Biology subjects

Chen, A.

Publications and source records attributed to Chen, A..

At least 73 records · Page 4Linked to original sources

bioRxiv2025

RNA Binding Proteins KhpA and KhpB Interact with Small Regulatory RNAs and Affect Global Gene Expression in Deinococcus radiodurans

Small regulatory RNAs (sRNAs) in bacteria often associate with RNA-binding proteins to gain intracellular stability and/or to enable regulatory efficiency. While much of the current knowledge about those sRNA binding proteins is derived from studies in Gram-negative organisms, the characterization of such proteins in Gram-positive species is still lagging behind. Here, we identified and characterized two sRNA binding proteins (KhpA and KhpB) in Deinococcus radiodurans, a Gram-positive bacterium that exhibits extreme resistance to radiation and other oxidative stressors. We demonstrate that KhpA and KhpB interact with key sRNAs in D. radiodurans and influence their stabilities. Although KhpA and KhpB interact with each other, they do not bind the sRNAs as a complex. Insightfully, KhpA and KhpB facilitate the interactions of the representative sRNAs PprS and Dsr9 in D. radiodurans with their respective mRNA targets, pprM and DR_1968. Through RNA-seq analysis, we further revealed that KhpA and KhpB have both overlapping and specific roles in a global gene regulation in D. radiodurans. Overall, this study expands our knowledge of posttranscriptional regulation in D. radiodurans and supports the growing consensus that KhpA and KhpB homologs constitute a new family of sRNA binding proteins in Gram-positive bacteria. IMPORTANCEThe bacterium Deinococcus radiodurans is the most radiation-resistant organism identified to date. Understanding the mechanisms of resistance of D. radiodurans is essential for leveraging this bacterium in biomedical and biomanufacturing applications. It was previously revealed that small regulatory RNAs (sRNAs) play crucial roles in the gene regulation of D. radiodurans. However, how these sRNAs are influenced by RNA binding proteins is poorly understood. Here we identified two conserved RNA binding proteins, KhpA and KhpB, as sRNA binding partners in D. radiodurans. These proteins affect the sRNA stability, sRNA-target interaction, and global gene regulation. Characterization of KhpA and KhpB will help us advance the understanding of how post-transcriptional network regulates the physiology and radioresistance of D. radiodurans.

molecular biology↗

bioRxiv2025

CSRefiner: A lightweight framework for fine-tuning cell segmentation models with small datasets

AO_SCPLOWBSTRACTC_SCPLOWRecent advances in spatial omics technologies have enabled transcriptome profiling at subcellular resolution. By performing cell segmentation on nuclear or membrane staining images, researchers can acquire single cell level spatial gene expression data, which in turn enables subsequent biological interpretation. Although deep learning-based segmentation models achieve high overall accuracy, their performance remains suboptimal for whole-tissue analysis, particularly in ensuring consistent segmentation accuracy across diverse cell populations. Existing fine-tuning approaches often require extensive retraining or are tailored to specific model architectures, limiting their adaptability and scalability in practical settings. To address these challenges, we present CSRefiner, a lightweight and efficient fine-tuning framework for precise whole-tissue single-cell spatial expression analysis. Our approach incorporates support for fine-tuning widely uaed segmentation models in the field of spatial omics, including recent published model Cellpose-SAM, while achieving high accuracy with very limited annotated data. This study demonstrates CSRefiners superior performance across various staining types and its compatibility with multiple mainstream models. Combining operational simplicity with robust accuracy, our framework offers a practical solution for real-world spatial transcriptomics applications.

bioinformatics↗

bioRxiv2025

Disruption of the developmental factor Otp in the adult forebrain reveals its diverse physiological functions

Orthopedia (Otp) transcription factor is a critical determinant in the development of the neuroendocrine hypothalamus, and its embryonic deletion results in lethality. Although Otp expression is maintained throughout life, its physiological function in adulthood is not well understood. Here, we generated a forebrain-specific, tamoxifen-inducible, conditional knockout mouse model to investigate the roles of Otp beyond development. Conditional deletion of Otp in two-month-old mice resulted in impaired stress responses, characterized by increased depressive-like behavior and elevated stress-induced cortisol levels. It also led to various metabolic changes, including reduced thyroid hormone levels and body temperature, a higher percentage of fat mass, and diminished responsiveness to ghrelin without affecting food intake, energy expenditure, or body weight. This composite metabolic phenotype was associated with reduced hypothalamic neuropeptides TRH, CRH, AgRP, and NPY expression. Our findings highlight the role of Otp in adult physiological functions as a key neuroendocrine integrator of adaptive stress response and energy balance.

neuroscience↗

bioRxiv2025

Live access to the emotional dynamics of REM sleep dreams in lucid dreamers with narcolepsy

Sleep helps regulate emotions, but it is still unclear whether -and how- the emotions we experience in dreams contribute to this regulation. To uncover the potential function of dream emotions, we must first understand what they are and how they unfold in dreams. The emotional content of dreams has mostly been studied using post-sleep dream reports, which provide a biased and static snapshot of a complex and dynamic experience. In this study, we took a more direct approach, accessing dream emotions in real-time. We asked twenty-four lucid dreamers with narcolepsy to report the emotional valence of their dreams, - positive, negative or neutral-, while still asleep, using predefined facial codes during daytime naps monitored with polysomnography. Of the 126 naps recorded, 62 contained at least one emotional code during REM sleep, yielding 191 codes in total. The ratios of positive and negative codes were evenly balanced per nap. The 33 naps with at least two codes allowed us to track the dream emotional dynamics. Over half of these naps showed opposite emotional valences (positive and negative). By measuring the time elapsed between codes, we estimated the average duration of a given dreams emotional valence in REM sleep to be about one minute. Positive emotions emerged on average earlier than negative ones during lucid REM sleep. These findings confirm the highly emotional nature of dreams and, more importantly, highlight that emotions in REM sleep dreams are fluid and fast-changing. Such emotional dynamics during REM sleep dreams may help us to better understand the mechanisms of the emotional regulatory function of dreams.

neuroscience↗

bioRxiv2025

Characterizing the Ion-Conductive State of the α7-Nicotinic Acetylcholine Receptor via Single-Channel Measurements and Molecular Dynamics Simulations

The 7-nicotinic acetylcholine receptor (7-nAChR) is a cation-selective Cys-loop receptor involved in diverse physiological processes and is an important therapeutic target. Multiple cryo-EM structures of putative open states are now available and their functional relevance is under active investigation. Here, we combined single-channel patch-clamp recordings with atomistic molecular dynamics (MD) simulations to assess the conductive properties of several 7-nAChR structures solved with different ligands. Simulations restrained to the respective cryo-EM structures produced only modest ion flux for all models, inconsistent with experiment, whereas fully unrestrained simulations revealed marked differences in their ability to relax into physiologically conductive ensembles. Two structures, 7KOX and ligand-bound 8V82, consistently stabilized into conductive states whose permeation properties matched our measured inward single-channel conductance. The conduction of 8V82 nearly stopped upon removing the modeled ligands. 8V80 showed only intermittent conduction with ligands and remained non-conductive without them. 7EKT collapsed into a non-conductive conformation upon relaxation, irrespective of whether the modeled ligands were retained or removed. 9LH5, despite having a transmembrane pore nearly identical to 7KOXs, exhibited approximately twofold higher conductance, likely due to a widened extracellular vestibule. Across models, permeation events followed Poissonian statistics with a characteristic entry lag captured by a double-Poisson model. Simulations of outward currents consistently overestimated the conductance compared to experiments, perhaps reflecting the absence of the full intracellular domain in available structural models and/or the presence of current-blocking concentrations of cytosolic Mg2+ in patch-clamp cell-attached recordings. These results identify the conformations most compatible with the physiological open state and underscore the importance of unrestrained MD, ligand stabilization, and extracellular-vestibule geometry in shaping 7-nAChR conduction.

biophysics↗

bioRxiv2025

Hydrogen Sulfide Suppresses Keratinocyte Migration in Scratch Assay

Cell migration is crucial for maintaining physiological functions in the body and is driven by mechanical, electrical, or chemical stimuli. Hydrogen sulfide (H2S), a gaseous signaling molecule, has been shown to regulate cellular migration in a cell type and context-dependent manner. Keratinocytes are the predominant cell-type in the epidermis and drive re-epithelialization during wound healing. However, the effect of H2S on keratinocyte migration remains incompletely defined. Here, we demonstrate that H2S suppresses keratinocyte migration in a serum-restricted in vitro scratch assay. Intracellular H2S levels were modulated using fast-releasing donors H2S donors (NaHS or Na2S) to achieve supraphysiological levels or by siRNA knockdown of endogenous H2S-producing enzymes cystathionine-{gamma}-lyase (CGL) and cystathionine-{beta}-synthase (CBS) to obtain infraphysiological levels. Treatment with NaHS or Na2S (10 {micro}M-5 mM, 24 h) reduced gap closure in a dose-dependent fashion, with negligible effects at [≤] 100 {micro}M and progressive suppression at higher concentrations. Conversely, siRNA knockdown of CGL or CBS boosted keratinocyte migration by 30% versus scrambled siRNA control. Supplementation with a slow-release H2S donor rescued the siCGL phenotype, restoring migration to control levels. Across experiments, intracellular H2S level is negatively associated with keratinocyte migration. These findings suggest that H2S can restrain keratinocyte motility under serum-restricted conditions, and that reports of accelerated wound closure with H2S donors in vivo may reflect effects on other cell types and/or inflammatory pathways rather than direct enhancement of keratinocyte migration. Significance StatementDysfunctional wound healing affects millions worldwide. While H2S donors can accelerate wound closure in animal models, the cell-type-specific effects remain unclear. We show that, H2S suppresses keratinocyte migration implicating non-keratinocyte mechanisms (e.g., endothelial, immune, fibroblast) in the in vivo benefit of H2S donors and refining where in the healing cascade H2S is most likely to help.

bioengineering↗

bioRxiv2025

The Amplified Burden of Depression in Drug-Resistant Epilepsy: Role of Limbic Network Dynamic Reconfiguration

ObjectivesTo explore potential associations between drug resistance and limbic network (LN) dynamic functional interactions in temporal lobe epilepsy (TLE), whether these LN alterations are associated with and potentially mediate comorbid depression severity, and their potential neurochemical and transcriptomic underpinnings. MethodsThis cross-sectional study included 49 patients in the Responsive group, 33 patients in the Resistant group, and 50 healthy controls (HCs). Using resting-state fMRI, we derived LN dynamic integration coefficients and used non-negative matrix factorization (NMF) to identify coactivation patterns. Depressive symptoms (Self-Rating Depression Scale, SDS), LN integration, and NMF pattern expression were compared across groups (ANOVA) and correlated. Mediation analysis tested if LN integration mediated the relationship between drug resistance status and SDS scores. LN integration differences were spatially correlated with neurotransmitter receptor density and Allen Human Brain Atlas transcriptomic data. ResultsPatients in the Resistant group reported significantly higher SDS scores than patients in the Responsive group (p = 0.008). LN integration with the dorsal attention network (DAN) and fronto-parietal network (FPN) was significantly lower in the Resistant group compared to the Responsive group (p = 0.001; p < 0.001). NMF analysis identified key LN-DAN (p = 0.013) and LN-FPN (p < 0.001) coactivation patterns. In TLE patients, DAN-LN integration was negatively correlated with depressive symptoms (r = -0.28, p = 0.01), and lower integration levels significantly mediated the relationship between group status (Resistant vs. Responsive) and increased symptom severity (p < 0.001). These LN integration coefficient differences (Responsive group vs. Resistant group) were associated with 5-hydroxytryptamine 1B (5-HT1B; r = 0.29, p = 0.036) and dopamine D2 (D2; r = -0.29, p = 0.034) receptor densities, and linked to gene expression pathways including telomerase activity regulation (p < 0.001). InterpretationThis study identifies dynamic LN dysregulation, supported by distinct neurochemical and transcriptomic profiles, as a core bridging mechanism potentially contributing to comorbid depression linked to drug response status in TLE patients.

neuroscience↗

bioRxiv2025

GLM-Prior: a nucleotide transformer model reveals prior knowledge as the driver of GRN inference performance

Gene regulatory network (GRN) inference depends on high-quality prior knowledge, yet curated priors are often incomplete or unavailable across species and cell types. We present GLM-Prior, a genomic language model fine-tuned to predict transcription factor (TF)-target gene interactions from nucleotide sequence, and benchmark it as a sequence-derived strategy for constructing TF-gene prior matrices. We integrate GLM-Prior with PMF-GRN, a probabilistic matrix factorization model, to create a dual-stage pipeline that combines sequence-derived priors with single-cell gene expression data for prior-conditioned GRN inference. Across six human, mouse, and yeast cell-line contexts, GLM-Prior performance scales with positive label abundance and TF coverage in training data, and shows abovechance agreement with independent reference networks in well-annotated mammalian contexts. We evaluate single-species, species-transfer, and multi-species training paradigms, finding that GLM-Prior can construct informative priors across related mammalian species, while transfer to yeast remains near chance. Comparisons with accessibility-based priors across multiple GRN inference methods show that GLM-Prior achieves the highest prior performance in four of five mammalian cell lines. Under these benchmarks, prior quality largely constrains achievable GRN inference performance, with expression-based inference providing prior-dependent refinement. Together, these results position GLM-Prior as a benchmarked workflow for transferable, sequence-derived prior construction in systems where matched experimental assays are unavailable.

genomics↗

bioRxiv2025

Uncovering the Regional and Cell Specific Bioactivity of Injectable Extracellular Matrix Biomaterials in Myocardial Infarction through Spatial and Single Nucleus Transcriptomics

Myocardial infarction (MI) remains a global health concern. To mitigate MI pathophysiology, we previously investigated a pro-reparative decellularized extracellular matrix (ECM) hydrogel for treating subacute and chronic MI. Despite increasing interest in biomaterial scaffolds, single cell and spatially resolved transcriptomics have not been used to probe their therapeutic activity in the heart. Here, we utilize spatial transcriptomics and single nucleus RNA sequencing to delineate the regional and cell-specific bioactivity of ECM biomaterials. ECM hydrogel subacute treatment induced cardiac resident macrophage preservation, fibroblast activation, and increased lymphatic, vasculature, smooth muscle and cardiomyocyte development as well as neurogenesis. Chronic treatment elicited macrophage polarization, cardiomyocyte and vasculature development, alongside fibroblast development. When comparing treatment timepoints, subacute administration had stronger immune modulation, while the chronic timepoint demonstrated higher cardiac development markers. Both subacute and chronic administration were associated with fibroblast activation and vasculature development. Thus, we elucidate undiscovered therapeutic targets of the ECM hydrogel, further demonstrating the potential of ECM biomaterials as an MI therapy.

bioengineering↗

bioRxiv2025

An atlas of cell type specific regulatory effects in cattle

Understanding the genetic and molecular architecture of complex traits and artificial selection is crucial for advancing sustainable precision breeding in cattle and other livestock. Yet, how genetic variation affects cellular gene expression remains elusive in cattle. Here, by integrating 8,866 bulk RNA-seq samples and 999,192 single cells of 81 cell types in 22 bovine tissues, we presented a comprehensive atlas of regulatory variants at the cell type resolution in cattle. By colocalizing with bulk-tissue expression quantitative trait loci (beQTL), we detected 57,043 novel cell-type stratified eQTL and cell-type/state interaction eQTL in 18,153 genes, which also exhibited a stronger tissue/cell-type specificity than beQTL. By examining genome-wide associations (GWAS) of 44 complex traits, these cell-resolved eQTL were colocalized with 505 (24%) additional GWAS loci compared to beQTL. Through integrating this resource with selection signatures between dairy and beef cattle, we provided tissue/cell-specific regulatory insights into cattle breeding. Overall, the current atlas of cell-type-specific regulatory variants will serve as an invaluable resource for cattle genomics and selective breeding.

genetics↗

bioRxiv2025

Metformin boosts mitochondria and neurogenesis via AMPK/mTOR/SIRT3 in POLG mutant organoids

BackgroundMutations in the POLG gene, encoding the catalytic subunit of mitochondrial DNA polymerase gamma, are the most common cause of mitochondrial diseases affecting the central nervous system. These mutations frequently result in neurodevelopmental disorders, yet the cellular and molecular mechanisms underlying POLG related encephalopathies remain poorly understood. In particular, how POLG mutations affect mitochondrial function, neural progenitor behavior, and early neurogenesis in the developing human brain has not been fully elucidated. MethodsTo investigate the impact of POLG mutations on human neurodevelopment, we generated 3D cortical brain organoids from induced pluripotent stem cells (iPSCs) derived from a patient carrying compound heterozygous POLG mutations (A467T/W748S). Organoid development was monitored using immunohistochemistry, transmission electron microscopy, and live-cell mitochondrial assays. Single-cell RNA sequencing (scRNA-seq) was performed to profile cellular diversity and transcriptional changes. Organoids were treated with metformin, a known mitochondrial modulator, and mitochondrial function was assessed by measuring membrane potential (TMRE), ATP production, and mtDNA copy number. Western blotting and immunofluorescence were used to investigate AMPK-SIRT3-mTOR signaling and markers of mitochondrial dynamics and mitophagy. Statistical analyses were performed using unpaired t-tests or ANOVA, with p-values < 0.05 considered significant. ResultsPOLG mutant cortical organoids exhibited impaired neural differentiation, with expansion of stress-associated neural progenitors and reduced neuronal populations. scRNA seq analysis revealed transcriptional signatures of oxidative stress, mitochondrial dysfunction, and suppressed neurogenesis, particularly in a specific neural progenitor subpopulation. Metformin treatment significantly improved mitochondrial membrane potential, ATP output, and mtDNA copy number in POLG organoids. It also promoted neuronal differentiation and reduced reactive progenitor states. Mechanistically, metformin activated AMPK and SIRT3, inhibited mTOR, enhanced expression of mitochondrial fusion and biogenesis markers (OPA1, PGC-1), and increased autophagic and mitophagic activity (LC3B, BNIP3). ConclusionsOur study demonstrates that POLG mutations disrupt early cortical development by impairing mitochondrial function and skewing progenitor fate. Metformin mitigates these effects by restoring mitochondrial homeostasis and promoting neurogenesis through the AMPK/SIRT3/mTOR axis. These findings offer mechanistic insights into POLG-related encephalopathy and support metformin as a candidate for therapeutic intervention in mitochondrial neurodevelopmental disorders.

neuroscience↗

bioRxiv2025

The biofuel producing legume tree, Pongamia pinnata, offers strong potential for carbon sequestration

Pongamia pinnata (Linn.) Pierre, also called Millittia pinnata or Indian beach, is a fast-growing legume tree that has gained considerable attention for its potential as a feedstock for biofuel. In this study, we assessed the growth characteristics and carbon sequestering potential of Pongamia grown under sub-tropical environments. At two different sites located in Queensland, Australia, Pongamia was found to produce considerable biomass, even when planted in nutrient-poor environments having low soil organic carbon. The seedlings planted were all derived directly from genetically diverse, yet related seed. Variability observed in growth appeared to be associated with the mother tree from which the seeds were sourced, underscoring the importance of planting elite genetic material for commercial purposes. Pongamia is known to be drought tolerant, with findings here demonstrating that even young seedlings have this characteristic. Collectively, our findings indicate that Pongamia can grow quickly, reaching a biomass of 13-19 kg over 3-4 years in our studies, and sequester high quantities of carbon, at 2.9 to 4.0 t of carbon per ha (assuming a tree density of [~]450/ha), even when planted in suboptimal growing conditions on marginal, nutrient-poor lands.

plant biology↗

bioRxiv2025

APOE4 homozygotes have less sleep fragmentation in late adulthood

Background and ObjectivesAPOE4, a genetic risk factor for Alzheimers Disease (AD), is associated with reduced functional connectivity of brain regions that regulate sleep, which may predispose persons to AD via altering sleep architecture. However, little is known about differences in sleep architecture by APOE genotype. MethodsThis cross-sectional study examined the association between APOE genotype and sleep architecture among middle-aged and older adults, using polysomnography (Sleep Heart Health Study, N=3,132). APOE genotype included: APOE4 heterozygotes, APOE4 homozygotes, APOE2 carriers, and APOE3 homozygotes. Macro sleep architecture was quantified using the percentage of time spent in rapid eye movement sleep (%REM), N1 (N1%), N2 (%N2), N3 (%N3), and arousal index. Micro sleep architecture was quantified as odds ratio product (ORP; a continuous measure of sleep depth) for overall sleep and each sleep stage, and spindle characteristics (power, density, and frequency). Linear regression was used, adjusting for covariates. ResultsThe mean age was 67, 53 percent were female, 24% were APOE4 heterozygotes, 2% were APOE4 homozygotes, 14% were APOE2 carriers, and 60% were APOE3 homozygotes. Macro sleep architecture did not vary across genotypes. Compared with APOE3 homozygotes, APOE4 homozygotes exhibited fewer arousals with age ({beta}=-0.33 per hour/year, p=0.04), resulting in significantly fewer arousals at age 70+. ORP decreased in a dose-response pattern with the number of APOE4 alleles during overall sleep and across all sleep stages (ORPAPOE3/3=0.94, ORPAPOE3/4=0.91, ORPAPOE4/4=0.87), and these group difference widened with each year of age. Finally, there was a trend for lower spindle density and power in APOE4 homozygotes relative to APOE3 homozygotes (ps=0.06). ConclusionsArousal threshold increased in a dose-response manner with each APOE4 allele, as evidenced by the findings on ORP. The differences in ORP between APOE3 homozygotes and APOE4 carriers widened further with age, paralleling age-related declines in arousal index among APOE4 homozygotes. Despite these indications of elevated arousal thresholds that might suggest less sleep fragmentation in APOE4 carriers, APOE4 homozygotes exhibited poorer sleep micro architecture, including trends toward reduced sleep spindle activity. Taken together, reduced arousability in APOE4 carriers may reflect abnormalities in cortical activation that blunt arousal rather than an indicator of healthier sleep.

physiology↗

bioRxiv2025

A Guanine-quadruplex located on the negative strand of the hepatitis C virus facilitates efficient genomic RNA synthesis

Guanine-rich nucleic acids can form a unique secondary structure called Guanine-quadruplexes (G4s). These G4s are conserved across all domains of life and in viruses, where they can play important regulatory roles in the viral lifecycle. In flaviviruses, the (-) strand 3 untranslated region (UTR) is essential for initiating genomic RNA synthesis. Within the (-) strand 3 UTR of hepatitis C virus (HCV), a highly conserved G4 is located within stem-loop IIy (SLIIy), spanning nucleotides 110-131--a region that is essential for efficient replication. Using bioinformatics, we demonstrate that nucleotides 110-131 are highly conserved across HCV genotypes 1-7, and this region within SLIIy can likely adopt a more energetically favorable G4 conformation rather than the predicted hairpin. From biophysical and cell biology experiments, we show that stabilizing the predicted hairpin structure in SLIIy disrupts RNA synthesis, while successive guanine-to-adenine mutations within the G4 sequence impair G4 formation and hinder replication. Combining our findings with prior studies, we propose that the SLIIy G4 plays a crucial role in recruiting NS3 helicase, which is necessary for unwinding RNA structures and facilitating access for NS5B polymerase. The G4 may serve as a regulatory switch that modulates helicase binding and replication efficiency. We propose that in the absence of a stable G4, NS3 recruitment or unwinding is impaired, leading to inefficient RNA unwinding and reduced polymerase activity, ultimately hindering viral replication. These results reveal a previously unrecognized role for G4 structures in the HCV replication cycle, highlighting their importance in regulating (+) strand RNA synthesis. ImportanceHepatitis C virus is a positive-sense single-stranded RNA virus that relies on an intermediate negative strand to generate new genomic RNA. While the last 157 nucleotides of the negative-strand 3 untranslated region are known to be essential for replication, the underlying regulatory mechanisms have remained poorly understood. Our study demonstrates that a highly conserved guanine-rich sequence (107-131 nt) located on the negative strand SLIIy forms a guanine-quadruplex secondary structure that plays a pivotal role in orchestrating viral RNA synthesis. This G4 not only promotes efficient replication but likely acts as a molecular target to recruit the NS3 helicase, thereby allowing access for the NS5B polymerase. Disruption of the G4 structure and stabilization of the stem-loop severely impairs viral replication. These findings reveal an unrecognized layer of post-transcriptional regulation in the HCV lifecycle and establish G4 RNA structures as a critical element in HCV genome replication.

biophysics↗

bioRxiv2025

The Mycobacterium tuberculosis ESX effector promotes pyroptosis-dependent pathogenicity and dissemination

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=126 SRC="FIGDIR/small/653593v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@8cd5e5org.highwire.dtl.DTLVardef@2ec22forg.highwire.dtl.DTLVardef@1ceb773org.highwire.dtl.DTLVardef@6dfba4_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphic AbstractC_FLOATNO C_FIG Mycobacterium tuberculosis (Mtb) remains a major global health threat, partly due to the extensive cytotoxicity induced during infection. Although GSDMD-mediated excessive pyroptosis promotes pathogen dissemination and tissue damage in tuberculosis, the mechanism remains poorly understood. Here, we identify the effector EccB5, a component of the Mtb ESX-5 secretion system, as a key driver of pyroptosis and hyperinflammatory responses. EccB5 enhances Mtb virulence by inducing pyroptosis of macrophages, promoting bacterial dissemination and exacerbating lung pathology. Conditional knockdown EccB5 increases host cell viability. Mechanistically, EccB5 directly interacts with GSDMD, strengthens its association with caspase-1, and facilitates caspase-1-mediated cleavage of GSDMD both in vitro and in vivo. In summary, our findings uncover a precise mechanism by which Mtb modulates host responses and advances its pathogenicity.

microbiology↗

bioRxiv2025

GDF15 reprograms the microenvironment to drive the development of uveal melanoma liver metastases

Uveal melanoma (UM) results in fatal liver metastasis, yet little is known about the interactions between UM and host cells in the tumor microenvironment that promote this distinctive proclivity. Here, we used single cell (sc)-RNA-Seq analysis of UM-hepatic stellate cell (HSC) co-cultures to demonstrate that HSCs enriched for UM cell states that expressed genes implicated in cell survival, metabolic reprogramming and angiogenesis. A lead candidate driver of HSC reprogramming was the TGF-{beta} family member GDF15, which was associated with a metastatic UM phenotype. Silencing of BAP1 in UM cells led to increased GDF15 expression and accumulation of H3K27ac marks at the GDF15 promoter. Treatment of HSCs with GDF15 led to increased expression extracellular matrix proteins, inflammatory cytokines and angiogenic factors, including IL-8. Both exogenous GDF15, IL-8 and conditioned media from UM-HSC co-cultures increased endothelial cell network formation in vitro, an effect that was blocked by anti-GDF15 antibodies. In multiple models of metastatic UM, silencing of GDF15 inhibited the outgrowth of metastatic lesions, associated with reduced deposition of extracellular matrix and recruitment of endothelial cells. UM liver metastasis development is dependent upon GDF15-mediated remodeling of the liver microenvironment leading to an angiogenic response and matrix deposition that supports tumor growth.

cancer biology↗

bioRxiv2025

Evidence for balancing selection on loci associated with host use in the genome of a generalist plant parasite

Most research on host-parasite interactions has focused on tightly coevolved species pairs, yet many parasites are generalists that infect a wide range of host taxa. Generalist parasites are subject to highly variable selection pressures across different hosts and the footprints that this heterogeneous selection leaves in the genomes of generalists have been largely unexplored. To address this knowledge gap, we studied patterns of host-associated genetic diversity in the generalist plant parasite, the northern root knot nematode Meloidogyne hapla. By leveraging whole genome amplification, we generated whole genome libraries of individual root-knot nematodes collected from three different host plant species in a spatially replicated design. Despite finding no evidence of host-associated population structure, we did observe several hundred host-associated SNPs putatively under differential host selection. Genomic regions containing these host-associated SNPs were enriched for signals of balancing selection. Additionally, host-associated SNPs were enriched for functions involved in plant pathogenesis, specifically the modification of plant cell walls. Host-associated SNPs were also found in genes with known roles in dampening host plant immune responses to infection. Together, our results suggest a model of balancing selection in which heterogeneous selection by different host taxa maintains adaptive diversity at loci involved in pathogenesis.

evolutionary biology↗

bioRxiv2025

Distinct yet Overlapping Functions of VMP1 and TMEM41B in Modulating Hepatic Lipoprotein Secretion and Autophagy in MASH

BackgroundTransmembrane protein 41B (TMEM41B) and vacuolar membrane protein 1 (VMP1) are endoplasmic reticulum (ER) scramblases that shuttle phospholipids between the inner and outer leaflets of the ER membrane. Both TMEM41B and VMP1 also play critical roles in regulating hepatic lipoprotein secretion and autophagy. Despite these similarities, whether TMEM41B and VMP1 exhibit different roles in very low-density lipoprotein (VLDL) secretion and autophagy in the pathogenesis of metabolic-associated steatotic liver disease (MASLD) remains unclear. MethodsWe created liver- and hepatocyte-specific single knockout (KO) and double knockout (DKO) mice for Tmem41b and Vmp1, as well as overexpression knock-in (KI) mice with hepatic overexpression of TMEM41B, Tmem41b KO/Vmp1 KI, and Vmp1 KO/Tmem41b KI. We conducted lipidomic, metabolomic, biochemical, and functional studies in these mice, fed either a chow diet or a MASLD diet. ResultsTMEM41B protein levels were decreased in the livers of human subjects with MASLD. The loss of hepatic Tmem41b impaired VLDL secretion, resulting in steatosis, inflammation, and fibrosis. Vmp1 KO mice exhibited similar phenotypes to DKO mice, displaying a more severe defect in VLDL secretion and greater liver injury than Tmem41b KO mice. Lipidomic analysis revealed decreased levels of phosphatidylcholine and phosphatidylethanolamine, along with increased neutral lipids in both Tmem41b KO and Vmp1 KO mice; however, these changes were generally more pronounced in Vmp1 KO mice. VMP1 and TMEM41B localized at the mitochondrial-associated membrane (MAM), and a reduction in mitochondria-ER contact was observed in hepatocytes deficient in either VMP1 or TMEM41B. Ultrastructural electron microscopy analysis showed increased accumulation of "lipid droplet" in the ER membrane bilayer and ER lumen in both Vmp1 KO and Tmem41b KO hepatocytes, with greater ER luminal "lipid droplet" accumulation in Tmem41b KO hepatocytes. The loss of hepatic Vmp1 or Tmem41b led to elevated levels of LC3-II and p62, with significantly higher levels of both markers in Vmp1 KO and DKO mouse livers compared to Tmem41b KO mouse livers. Restoring Vmp1 in Tmem41b KO mice partially improved defective VLDL secretion; however, high expression levels of VMP1 did not correct the hepatic autophagy defect. In contrast, restoring Tmem41b in Vmp1 KO mice dose-dependently enhanced both defective VLDL secretion and autophagy. Importantly, overexpression of hepatic TMEM41B mitigated diet-induced MASLD in mice. ConclusionThe loss of hepatic Vmp1 or Tmem41b decreases hepatic MAM and phospholipid content, leading to decreased VLDL secretion and promoting MASLD. While VMP1 and TMEM41B have overlapping functions, VMP1 appears to play a more critical role in regulating VLDL secretion and autophagy in mouse livers than TMEM41B.

cell biology↗