Biology subjects

Huang, J.

Publications and source records attributed to Huang, J..

At least 199 records · Page 11Linked to original sources

bioRxiv2025

AIPAMPDS: an AI platform for antimicrobial peptide design and screening

Antimicrobial resistance (AMR) presents a pressing global health crisis, particularly in the post-pandemic era, underscoring the urgent need for innovative therapeutics. Antimicrobial peptides (AMPs) hold promises for combating multidrug-resistant infections but are hindered by challenges in clinical translation, including limited hemocompatibility and inefficient design processes. To overcome these obstacles, we developed AIPAMPDS, an AI-driven computational platform that integrates a deep generative model with multi-stage screening to design highly active, non-haemolytic AMPs. The platform features two key modules: (1) a Generative Pre-trained Transformer (GPT)-based framework for target-specific AMP generation, enabling rapid exploration of novel peptide sequences, and (2) a multi-neural network screening pipeline for dual evaluation of antimicrobial activity and toxicity. By systematically evaluating both AI-generated and natural candidate AMPs, we identified ten lead peptides for experimental validation, of which 90% exhibited negligible haemolytic activity while maintaining robust antimicrobial potency. AIPAMPDS bridges computational design and therapeutic application, significantly advancing AMP discovery and providing a powerful tool for next-generation antimicrobial development. The platform is freely accessible via a web server at https://amps.pianlab.cn/AIPAMPDS/. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=94 SRC="FIGDIR/small/644774v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@14e9a20org.highwire.dtl.DTLVardef@168e811org.highwire.dtl.DTLVardef@5bd9beorg.highwire.dtl.DTLVardef@344c34_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Compositional data modeling of high-dimensional single cell RNA-seq (CoDA-hd): its advantages over commonly used normalization approaches

Compositional data analysis (CoDA) is an emerging statistical framework and has been extended to microbiome, bulk RNA-seq, and cell type proportions in single-cell RNA-seq (scRNA-seq) (with 50-200 components). Here, we explore the high-dimensional application of CoDA (CoDA-hd) and its various log-ratio (LR) transformations to raw count matrices of scRNA-seq which has over 20,000 components (e.g., protein coding genes). scRNA-seq matrices are typically sparse and high-dimensional. Common approaches of normalization such as log-normalization may lead to suspicious findings as previously shown for trajectory inference. Although RNA-seq is compositional data by nature, the geometry of CoDA in high-dimensional simplex is not compatible with most downstream analyses of scRNA-seq which are based on Euclidean space. In this study, we explored if CoDA is adaptable with scRNA-seq in various downstream applications. Specifically, we attempted to study: (1) CoDA adaptability to scRNA-seq; (2) handling of zero data: prior-log-normalized, imputation or with specific count addition; (3) transformation to Euclidean space and compatibility with downstream analyses. Our results suggested that (1) the innovative count addition schemes (e.g., SGM) enable the application of CoDA to high dimensional sparse data (i.e., scRNA-seq); (2) log-normalized data could be transformed to CoDA LR transformation as an approximation; (3) CoDA LR transformations such as count-added centered-log-ratio (CLR) had some advantages in dimension reduction visualization, clustering, and trajectory inference in the tested real & simulated datasets. CLR provided more decent and separated clusters in dimension reductions, improved the Slingshot trajectory inference, and eliminated the suspicious trajectory that is probably caused by the dropouts. We therefore concluded that CoDA may be a preferred scale-free model to handle scRNA-seq data for these downstream applications. Additionally, an R package CoDAhd was developed for conducting CoDA LR transformations for high dimensional scRNA-seq data. The code for implementing CoDA-hd and some example datasets were placed at https://github.com/GO3295/CoDAhd.

bioinformatics↗

bioRxiv2025

ZBP1 Senses Splicing Aberration through Z-RNA to promote Cell Death

RNA splicing, a highly regulated process performed by the spliceosome, is crucial for eukaryotic gene expression and cellular function. Numerous cellular stresses including oncogenic insults dysregulate RNA splicing, often provoking inflammatory responses and cell death. However, the molecular signal produced by spliceosome aberration and how cell sensed and respond to it remain elusive. Here we show that spliceosome inhibition induces the widespread formation of unique, left-handed nucleic acids, Z-form nucleic acids (Z-NAs) from the nucleus. These Z-NAs were double-stranded RNA (dsRNA), rather than DNA-RNA hybrids, that were predominantly derived from transcripts of mis-spliced intronic RNA. Spliceosome inhibition induced the egress of these Z-RNA from the nucleus to the cytoplasm in an active manner. Sensing of the accumulation of Z-RNA in the cytosol by the host sensor ZBP1 triggered cell death, mainly for RIPK3-MLKL dependent necroptosis. Collectively, these findings delineate a previously uncharacterized mechanism in which Z-NA sensing by ZBP1 responds to global aberrations of RNA splicing to trigger inflammatory cell death.

cell biology↗

bioRxiv2025

Resilience and vulnerabilities of tumor cells under purine shortage stress

Purine metabolism is a promising therapeutic target in cancer; however how cancer cells respond to purine shortage,particularly their adaptation and vulnerabilities, remains unclear. Using the recently developed purine shortage-inducing prodrug DRP-104 and genetic approaches, we investigated these responses in prostate, lung and glioma cancer models. We demonstrate that when de novo purine biosynthesis is compromised, cancer cells employ microtubules to assemble purinosomes, multi-protein complexes of de novo purine biosynthesis enzymes that enhance purine biosynthesis efficiency. While this process enables tumor cells to adapt to purine shortage stress, it also renders them more susceptible to the microtubule-stabilizing chemotherapeutic drug Docetaxel. Furthermore, we show that although cancer cells primarily rely on de novo purine biosynthesis, they also exploit Methylthioadenosine Phosphorylase (MTAP)-mediated purine salvage as a crucial alternative source of purine supply, especially under purine shortage stress. In support of this finding, combining DRP-104 with an MTAP inhibitor significantly enhances tumor suppression in prostate cancer (PCa) models in vivo. Finally, despite the resilience of the purine supply machinery, purine shortage-stressed tumor cells exhibit increased DNA damage and activation of the cGAS-STING pathway, which may contribute to impaired immunoevasion and provide a molecular basis of the previously observed DRP-104-induced anti-tumor immunity. Together, these findings reveal purinosome assembly and purine salvage as key mechanisms of cancer cell adaptation and resilience to purine shortage while identifying microtubules, MTAP, and immunoevasion deficits as therapeutic vulnerabilities.

cancer biology↗

bioRxiv2025

DiCE: Differential Centrality-Ensemble Analysis Based on Gene Expression Profiles and Protein-Protein Interaction Network

Uncovering key genes that drive diseases and cancers is crucial for advancing understanding and developing targeted therapies. Traditional differential expression analysis often relies on arbitrary cutoffs, missing critical genes with subtle expression changes. Some methods incorporate protein-protein interactions (PPIs) but depend on prior disease knowledge. To address these challenges, we developed DiCE (Differential Centrality-Ensemble), a novel approach that combines differential expression with network centrality analysis, independent of prior disease annotations. DiCE identifies candidate genes, refines them with an information gain filter, and reconstructs a condition-specific weighted PPI network. Using centrality measures, DiCE ranks genes based on expression shifts and network influence. Validated on prostate cancer datasets, DiCE identified genes over-represented in key pathways and cancer fitness genes, significantly correlating with disease-free survival (DFS), despite DFS not being used in selection. DiCE offers a comprehensive, unbiased approach to identifying disease-associated genes, advancing biomarker discovery and therapeutic development. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/638654v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@1ce9b81org.highwire.dtl.DTLVardef@14352e1org.highwire.dtl.DTLVardef@f63caeorg.highwire.dtl.DTLVardef@a0086_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Convergent strigolactone perception via independent horizontal gene transfer of pre-adapted α/β-hydrolases

Strigolactones (SLs) are not only phytohormones that influence multiple aspects of plant growth and development, but also signaling molecules for interactions between plants and certain fungi or bacteria. In plants, the SL receptor is an /{beta}-hydrolase (ABH) encoded by the D14/KAI2 gene family, which is known to be derived from proteobacterial RsbQ through horizontal gene transfer (HGT). Here we show that the CpD14-like (CDL) gene family, which encodes another ABH exhibiting SL receptor functions in fungi, was acquired from actinobacteria via an independent HGT event. X-ray crystallographic experiments reveal that CDL and D14/KAI2/RsbQ (DKR) proteins have distinctive lid structures but maintain the same core /{beta} fold and catalytic triad. Biochemical assays further show that both actinobacterial CDL and proteobacterial RsbQ can recognize and hydrolyze SLs, suggesting that they are pre-adapted for SL perception. This work demonstrates that independent HGT of pre-adapted genes can trigger convergent evolution of key innovations and provides insights into how SLs are sensed by organisms across kingdoms.

evolutionary biology↗

bioRxiv2025

Hyper-responsiveness of cancer stem cells to microenvironmental cues controls metastasis and therapy response through YAP/TAZ/TEAD

Cancer stem cells (CSCs) are key drivers of metastasis and therapy resistance but have been challenging to visualize and study in situ. Using a fluorescent CSC reporter, we observed very different population dynamics for CSCs and nonCSCs during metastatic lung colonization in breast cancer models. CSC expansive self-renewal drives early lesion formation before switching to a maintenance mode of balanced self-renewal and differentiation, whereupon nonCSC proliferation takes over as the main driver of metastatic expansion. Mechanistic analyses showed that CSCs are hyper-responsive to microenvironmental cues such as cell crowding and nutrient availability, suggesting a novel role for CSCs as sensors and early responders to fluctuating local conditions in the tumor. Incoming signals converge on YAP/TAZ/TEAD, with heightened CSC sensitivity and response supported by elevated receptor expression and increased chromatin accessibility around enhancers with TEAD binding sites. Targeting inputs to the YAP/TAZ/TEAD node reversed chemotherapy-induced enrichment of CSCs in lung metastases. HighlightsO_LIDifferent population dynamics for breast cancer stem cells (CSCs) and their differentiated progeny in early metastatic colonization C_LIO_LICSCs are hyper-responsive to microenvironmental cues and serve as sensors of local conditions for the tumor C_LIO_LIMany microenvironmental inputs converge on YAP/TAZ to regulate self-renewal vs differentiation fate decisions in the CSC C_LIO_LITargeting YAP/TAZ input pathways blocks chemotherapy-induced enrichment of CSCs C_LI

cancer biology↗

bioRxiv2025

Lipocalins are versatile regulators of development and stress response in mosses

Major physiological and developmental innovations, such as stress tolerance strategies and three-dimensional growth, are key to land plant adaptation and evolution, but their genetic basis remains to be fully understood. Although lipocalins are known for their role to optimize adaptation and development in animals, they are heavily understudied in plants. Here we show that lipocalins in the moss Physcomitrium patens are versatile regulators of miscellaneous key physiological and developmental processes. Phylogenetics analyses revealed three major groups of plant lipocalins, including a newly identified group that is restricted to seedless plants. The temperature-induced lipocalin gene in P. patens (PpTIL) is functionally conserved with flowering plants in response to various abiotic stresses (e.g., heat, cold, salt, and dehydration). We demonstrate that PpTIL not only regulates protonemal development and the transition from two-dimensional to three-dimensional growth, but also affects many other processes such as lipid transport and metabolism, auxin biosynthesis and transport, and chlorophyll catabolism. These findings provide major insights into the role of lipocalins in land plant evolution.

evolutionary biology↗

bioRxiv2025

Comprehensive profiling of N6-methyladnosine (m6A) readouts reveals novel m6A readers that regulate human embryonic stem cell differentiation

N6-methyladenosine (m6A) methylation has emerged as a prevalent RNA modification that extensively impacts various physiological and pathological processes via various post-transcriptional readout effects in mammals. High-throughput methylome profiling has outlined the landscape of m6A modification sites, but their downstream readouts require comprehensive investigation. To this end, we systematically assessed the effects of m6A on mRNA half-life, translation efficiency, and alternative splicing across five cell lines (A549, HEK293T, HUVEC, JURKAT, and human embryonic stem cells (hESCs)) using actinomycin D-disrupted temporal transcriptome sequencing, ribosome sequencing, and ultra-high-depth transcriptome sequencing, respectively. Our analysis, coupled with the integration of public and re-profiled m6A methylome data, revealed high cell type specificity in m6A readouts where m6A level alone is insufficient to predict m6A readouts. Machine learning models focused on the RNA binding protein (RBP) binding context of m6A sites demonstrated substantial predictive ability of m6A readouts while prioritizing putative m6A readers from their informative RBP features. Four novel m6A readers (DDX6, FUBP3, FXR2, and L1TD1) were identified and validated through m6A RNA pull-down assays and transcriptome-wide RBP binding site mapping. Notably, FUBP3, FXR2 and L1TD1 were found to regulate hESC differentiation without impairing self-renewal, underscoring their critical roles in stem cell biology. Together, this study bridges the gap in understanding m6A functional readouts and lays the groundwork for future research on m6A-mediated stem cell fate decisions.

genomics↗

bioRxiv2025

Chimeric Collagen-like Proteins with Tunable Structural Heterogeneity for Precise Control and Targeted Doxorubicin Delivery

Protein-based drug delivery systems offer advantages such as genetic tunability, structural homogeneity, and high biocompatibility. However, precisely controlling the assembly of protein nanoparticles and establishing correlations between their macroscopic properties and molecular architecture remain significant challenges. Here, inspired by chimeric collagen-like proteins, we rationally designed a series of de novo chimeric proteins comprising an N-terminal globular domain fused to a linear collagen-like domain. These structurally heterogeneous proteins were conjugated with doxorubicin (DOX) and homogenized to form chimeric collagen-like protein-DOX nanocomplexes (CDCNs). By fine-tuning the protein structure and domain organization, we achieved precise control over CDCN architecture and drug release kinetics. Furthermore, to enhance tumor specificity, triple-negative breast cancer (TNBC)- and glioblastoma (GBM)-targeting peptides were genetically fused to the chimeric proteins, respectively. In both in vitro and in vivo models of TNBC and GBM, CDCNs facilitated selective tumor accumulation, enhanced cellular uptake, and promoted apoptosis while minimizing off-target toxicity. This work establishes a strategy for designing protein-based nanoplatforms with programmable structures and tunable functionalities, offering promising potential for precisely controlling of protein-based delivery system.

bioengineering↗

bioRxiv2025

Two-Stage CD8+ CAR T-Cell Differentiation in Patients with Large B-Cell Lymphoma

Chimeric antigen receptor (CAR) T-cell therapy has expanded therapeutic options for patients with diffuse large B-cell lymphoma (DLBCL). However, progress in improving clinical outcomes has been limited by an incomplete understanding of CAR T-cell differentiation in patients. To comprehensively investigate CAR T-cell differentiation in vivo, we performed single-cell, multi-modal, and longitudinal analyses of CD28-costimulated CAR T cells from infusion product and peripheral blood (day 8-28) of patients with DLBCL who were successfully treated with axicabtagene ciloleucel. Here, we show that CD8+ CAR T cells undergo two distinct waves of clonal expansion. The first wave is dominated by CAR T cells with an exhausted-like effector memory phenotype during the peak expansion period (day 8-14). The second wave is dominated by CAR T cells with a terminal effector phenotype during the post-peak persistence period (day 21-28). Importantly, the two waves have distinct ontogeny and are biologically uncoupled. Furthermore, lineage tracing analysis via each CAR T cells endogenous TCR clonotype demonstrates that the two waves originate from different effector precursors in the infusion product. Precursors of the first wave exhibit more effector-like signatures, whereas precursors of the second wave exhibit more stem-like signatures. These findings suggest that pre-infusion heterogeneity mediates the two waves of in vivo clonal expansion. Our findings provide evidence against the intuitive idea that the post-peak contraction in CAR abundance is solely apoptosis or extravasation of short-lived CAR T cells from peak expansion. Rather, our findings demonstrate that CAR T-cell expansion and persistence are mediated by clonally, phenotypically, and ontogenically distinct CAR T-cell populations that serve complementary clinical purposes.

immunology↗

bioRxiv2025

NtLLG4-mediated unconventional polar exocytosis of NtPPME1 coordinates cell wall rigidity and membrane dynamics to control pollen tube integrity

Plant fertilization relies on controlled pollen tube growth that integrates membrane dynamics and cell wall expansion. We previously identified an unconventional exocytic pathway wherein Golgi-derived secretory vesicles (GDSVs) bypass the trans-Golgi network to deliver Nicotiana tabacum pectin methylesterase 1 (NtPPME1), thereby modulating cell wall rigidity. However, the mechanisms linking this patwhay with membrane dynamics and signaling remain elusive. Here, we used cryo-FIB-SEM and 3D tomography to identified GDSVs as a distinct vesicle population at the pollen tube tip. We further demonstrated that tobacco LORELEI-like-GPI-anchored protein 4 (NtLLG4), a key signaling molecule controlling membrane dynamics and integrity, functions as a receptor for NtPPME1, regulating its polar exocytosis via GDSVs to control cell wall stiffness. Furthermore, we identified trafficking signals which direct the unconventional exocytosis of NtPPME1 across intracellular organelles. Our findings reveal a crucial mechanism coupling cell wall rigidity with membrane signaling to control pollen tube growth and integrity during fertilization. TeaserWe reveal a mechanism coupling cell wall rigidity with member signaling to control pollen tube growth and integrity during plant fertilization.

plant biology↗

bioRxiv2025

Unraveling the Neurocognitive Mechanisms of Delayed Punishment in Second- and Third-Party Contexts

Second-party punishment (SPP) and third-party punishment (TPP) are essential in regulating social behavior and maintaining social norms, yet the effectiveness may wane when punishment is delayed. Currently, the decision-making and neural mechanisms underlying SPP and TPP under temporal delays remain largely unexplored. This study investigated both SPP and TPP punishment decisions with time-delay (immediate vs. delayed) using a hypothetical criminal scenario assessment task with fMRI. Results showed increased activity in the bilateral precuneus and left temporoparietal junction (TPJ) and stronger left TPJ-dorsolateral prefrontal cortex (dlPFC) connectivity in SPP compared to TPP. Interestingly, third-parties imposed more severe punishment in delayed conditions than in immediate ones, accompanied by enhanced neural activity in dorsomedial prefrontal cortex (dmPFC), left TPJ, ventrolateral prefrontal cortex (vlPFC), ventromedial prefrontal cortex (vmPFC), and caudate nucleus. This delay effect was evident only in misdemeanor and felony cases, with no differences in capital offenses, suggesting a moderating effect of criminal severity. In contrast, SPP showed no significant changes in punishment or neural response across immediate and delayed conditions. Multivariate pattern analysis further indicated that dmPFC, TPJ, vlPFC, vmPFC and caudate function together to encode punishment severity in TPP contexts, a pattern not observed in SPP. These findings underscore distinct decision-making mechanisms between SPP and TPP under temporal delays, with implications for understanding justice-related processing in the human brain under time constraints. Significance statementWithin the justice system, punishing norm violators is often hindered when they flee, resulting in delayed punishments. However, little is known about how second-party (SPP) and third-party (TPP) implement punishment with time delays. The current study investigated this using real criminal scenarios with time delays under fMRI scanning. We found that third parties imposed more severe punishment on defendants in delayed conditions than in immediate ones, reflected by enhanced neural activity in the dorsomedial prefrontal cortex, left temporoparietal junction, ventrolateral prefrontal cortex, ventromedial prefrontal cortex, and caudate nucleus. In contrast, no significant differences were observed in second-party punishment. We also found a moderating effect of criminal severity: the difference between immediate and delayed sanctions by third-parties was present in misdemeanor and felony cases but not in capital offenses. These results shed light on the decision-making mechanisms of SPP and TPP in contexts involving temporal delay, which may have deep impact in different disciplines, e.g., social science, cognitive neuroscience and legal system.

neuroscience↗

bioRxiv2025

Structural and Functional Insights into GGCX-FIX Interaction: Implications for Vitamin K-Dependent Bleeding Disorders

Gamma-carboxylation, catalyzed by {gamma}-glutamyl carboxylase (GGCX), is a critical post-translational modification essential for the biological activity of vitamin K-dependent proteins (VKDPs). Mutations in GGCX, depending on their specific location, result in vitamin K-dependent coagulation factor deficiency type 1 (VKCFD1), which encompasses a broad spectrum of clinical manifestations ranging from mild to severe, including bleeding disorders, osteoporosis, and vascular calcification. The limited knowledge of GGCXs structure and functional regions hinders our understanding of the consequences of GGCX mutations and the treatment for VKCFD1. This study aimed to identify key functional regions of GGCX and their interactions with VKDPs to better elucidate the molecular mechanisms underlying these diverse clinical symptoms. Using AlphaFold 3 and molecular dynamics simulations, we developed a complex binding model of GGCX, FIX, and reduced vitamin K, which revealed critical regions and residues involved in their interaction. Site-directed mutagenesis and cell-based assays further validated the model, confirming that multisite and regional cooperative binding of FIX to GGCX plays a key role in modulating {gamma}-carboxylation efficiency. Additionally, novel residues (I296, M303, M401, M402) were identified as essential for GGCXs dual enzymatic activities: carboxylation and vitamin K epoxidation. We further demonstrated that the spatial proximity of these active sites supports the hypothesis that GGCXs carboxylation and vitamin K epoxidation centers are interconnected, ensuring the efficient coupling of these processes. Our GGCX-FIX binding and carboxylation model aligns with known pathogenic GGCX mutations, providing valuable insights into the molecular basis of coagulation disorders caused by GGCX mutants.

pathology↗

bioRxiv2025

Stem Cell-Parenchymal Fusion: Communication and Gene Regulation

Cell fusion, a natural process occurring between similar or dissimilar cell types, often confers new or enhanced functionality, yet its impact on cellular communication and gene regulation remains poorly understood. Here, we used recent analytical frameworks to investigate accidental cell fusion between murine cardiomyocytes (mHL1) and murine mesenchymal stromal/stem cells (mMSC) leveraging previously published single-cell RNA sequencing data. After fusion, we observed a biased distribution of gene expression and acquired phenotypes in fused hybrids. Trajectory inference showed that hybrids with a more mMSC-like transcriptome diverged more significantly from parental cells relative to hybrids with a more mHL1-like transcriptome. We also observed dynamic changes in cell-cell communication, with early (Day 1) downregulation of Wnt signaling and Melanogenesis evolving into the upregulation of pathways like Endocrine resistance and Focal adhesion by Day 3. Notably, ECM (extracellular matrix)-receptor interactions were largely consistent whether annotation or unsupervised Clustree methods were used. Furthermore, our analysis indicated the emergence of various cancer-associated signaling mechanisms. Our findings highlight the remarkable plasticity of cellular identity following fusion and lay the groundwork for future research into the precise molecular mechanisms driving these transformations and the potential of cell fusion for generating novel cell types.

systems biology↗

bioRxiv2025

Homeodomain protein Sxi1α regulates cell-cell fusion during distinct sexual reproduction modes in Cryptococcus deneoformans

Sex-specific homeodomain (HD) proteins are key regulators of cell identity and sexual development in fungi, typically functioning as heterodimers to govern transcription. In the human fungal pathogens Cryptococcus neoformans and Cryptococcus deneoformans, the HD proteins Sxi1 and Sxi2a (Sex-inducer 1 and 2a) have been characterized as interacting components that play critical roles in sexual development during x a sexual reproduction. cells are the predominant mating type in natural populations of Cryptococcus, and unisexual (same-sex) mating can also occur in certain genetic backgrounds. The roles of Sxi1 and Sxi2a in unisexual reproduction are not fully understood. To elucidate the functions of Sxi1 and Sxi2a, we first applied AlphaFold3 prediction, which identified potential heterodimeric and homodimeric complexes. Formation of a Sxi2a homodimer was then experimentally validated through yeast two-hybrid assays. We subsequently deleted SXI1 and SXI2a in the hyper-filamentous self-fertile C. deneoformans strains XL280 and XL280a. Disruption of these genes did not result in noticeable defects in vegetative growth, virulence-associated traits, colony morphology, sporulation, or competitive fitness during unisexual crosses. Interestingly, both bilateral (mutant x mutant) and unilateral (mutant x wildtype) crosses involving the sxi1{Delta} mutant significantly increased - cell fusion efficiency, suggesting a previously unrecognized inhibitory role for Sxi1 in regulating same-sex cell fusion. Consistently, genes encoding mating pheromones and the -pheromone receptor Ste3 were upregulated in the sxi1{Delta} fusion assays. Transcriptomic analysis of sxi1{Delta} and sxi2a{Delta} mutants led to the identification of unique subsets of genes negatively regulated by each transcription factor during unisexual reproduction. Additionally, x a crosses between null mutants of sxi1{Delta} and sxi2a{Delta} revealed differential regulation of mating-type (MAT) loci genes dependent only on Sxi1 or Sxi2a. Together, our findings reveal a novel role for Sxi1 in governing cell fusion and demonstrate that Sxi1 and Sxi2a have distinct transcriptional control during unisexual and x a sexual reproduction, potentially exerting opposing regulation of sex-specific MAT genes. Author SummaryAnalogous to processes observed in many eukaryotic organisms, fungal sexual reproduction reshuffles genetic material, producing novel genetic variants with increased fitness and/or adaptive potential. The human fungal pathogen Cryptococcus deneoformans is widespread in the environment and can cause severe infections in immunocompromised individuals. Sexual reproduction in this species occurs between cells of either opposite mating types ( x a mating) or the same mating type ( x and a x a unisexual reproduction). The mating type-specific homeodomain transcription factors Sxi1 and Sxi2a are known to form a complex that broadly controls sexual development following and a cell fusion. Here, we find that Sxi1 inhibits cell-cell fusion during unisexual reproduction, while its partner, Sxi2a, shows only limited inhibition in certain genetic contexts. Further, we show that Sxi1 and Sxi2a regulate transcription of distinct gene sets during both x a and unisexual reproduction, including many within the mating-type (MAT) loci, often in opposing manners. Together, these findings reveal that Sxi1 and Sxi2a function both cooperatively and independently, thereby allowing for a more flexible and finely-tuned mating control system than previously recognized, providing insight into how this organism generates genetic variation and adapts to changing environments.

microbiology↗

bioRxiv2025

Complex Human Hair Bearing Skin Organoids as Model for Herpes Simplex Virus 1 Infection in the Skin

Epithelial surfaces are the initial site of Herpes Simplex Virus 1 (HSV-1) infection. After latency establishment in the peripheral nervous system, reactivation can lead to skin pathologies. To reflect the complexity of skin we used complex human hair bearing skin organoids as a new model system for HSV-1 infection. We analyzed spread of the infection through the three-dimensional tissue and the cell type specific host response in a spatial temporal manner. Single-cell-resolution spatial transcriptomics and imaging shows a restricted viral infection in keratinocytes of the epidermis and hair follicles. The host factor IER3 is upregulated in papillary fibroblasts of the dermis as the infection progresses through the tissue. In keratinocytes of the stratum basale, we find IRF3 upregulated in infected but also virus negative cells, suggesting a signaling mechanism to initiate antiviral responses. Furthermore, we find a cell type specific inflammatory response with an upregulation of TNFSF9 in dermal fibroblasts and TNF in keratinocytes. TNF target genes are induced in neighboring uninfected cells suggesting the induction of a localized inflammatory reaction to suppress viral infection. In summary, our findings position human hair-bearing skin organoids as a highly physiological model system for studying HSV-1 pathogenesis and reveal spatially organized host responses.

microbiology↗

bioRxiv2025

Age-Dependent Patterns of Gut Bifidobacterium Microbiome Diversity across child Cohorts in a Multi-Ethnic population, Western China

High-throughput sequencing has provided unprecedented opportunities to elucidate the role of human gut microbiota. However, current studies have primarily focused on infant and adult cohorts, leaving biomarkers associated with the development of transitional children unclear, particularly in revealing geographic, multi-ethnic group underrepresented. Here, we analyzed fecal samples from 776 infants and children aged 0 to 15 years with amplicon sequencing of the 16S rRNA and groEL gene, representing multi-ethnic groups from different residential locations in the Yining region of Xinjiang. First, we systematically characterized the gut microbiota Spectrum of children in the Ili region. As age increases, the diversity and functional composition of the microbiota become richer and more stable, with Bifidobacterium consistently dominating the community. Utilizing Bayesian regression models, we identified a cluster of microbiota strongly associated with growth and development in children, which phylogenetically corresponds to Bifidobacterium breve. Finally, we included another similar study in the species-level of Bifidobacterium, and metrics of the age-related effect sizes showed heterogeneity across ethnic groups (Uyghur vs Han: {beta}Deming = 4.40 and Kazakh vs Han: {beta}Deming = 4.33). In conclusion, this study provides novel insights into the developmental trajectories of the gut microbiome in children and its relationship with health. IMPORTANCEThe intestinal microbial markers associated with the development of geographically diverse, multi-ethnic children, particularly those represented by Bifidobacterium species, have been elucidated in this study. Through the application of Bayesian regression models, we identified a microbiome profile closely linked to childhood growth and development, specifically phylogenetically aligned with Bifidobacterium breve. When incorporating a comparable study conducted at the species level of another Bifidobacterium, it was observed that age-related effect sizes varied among different ethnic groups, offering novel insights into the developmental trajectory of childrens gut microbiota and its implications for health.

microbiology↗