Biology subjects

Kim, H. J.

Publications and source records attributed to Kim, H. J..

At least 19 recordsLinked to original sources

bioRxiv2026

Single-Cell and Spatial Analysis Reveals Retention of FCRL4⁺ Memory B Cells in Pediatric Tonsillar Hypertrophy

Pediatric tonsillar hypertrophy is a leading cause of sleep disordered breathing in children, yet the immune mechanisms sustaining tonsillar enlargement remain unclear. Using integrated single cell and spatial profiling of pediatric tonsils in a discovery cohort (n = 45) and independent validation cohort (n = 32), we identify a selective expansion and follicular retention of FCRL4+ memory B cells within the mantle zone of hypertrophic tonsils. These cells exhibit a quiescent, tissue-adapted phenotype characterized by NFATC1 repression and reciprocal RUNX1/RUNX2 regulation. Their accumulation is orchestrated by follicular type 1 regulatory T (Tr1) cells, which engage CTLA-4/CD86 checkpoint interactions to suppress plasma cell differentiation and reinforce a local immune tolerance program. Together, these interactions establish a persistent follicular niche enriched in regulatory FCRL4+ memory B cells. Our findings delineate a mechanistic Tr1/FCRL4+ B cell axis underlying follicular hypertrophy and highlight potential immunomodulatory targets for nonsurgical management of pediatric sleep-disordered breathing.

immunology↗

bioRxiv2026

Sonic Hedgehog Is An Important Regulator Of Intervertebral Disc Homeostasis And Rejuvenation

Intervertebral disc degeneration and associated neurological symptoms constitute a global health burden, yet no cure is currently available (1,2). Each disc consists of a central nucleus pulposus, surrounded by annulus fibrosus, and end plates connecting it to the growth plates of adjacent vertebral bodies. The notochord-descendant nucleus pulposus continues to express sonic hedgehog, which regulates the proliferation and differentiation of all components of neonatal mouse discs (3-7). Sonic hedgehog expression and function decline with age and are associated with disc pathologies, including terminal differentiation of nucleus pulposus cells to chondrocyte-like phenotype (7,8). Here, we used fate-mapping and conditional genetic mouse models to test the role of sonic hedgehog in the homeostasis of aging discs. We found that loss of NP-derived Shh is sufficient to accelerate multiple features of age-associated disc degeneration, including reduced NP cell number, altered matrix turnover, and induction of inflammatory, angiogenic and neurotrophic programs. Age-related disc pathologies were more prevalent in the lumbosacral discs of mice, like in humans (8-13). Also, like mice, the expression of sonic hedgehog and its targets by human nucleus pulposus cells declines with age and pathological degeneration. Moreover, pharmacologic Hedgehog activation partially restored anabolic gene expression and reduced catabolic, inflammatory and neurotrophic mediators in degenerated human NP explants ex vivo. These findings indicate that Sonic hedgehog, a developmental signal retained in the adult NP, is functionally active and required for disc homeostasis during aging and supports activation of hedgehog signaling as a candidate disease-modifying pathway for intervertebral disc pathologies.

developmental biology↗

bioRxiv2026

Ultrastructural and Proteomic Signatures of Mechanoadaptive Fibroblast Remodeling across Microphysiological and Mesoscale Shear Platforms

Mechanobiological cues in the tissue microenvironment increasingly drive pathological fibroblast activation in inflammatory bowel disease (IBD), yet engineered platforms modeling this transition remain limited. Here, a microfluidic gut-on-a-chip microphysiological system and a mesofluidic rotary shaker are used to demonstrate that sustained fluid shear stress alone is necessary and sufficient to drive an irreversible, profibrotic phenotypic switch in primary normal human intestinal fibroblasts. Across both platforms, normal fibroblasts from small and large intestine reproducibly self-organize into three-dimensional (3D) multicellular aggregates within 72 h, independent of shear delivery format, indicating that the transition is governed by mechanical dose rather than device geometry. The resulting aggregates acquire robust -smooth muscle actin (-SMA) expression with aligned stress fibers, contrasting with the -SMA-negative parental population. Scanning electron microscopy (SEM) resolves densely packed cellular microarchitecture embedded in a microfibrillar extracellular network, while ultrastructural serial block-face 3D EM reveals expansive intercellular spaces, stochastic fibrillar extrusions, and electron-dense cytoplasmic material at cell boundaries. Proteomic profiling confirms enrichment of core matrisome components, including collagen subtypes and matrix metalloproteinases. Together, these results establish fluid shear stress as a platform-independent, sufficient mechanical trigger for fibroblast-to-mechanoadaptive transition, positioning microphysiological shear platforms as tractable tools for modeling and targeting early fibrogenesis in IBD.

bioengineering↗

bioRxiv2026

Competing Molecular Interactions Govern the Dynamical Arrest of G3BP1 Condensates

G3BP1 is a central scaffold of stress granules (SGs). Upon cellular stress, G3BP1 forms complex coacervates with translationally repressed mRNAs and recruits multiple RNA-binding proteins to form reversible biomolecular condensates. Persistent SGs are linked to age-dependent dynamical arrest and impaired disassembly. Here, we employ active and passive nanoscale rheology with optical tweezers to show that G3BP1 condensates evolve from being dominantly viscous fluids to dynamically arrested network glasses characterized by nanoscale caging and elastic memory. Integrating atomistic and coarse-grained simulations with experiments, we find that electrostatic interactions between the oppositely charged intrinsically disordered regions drive condensate ageing. RNA modulates these interactions in a length-and structure-dependent manner, delaying dynamic arrest, whereas Caprin-1 binding to the NTF2L domain has little effect. Together, these findings reveal how competing inter-IDR and IDR-RNA interactions govern condensate ageing and material-state transitions. The findings have broader implications for the regulation of SG dynamics in cells.

biophysics↗

bioRxiv2026

Long-read sequencing reveals novel mitochondrial genome variants undetected by short-read sequencing in Korean population

While long-read sequencing technologies (e.g., PacBio Revio, ONT) have revolutionized high-quality genome assembly for the human pangenome, mitochondrial genome (mtDNA) analysis still largely relies on short-read and Sanger sequencing. However, short-read sequencing often lacks the resolution required to resolve complex variations due to the unique features of mtDNA, such as high mutation rates and repetitive homopolymeric regions, which frequently lead to alignment artifacts and mapping ambiguities. To address this, we evaluated whether applying long-read sequencing to mtDNA improves analytical quality in empirical data. Through comprehensive bioinformatics analyses, we compared the performance of long-read sequencing against short-read sequencing and microarrays. Our results revealed that long-read sequencing detected the highest number of variants (n = 533), significantly outperforming both short-read sequencing (n = 525) and microarrays (n = 49). Notably, both sequencing methods provided significantly higher resolution in haplogroup assignment compared to microarrays in terms of phylogenetic depth (p < 0.05). Long-read sequencing demonstrated superior detection power, particularly for InDels. We identified two novel non-synonymous variants, including a unique InDel detected exclusively by long-read sequencing. Protein modeling and stability analysis validated that this InDel causes structural instability (RMSD > 2.0 [A],{Delta}{Delta} G = -45.21 kcal/mol). Furthermore, we confirmed that this novel InDel is shared among haplogroup A samples in both the 1000 Genomes Project ONT dataset and the Korean population, highlighting the practical implications of long-read sequencing for molecular biology and population genetics.

genomics↗

bioRxiv2026

The anatomical compartment defines distinct immune remodeling within human visceral adipose tissue during aging

Age-associated inflammation varies across tissues, but whether distinct visceral adipose tissue (VAT) depots undergo inflammatory remodeling in a depot-specific manner remains unclear. Here we profiled human peri-organ VAT, comparing kidney-associated fat (KF, from kidney transplantation donors) and gallbladder-associated fat (GBF, from asymptomatic cholecystectomy patients with incidental polyps), using single-cell RNA sequencing, flow cytometry, intracellular protein profiling and in situ analysis. GBF showed broad tissue-residency and granzyme K-associated remodeling across conventional, regulatory and innate-like lymphocyte compartments, whereas KF showed stronger B cell remodeling, greater myeloid representation and more compartmentalized changes within resident effector-like CD8+ T cell states. We further identified age-associated CD20+ T cells with features consistent with local B-T cell interaction and an antigen-experienced, granzyme K-associated inflammatory memory phenotype. In situ analysis revealed age-associated myeloid accumulation and crown-like structure remodeling, accompanied by distinct myeloid inflammatory programs in KF and GBF. Finally, depot-specific immune signatures associated with clinical indices of adjacent kidney and liver function. These findings indicate that age-associated distinct immune programs within peri-organ VAT depots track with local tissue context and the state of the adjacent organ.

immunology↗

bioRxiv2026

The Spatial Landscape of Extracellular Matrix Gene Expression in Healthy and Type 2 Diabetic Human Pancreas

The unique peri-islet and double-layered vascular basement membrane (BM) of the human pancreatic islet are critical regulators of beta cell survival and function. While animal models imply that endothelial cells (ECs) are the exclusive source of islet BM, the precise cellular origins and spatial organization of the human islet matrisome remain poorly defined due to overlap in genes that mark non-epithelial cell populations and loss of spatial context during single-cell dissociation. In this study, we combine computational integration of whole-pancreas single-cell transcriptomes using CONCORD with high-resolution MERFISH spatial genomics to map the extracellular matrix (ECM) landscape across 251,477 spatially resolved cells from seven non-diabetic and five type 2 diabetic human donors. Contrary to an endothelial-centric paradigm, our data support a cooperative division of labor in the provision of BM, where pericytes represent the dominant transcriptional source of structural BM collagens (COL4A1, COL4A2) and ECs selectively express complementary matrix factors (HSPG2, LAMA5). Spatial neighborhood analysis further resolves a specialized population of islet-associated fibroblasts enriched at the islet boundary that are characterized by expression of peri-islet laminin genes. In type 2 diabetes, this homeostatic perivascular niche changes composition, marked by a significant increase in the islet fibroblast-to-pericyte ratio. Concurrently, islet pericytes undergo pro-fibrotic reprogramming characterized by the loss of canonical identity markers (PDGFRB), altered expression of ECM genes including COL1A2 and COL18A1, and upregulation of contractile machinery (MYL9). In the non-diabetic pancreas, pericytes constitute the principal vascular BM-expressing population within islets, whereas type 2 diabetes is associated with coordinated, compartment-specific remodeling of vascular-supportive stromal populations. Research in ContextO_ST_ABSWhat is already known about this subject?C_ST_ABSO_LIExtracellular matrix (ECM), and in particular basement membrane (BM), are essential structural and signaling components of the pancreatic islet microenvironment that contribute to beta cell function and survival. C_LIO_LIIslet capillaries are closely associated with endocrine cells and are surrounded by specialized BMs; however, the cellular sources of these BM components in the adult human pancreas remain incompletely defined. C_LIO_LIType 2 diabetes is associated with islet fibrosis and vascular dysfunction, but cell type-specific alterations in ECM-producing populations have not been comprehensively characterized in situ. C_LI What is the key question?O_LIWhich cell populations produce the components of ECM, including BM, within the adult human islet, and how are these populations altered in type 2 diabetes? C_LI What are the new findings?O_LISpatial transcriptomics identifies pericytes as the predominant vascular-associated source of ECM, including BM, gene expression in human islets, whereas endothelial cells exhibit complementary but more limited matrix-producing programs. C_LIO_LISpatial transcriptomics identifies an islet-associated fibroblast population enriched for fibrillar collagen and BM-associated genes that localizes preferentially to the islet surface niche. C_LIO_LIType 2 diabetes is associated with remodeling of perivascular ECM programs, including reduced expression of vascular basement membrane genes, a shift from a pericyte to smooth muscle-like identity, and increased expression of matrix-remodeling and fibrosis-associated genes. C_LI How might this impact clinical practice in the foreseeable future?O_LIDefining the cellular sources and disease-associated remodeling of the human islet ECM may inform the development of therapies aimed at preserving or restoring the islet microenvironment in type 2 diabetes. C_LIO_LIIncluding key subtypes of islet-associated ECM-producing cells may be important in improving current protocols to generate replacement islets from human pluripotent stem cells for cell replacement therapy for diabetes. C_LI

cell biology↗

bioRxiv2026

Spatial tumor microenvironmental architecture of chemoradiotherapy-resistant residual esophageal squamous cell carcinoma

PurposeResidual ESCC after neoadjuvant CCRT reflects incomplete treatment response and carries a high risk of recurrence. We sought to define the spatial tumor microenvironmental features of CCRT-resistant residual ESCC -- using tumor regression grade (TRG) 2-3 tumors as a model of poor pathologic response -- with mechanistic refinement at single-cell resolution and outcome assessment in internal and external cohorts. Experimental DesignWe profiled post-CCRT ESCC tissue microarray (TMA) cores by Visium FFPE whole-transcriptome spatial transcriptomics and orthogonal Xenium single-cell-resolution in situ profiling on serial sections from the same TMA blocks. Spot-level cell-type composition was inferred by CellDART deconvolution against a published ESCC single-cell RNA sequencing (scRNAseq). Local malignant-cell-enriched regions were defined by CancerFinder. We additionally re-analyzed the scRNAseq dataset to characterize SPP1 and CXCL5 macrophage states and their ligand-receptor signaling using CellChat. Outcome associations were evaluated in the SNUH cohort and externally tested in the TCGA ESCC cohort. ResultsTRG 2-3 residual tumors exhibited effector immune-cell exclusion from malignant-cell-enriched regions, SPP1/CXCL5 macrophage accumulation, microvascular rarefaction, and proliferative-metabolic reprogramming. Single-cell re-analysis confirmed that CXCL5 macrophages constitute a transcriptional subset of the broader SPP1 macrophage population. CellChat showed that SPP1 macrophages dominantly signal through SPP1-CD44 and SPP1-integrin axes to stromal and epithelial targets, and additionally engage immunosuppressive NECTIN2-TIGIT, CD86-CTLA4, and LGALS9-HAVCR2 programs with CD8 T cells, providing a mechanistic context for local immune exclusion. Visium LIANA and Xenium distance-gradient profiling localized SPP1-associated signaling preferentially to tumor cells and CAFs. Higher SPP1 expression and lower endothelial abundance were associated with shorter disease-free survival in the SNUH cohort. Higher SPP1 expression was also associated with shorter disease-free survival in the independent TCGA ESCC cohort. ConclusionsCCRT-resistant residual ESCC is characterized by a spatially organized tumor microenvironmental niche centered on SPP1-associated macrophage programs and microvascular rarefaction. These spatially resolved findings identify candidate macrophage- and vasculature-targeted axes for overcoming treatment resistance. Translational RelevancePatients with esophageal squamous cell carcinoma (ESCC) who harbor residual disease after neoadjuvant chemoradiotherapy (CCRT) remain at high risk of recurrence, yet the spatial biology of this resistant residual state has been poorly defined. Here, we combine Visium whole-transcriptome and Xenium single-cell-resolution spatial transcriptomics with single-cell RNA-seq re-analysis and external TCGA context to define the tumor microenvironmental architecture of CCRT-resistant residual ESCC. The resistant niche is characterized by effector immune-cell exclusion from malignant-cell-enriched regions, accumulation of SPP1/CXCL5 macrophage programs that signal to tumor cells and cancer-associated fibroblasts through SPP1-CD44 and SPP1-integrin axes, microvascular rarefaction, and proliferative-metabolic reprogramming. These findings define a coordinated spatial tumor microenvironmental signature of CCRT-resistant residual ESCC and nominate macrophage-tumor/stromal interactions and vascular injury as biologic axes that may help guide future strategies to overcome treatment resistance.

cancer biology↗

bioRxiv2026

CD1a-Mediated Presentation of Canonical Microbial Peptides to T Cells

Langerhans cells express the nonpolymorphic antigen-presenting molecule CD1a, positioning them as contributors to host immunity against Mycobacterium leprae in human leprosy. CD1a was originally shown to present non-canonical lipopeptide antigens such as dideoxymycobactin and chemically diverse hydrophobic ligands. Here, we generated CD4 T cell lines from leprosy lesions that recognized M. leprae in a CD1a-restricted manner. Unexpectedly, antigen recognition was protease-sensitive, prompting biochemical purification that identified two microbial protein antigens: LppX, a 25-kDa lipoglycoprotein, and Ag85A, a 30-kDa secreted protein with no known lipid modification. Recombinant proteins activated the corresponding T cell lines in a CD1a-dependent manner. Epitope mapping identified 12-mer peptides that fully reconstituted antigenicity, were conserved between M. leprae and M. tuberculosis, and elicited robust, dose-dependent IFN-{gamma} production and T cell proliferation, establishing that DNA-encoded, ribosomally translated peptides serve as CD1a-restricted cognate antigens. Biochemical analyses showed peptide binding to CD1a, supported by isoelectric focusing and surface plasmon resonance (KD [~]75 M for Ag85A). CD1a-peptide tetramers specifically stained cognate T cells, soluble CD1a was sufficient to present peptide antigen, and transfer of the LppX-specific TCR into naive T cells restored antigen responsiveness. Using CD1a-peptide tetramers, we identified antigen-specific T cells enriched in patients undergoing reversal reactions compared with patients with lepromatous leprosy and healthy donors. The CD1a-restricted T cell lines secreted IFN-{gamma} and IL-26, cytokines with established antimicrobial activity. Together, these findings demonstrate that CD1a can present canonical microbial peptides as part of a cell-mediated immune response in leprosy, extending the known spectrum of CD1a ligands. Because CD1a is nonpolymorphic and presents antigens to antimicrobial T cells, CD1a-peptide complexes may provide a broadly applicable platform for studying, detecting, and potentially targeting mycobacterial immunity.

immunology↗

bioRxiv2026

ADAR-Sense: an open-access, species-agnostic web tool for automated, user-customisable ADAR-based RNA sensor design

BackgroundEngineered synthetic RNAs enable cellular control by sensing and responding to intracellular biomolecules. Recently developed sense-edit-switch RNAs (sesRNAs) based on Adenosine Deaminase Acting on RNA (ADAR)--which edits a stop codon to switch on custom payload translation in the presence of a target RNA--are consistently functional across different species. The ability of sesRNAs to couple bespoke payload translation to the presence of cell type-specific transcripts will usher in an era of precise cell-targeted biotechnological interventions. ResultsTo expedite the generation of sesRNAs, we develop ADAR-Sense--a universal web tool for automated sensor design based on user-defined sensor length, sensor-target RNA mismatch number, mismatch proximity to the ADAR-editable stop codon, and targeted custom element inclusion for improved ADAR recruitment and subsequent payload induction. ConclusionsCompared to current tools, the simplicity and flexibility of ADAR-Sense will streamline the design and screening of sesRNAs in new cell types and conditions, supporting the swift adoption of this sensing platform in both basic and translational research.

synthetic biology↗

bioRxiv2026

Advanced Fabrication Protocol of an Elastic Porous Membrane for Organ-on-a-chip Applications

Elastic porous membranes are essential components of mechanically active organ-on-a-chip and microphysiological system (MPS) platforms, where cyclic strain is required to recapitulate physiologically relevant tissue mechanics. However, existing fabrication methods are often difficult to reproduce, low throughput, or dependent on specialized infrastructure, limiting their adoption across laboratories. Many protocols also lack quality control steps for ensuring device assembling and reproducibility. In this paper, we present a robust and accessible fabrication and quality control workflow for the consistent production of elastic porous PDMS membranes. The method uses commercially available heat presses, release liners, and pre-patterned membrane wafers to enable rapid membrane molding. We describe a quality control framework, including visual verification of porous regions and wettability testing for surface activation, to ensure irreversible PDMS bonding and reliable device assembly. Together, this workflow improves fabrication yield, reduces device failure, and supports reproducible implementation of elastic porous membrane in organ-on-a-chip applications.

bioengineering↗

bioRxiv2025

Alphaviral nonstructural protein-host RBP co-condensation as a mechanism to sustain virus replication

It has long been recognized that the intracellular replication of alphaviruses critically relies on several key host RBPs, including G3BP1/2 and FXR1/FXR2/FMR1, but how these RBPs modulate alphaviral replication and whether it would be possible to target these RBPs for antiviral treatment are less explored. Here, using SFV as a model, we report that SFV nsP3 exploits G3BP for its condensation and transforms antiviral stress granules into proviral nsP3-G3BP co-condensates. The gel-like co-condensates of nsP3 and G3BP enrich and protect viral genomic RNAs from host RNase degradation and serve as viral translational hubs to promote viral replication. The mode of nsP3-RBP co-condensation is prevalent across alphaviruses, and disruption of nsP3 condensates is an efficient antiviral approach. Thus, these findings uncover a general anti-alphavirus strategy based on the conserved reliance of nsP3-RBP co-condensation.

cell biology↗

bioRxiv2025

Variable Schwann cell merlin inactivation is targetable with TEAD1 inhibition in schwannomas.

Schwann cell tumors occur frequently in association with the vestibular nerves, leading to sensorineural hearing loss, and brainstem compression. In humans, unilateral vestibular schwannomas (VS) occur sporadically (VSspo)1, or bilaterally with neurofibromatosis type 2 syndrome (NF2) - VSnf2.2 VS formation is driven by sub-haploid NF2 gene dosage3, typically by biallelic loss.4,5 Loss of merlin promotes hippo/TEAD dependent transcriptional reprogramming, proliferation, and paracrine signaling that varies across time, and tumor volume.4,6 These variations lead to a clinically unpredictable course, and incomplete response to treatment. We hypothesized that Schwann cell merlin inactivation state determines cell-wise hippo/TEAD dependency and drives schwannoma pathogenesis. We analyzed clinical samples from VSspo and VSnf2 with a multi-omics approach and detected variation in merlin activity within tumor Schwann cell population. We found that tumor-driving merlin-depleted Schwann cells (Schwannmer-) exhibited elevated hippo activity that was predominantly driven by TEAD1. In-silico TEAD1 perturbation led to a reversal to merlin intact Schwann phenotype. These findings, and tumor cell growth suppression were confirmed in NF2fl/fl;Peri-Cre mouse model7, and in human derived schwannoma cells treated with a pan-TEAD auto palmitoylation inhibitor VT3989.8 Our computational and experimental results confirm that TEAD1 inhibition could be a potent, targeted strategy for schwannomas.

cancer biology↗

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

Unraveling diel regulation of cuticle biosynthesis

The plant cuticle is vital for growth and adaptation to environmental stresses. Although cuticle biosynthesis is dynamically regulated by environmental cues, the molecular mechanisms integrating these external signals with intracellular responses remain largely unknown. Here, we discovered that cuticle biosynthesis is precisely orchestrated by two distinct regulatory modules acting in synchrony with the diel cycle. Daylight is perceived by phytochrome B (phyB), which suppresses cuticular wax biosynthesis by degradation of PIF4, a phytochrome-interacting bHLH factor that activates wax biosynthetic genes. This suppression is alleviated when phyB itself is degraded by the E3 ubiquitin ligase LRB, leading to diurnal activation of PIF4. In contrast, loss-of-function and transcriptional assays of CFLAP1 demonstrated its direct negative role in cutin biosynthesis. At night, the E3 ubiquitin ligase COP1 mediates proteasomal degradation of CFLAP1, thereby promoting cutin accumulation via activation of BDG1. Together, these results reveal that two regulatory modules, LRB-phyB-PIF4 and COP1-CFLAP1 coordinate the diel regulation of cuticle formation, ensuring timely assembly of the protective barrier.

plant biology↗

bioRxiv2025

Roles of G-protein coupled receptors and mechanosensitive ion channels in pressure-induced chronotropy of lymphatic vessels

Active lymph pumping relies on the spontaneous contractions of collecting lymphatic vessels, whose contraction frequencies are exquisitely sensitive to changes in intraluminal pressure. This homeostatic and mechanosensitive mechanism, termed pressure-induced lymphatic chronotropy, enables lymph transport to be matched to the filling state of the lymphatic capillary network. The mechanistic basis of pressure-induced chronotropy was investigated using ex vivo contraction assays of mouse popliteal collecting vessels, in which contraction frequency increases >10-fold with pressure changed from 0.5 to 5 cmH2O. The contractile, electrophysiological and transcriptional similarities between lymphatic muscle cells (LMCs) and arterial smooth muscle led us to hypothesize that pressure-dependent chronotropy shares a parallel signaling process with pressure-induced arterial depolarization/constriction. Thus, we probed two major mechanisms: 1) pressure-induced activation of mechanosensitive cation channels, including TRPC6, TRPM4, PKD1/2, TRPV2 and ENaC, and 2) mechano-activation of GNAQ/GNA11-coupled G-protein receptors (GPCRs) that would generate second messengers to activate those channels. Contraction assays were combined with scRNAseq analysis of the respective targets, with maximum use made of transgenic mice to avoid non-specific effects of pharmacological inhibitors, particularly those used to block TRP channels. Our findings rule out significant roles for the above TRP channels and other putative mechanosensitive channels implicated in arterial myogenic constriction, as well as channels implicated in ionic pacemaking of other tissues. In contrast, smooth-muscle specific knock out or inhibition of ANO1 or IP3R1 significantly blunted the effect of pressure on frequency. Pressure-induced chronotropy was suppressed by [~]70-90% at all pressures in GNAQ/GNA11 double knockout vessels, but with responsiveness partially maintained at pressures above 5 cmH2O. Pressure-induced chronotropy was also suppressed after acute Gq/11 inhibition with YM254890, but was normal in vessels from GNA12/GNA13 double knock out mice. These results support a scheme whereby mechano-activation of one or more GNAQ/GNA11-coupled GPCRs generates IP3, which induces SR Ca2+ release through IP3R1 and drives depolarization through the activation of ANO1 Cl- channels. The major GPCRs expressed in LMCs were subsequently identified and ranked by scRNAseq analysis but knock out or pharmacological inhibition of each of the top 7 candidates failed to significantly affect pressure-induced chronotropy. Our results strongly implicate one or more GNAQ/GNA11-coupled GPCRs in mediating this homeostatic process; however, the specific mechanosensitive GPCRs remain to be identified.

physiology↗

bioRxiv2025

Integrative multiomic analysis links TDP-43-driven splicing defects to cascading proteomic disruption of ALS/FTD pathways

Loss of nuclear TDP-43 is a hallmark of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Although TDP-43 is known to regulate RNA processing, including repression of cryptic exons, we currently lack a systems-level understanding of the consequences of TDP-43 loss. To address this, we generated multiomic datasets, including RNA-seq and proteomics, from human iPSC-derived neurons depleted of TDP-43. We found that differentially spliced genes, many expressing cryptic exons, had the greatest protein reductions. Surprisingly, nearly half of differentially expressed proteins were neither mis-spliced, nor differentially expressed genes; most of these also had no reported mis-splicing in seven additional post-mortem and iPSC-derived neuron datasets. Integrative network analysis identified a high-confidence disease-specific subnetwork of over 700 interacting proteins, enriched for mRNA processing, synaptic function, and autophagy. Comparison with post-mortem ALS and FTD samples revealed convergent protein and pathway disruptions. We experimentally validated network-predicted effects of cryptic splicing in ATG4B, STMN2, and DAPK1. Our analyses reveal new TDP-43-dependent molecular cascades and nominate central genes as potential ALS/FTD therapeutic targets.

neuroscience↗

bioRxiv2025

A point mutation in the nuclear speckle protein and splicing factor SRRM2 is associated with amyotrophic lateral sclerosis and causes dysregulation of synapse-associated genes

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the progressive loss of motor neuron function. ALS is a multifactorial disease which can originate from complex genetic and environmental factors. The identification of risk factors and their molecular contribution to ALS expand our understanding of the disease mechanism. Here, we describe a family with dominantly inherited degeneration, which carries a mutation in the serine/arginine repetitive matrix 2 gene (SRRM2). SRRM2 is essential for nuclear speckle formation and a constitutive member of the RNA splicing machinery. To investigate how the mutation in SRRM2 contributed to the ALS pathogenesis, we examined its effect on a model cell line, where the point mutation was introduced in the endogenous gene. Surprisingly, we found that the resulting single amino acid exchange led to the loss of one protein-protein interaction, between SRRM2 and the splicing factor ACIN1. Transcriptome studies further revealed wide-spread differential gene expression, which converged on the dysregulation of synapse-associated pathways. Together, our findings identify SRRM2 as a novel ALS risk factor and provide mechanistic insights into how its mutation can be linked to ALS pathology.

molecular biology↗