~]108 reconstructed 5 kb-resolution single cell structures, spanning over 60,000 genomic loci across 50 human cell types and experimental conditions. ChromPolymerDB features an interactive web interface with tools for 3D structural analysis and multi-omics integration. Users can explore associations between chromatin conformation and gene expression, epigenetic modifications, and regulatory elements. The platform also supports comparative analyses to identify structural changes across cell types, developmental stages, or disease contexts. ChromPolymerDB offers a unique resource for researchers studying the relationship between genome architecture and gene regulation, and for advancing comparative 3D genomics. ChromPolymerDB is available online at https://chrompolymerdb.bme.uic.edu/. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=85 SRC="FIGDIR/small/678816v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@110c807org.highwire.dtl.DTLVardef@bc713forg.highwire.dtl.DTLVardef@1af0ea9org.highwire.dtl.DTLVardef@1eba638_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

CASP3-GSDME-mediated trophoblast pyroptosis contributes to systemic inflammation

Early-onset preeclampsia (EOPE) is associated with excessive apoptosis and inflammation, but the mechanistic link between these processes remains enigma. Here, we report elevated circulating pro-apoptotic proteins in EOPE patients at early pregnancy, along with concurrent CASP3 activation and GSDME cleavage in a subset of EOPE placentas. Using multiple trophoblast cell lines, we demonstrate that trophoblast cells, which highly express GSDME, undergo a shift from apoptosis to CASP3-dependent pyroptosis, driving inflammation. Notably, pyroptotic trophoblasts further induce pro-inflammatory macrophage polarization within placental villi organoids, establishing a feedback loop that amplifies both trophoblast pyroptosis and inflammatory responses in trophoblast organoids-macrophage assembloids. In vivo, CASP3-GSDME-mediated trophoblast pyroptosis contributes to systemic inflammation in wild-type pregnant mice but not in Gsdme-/- mice. Screening of EOPE prevention drugs reveals Vitamin D as a suppressor of GSDME activation and pyroptosis in trophoblast cells. These findings highlight the CASP3-GSDME axis as a promising therapeutic target for preeclampsia prevention.

cell biology↗