Biology subjects

Xie, F.

Publications and source records attributed to Xie, F..

At least 19 recordsLinked to original sources

bioRxiv2026

PC1-guided transcriptomic stratification reveals hepatic transcriptional heterogeneity and defines a myeloid-associated 20-gene signature

Hepatic lipid-associated inflammation contributes to metabolic liver disease and cardiometabolic complications. Treatment-based transcriptomic comparisons can obscure inter-individual heterogeneity when animals exposed to the same experimental condition show divergent molecular responses. In the public hyperlipidemic liver transcriptomic dataset GSE338111, conventional sex-adjusted comparison of Amlexanox versus DMSO identified only 21 differentially expressed genes at FDR < 0.05 and |log2FC| [≥] 1, and submission of this DEG set to Metascape yielded no GO Biological Process enrichment result. We therefore applied treatment-independent, PC1-guided transcriptomic stratification based on the 500 most variable genes. This analysis resolved three PC1-derived groups and enabled derivation of a myeloid-associated 20-gene signature from the G2-versus-G1 contrast. Independent bulk-transcriptomic cohorts supported responsiveness of the signature to dietary challenge and pharmacologic intervention, while single-cell analysis localized its expression predominantly to hepatic myeloid populations. Human cis-eQTL Mendelian randomization and colocalization further identified TAGLN2 as the signature gene with the strongest genetic support for coronary heart disease. Together, these findings show that PC1-guided stratification can improve resolution of heterogeneous hepatic transcriptional responses and provide a cross-cohort molecular signature for subsequent mechanistic and translational evaluation.

genomics↗

bioRxiv2026

Dormant Bacteria's Fatal Attraction to RNA Bacteriophages

Phages are the most abundant biological entities on Earth, yet RNA phages are strikingly scarce compared to their DNA counterparts--a long-standing mystery in phage biology. Here, we use dsRNA phage phiYY as a model to demonstrate that while RNA phages efficiently infect growing bacteria, they are gradually eliminated inside dormant bacteria through weak and time-dependent ROS-mediated damage. The RNA phages decline over days rather than through immediate clearance inside dormant bacteria. Accordingly, scavenging ROS with mannitol or overexpressing ROS degradation enzymes AhpB/TrxB2 rescues RNA phages from elimination. Crucially, because the underlying ROS-mediated RNA damage is minimal, RNA phage survival hinges on genomic redundancy. In single-phage infections, the lone RNA genome is highly vulnerable to cumulative damage and is eventually inactivated. In contrast, during co-infection by multiple RNA phages, the presence of multiple genome copies provides functional redundancy, thereby allowing a fraction of RNA phages to survive inside dormant bacteria. Given that most environmental bacteria are dormant and subject to heterogeneous phage infection, this copy number-dependent vulnerability offers a possible explanation for the scarcity, but not perish, of RNA phages in nature.

microbiology↗

bioRxiv2026

CIPHER: An end-to-end framework for designing optimized aggregated spatial transcriptomics experiments

MotivationMost imaging-based spatial transcriptomics methods measure individual genes, which limits scalability and typically requires integration with scRNA-seq to recover full cellular states. Recent approaches such as CISI, FISHnCHIPs, and ATLAS address this limitation by measuring aggregate transcriptional signatures, where multiple genes are pooled into each channel to increase throughput. While aggregate measurements improve scalability, they shift the problem from gene selection to feature design. For effective integration with scRNA-seq, these signatures must be not only discriminative in transcriptional space but also straightforward to measure, with balanced signal, sufficient dynamic range, and robustness to experimental noise. By optimizing decoding accuracy in isolation, existing methods leave substantial performance on the table. ResultsWe present CIPHER (Cell Identity Projection using Hybridization Encoding Rules), a neural-network framework that jointly optimizes the experimental encoding matrix, i.e., the way that genes are aggregated to signatures, and the downstream cell embedding. CIPHER integrates the physical limits of imaging assays directly into its loss function, shaping the latent space to maximize discriminability while maintaining robustness to measurement noise and signal constraints. Using a large-scale mouse brain scRNA-seq reference, we show that CIPHER-designed encodings yield latent spaces with improved cell-type separability, uniform signal utilization, and greater resilience to hybridization variability, resulting in higher decoding accuracy from both simulated and experimental data. ConclusionCIPHER formulates aggregate signature design as a joint optimization problem over decoding accuracy and experimental measurability. This enables systematic, scRNA-seq-aligned feature design for scalable spatial transcriptomics based on aggregate measurements. AvailabilityCode and documentation are available at https://github.com/wollmanlab/Design/. Author summarySpatial transcriptomics reveals how cells are organized within tissues by mapping where genes are expressed. To achieve both scale and resolution, many approaches now combine spatial imaging with single-cell RNA-seq references to reconstruct complete transcriptomes in situ. New methods such as CISI, FISHnCHIPs, and ATLAS accelerate this process by measuring combinations of co-expressed genes rather than each gene individually. These aggregate measurements simplify experiments but introduce a new challenge: deciding which genes to combine so that the resulting features are both experimentally reliable and computationally informative for integration with scRNA-seq data. We developed CIPHER, a computational framework that learns how to design and decode these aggregate measurements optimally. By integrating experimental constraints and decoding accuracy into a unified neural-network model, CIPHER provides a principled approach for designing signature-based spatial transcriptomics experiments that enable efficient and accurate reconstruction of cellular transcriptomes.

bioinformatics↗

bioRxiv2026

Latent Regulatory Programs Generate Synthetic T Cell States with Enhanced Therapeutic Potential

Transcription factors (TFs) govern cell fate through coordinated gene-regulatory networks, yet the full potential of these networks to generate non-native, therapeutically advantageous cell states in vivo remains largely unexplored. We hypothesized that systematic gain-of-function (GOF) overexpression of TFs in CD8 T cells, central mediators of immune protection, could reveal latent, or "hidden," regulatory programs capable of generating synthetic T cell states with therapeutic utility. To test this, we developed single-cell GOF sequencing (scGOF-seq), a multiplexed platform for unbiased, in vivo mapping of GOF effects on T cell fate in immunocompetent mouse models of infection and cancer. scGOF-seq uncovered unexpected regulators of T cell differentiation and accumulation, including SOX2, OCT4, and GATA2, which are normally silenced during T cell differentiation. Notably, outside its native regulatory context, supraphysiologic cMyc GOF reprogrammed CD8 T cells into a synthetic stem-effector hybrid state, enabling >5,000-fold antigen-dependent expansion and antitumor activity, contrasting sharply with its native function in driving terminal differentiation. scGOF-seq further identified TF modules that cooperate with cMyc GOF to promote robust CD8 T cell responses in solid tumors. Together, these findings establish GOF perturbation as a powerful strategy for revealing latent immune regulatory programs and engineering synthetic immune states with therapeutic potential. One-Sentence SummaryIn vivo single-cell gain-of-function screening reveals latent transcriptional programs that can reprogram T cells into highly functional synthetic states.

immunology↗

bioRxiv2025

Gasdermin E couples mitochondrial damage to pyroptotic neurodegeneration in Parkinson disease

Parkinsons disease (PD) features progressive loss of nigrostriatal dopamine neurons, but how mitochondrial damage engages programmed cell death pathways remains unresolved. Here, we identify gasdermin E (GSDME), the caspase-3-activated executor of pyroptosis, as a critical mediator of neurodegeneration in toxin-based PD models. In primary neurons and SH-SY5Y cells, the mitochondrial complex I inhibitor MPTP/MPP triggered caspase-3 activation, GSDME cleavage, and lytic membrane rupture. Genetic silencing of Gsdme or its transcriptional regulator SP1 reduced neuronal death. In vivo, Gsdme deficiency preserved substantia nigra pars compacta dopaminergic neurons, improved motor performance, and mitigated anxiety- and depression-like behaviors after MPTP administration. Loss of Gsdme also dampened microglial and astrocytic activation and lowered proinflammatory cytokines in striatum and substantia nigra. Mechanistically, cleaved GSDME localized to mitochondria, disrupted membrane potential, increased reactive oxygen species, and precipitated organelle injury, thereby coupling mitochondrial dysfunction to pyroptotic cell death. These findings identify GSDME-mediated pyroptosis as a mechanistic link between mitochondrial toxicity and neuroinflammation in PD and nominate GSDME as a therapeutic entry point to slow disease progression. SignificanceHow mitochondrial injury kills dopamine neurons in Parkinsons disease is a central unresolved question. We show that the pyroptosis executor GSDME is required for neurodegeneration and neuroinflammation in MPTP models, mechanistically linking caspase-3 activation and mitochondrial damage to lytic cell death. Targeting GSDME may provide a strategy to protect vulnerable neurons in PD. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=191 SRC="FIGDIR/small/694264v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@15364e7org.highwire.dtl.DTLVardef@57b65org.highwire.dtl.DTLVardef@1e85108org.highwire.dtl.DTLVardef@1200d34_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsGSDME drives PD-linked neuronal loss. Silencing GSDME (or its transcriptional factor Sp1) ameliorates MPTP-triggered neuronal cell death. Genetic ablation of Gsdme is protective in the MPTP-induced PD mouse model. Gsdme-/-mice retain nigrostriatal DA neurons, show improved motor and affective behaviors, and exhibit reduced microglial and astrocytic activation and cytokines release upon MPTP treatment. Cleaved GSDME accumulates on mitochondria, collapses {Delta}{Psi}m, raises ROS, and links mitochondrial toxicity to pyroptosis, thus positioning GSDME as a tractable target to slow PD progression.

neuroscience↗

bioRxiv2025

Streptavidins play a multifunctional role within a biotin-pathway antibiotic network encoded in a biosynthetic supercluster

Streptavidin, the well-known biotin-binding protein extensively used in biotechnology, is naturally co-produced in Streptomyces avidinii alongside stravidins--inhibitors of biotin biosynthesis. Here, we uncover and activate a conserved genomic region flanked by two streptavidin genes, revealing multiple biosynthetic gene clusters that produce diverse biotin-related metabolites, including stravidins, acidomycin, -methyl-KAPA, -methyldesthiobiotin, and the novel non-proteinogenic amino acid ANDA. We show -methyldesthiobiotin arises from -methyl-KAPA, illustrating how methylated analogues interfere with distinct steps of biotin biosynthesis. Contrary to the view that streptavidin functions solely by sequestering biotin, our biochemical, structural, and bioactivity analyses demonstrate that it also binds acidomycin, neutralizing its antibacterial activity to protect the producer while likely facilitating compound secretion. The crystal structure of the streptavidin-acidomycin complex reveals the molecular basis for this dual functionality. Our findings establish a multifunctional streptavidin that integrates biotin sequestration and self-resistance to balance an antibiotic network targeting the biotin pathway in microbial competition.

microbiology↗

bioRxiv2025

Accelerating scRNA-seq analysis: automated cell type annotation using representation learning and vector search

Cell type annotation in single-cell RNA sequencing (scRNA-seq) experiments is the fundamental step of assigning cell types to individual cells or clusters of cells based on their gene expression profiles. This process is crucial for developing biological insights from scRNA-seq experiments. We present a service that automates cell type annotation for 10x Genomics single-cell gene expression samples, enabling researchers to rapidly and accurately categorize cells within a sample. This service operates on the basis of reverse search: it compares each cells gene expression profile against the Chan Zuckerberg CELL by GENE (CZ CELLxGENE) Census, a comprehensive repository of published scRNA-seq datasets enriched with community-annotated cell types, and yields cell type annotations through summarizing the labels associated with similar cells. The annotation algorithm employed in this service avoids reliance on predefined marker genes or tissue-specific references, providing both fine-grained and coarse annotations. These initial annotations can be further refined by investigators to suit their specific research needs.

bioinformatics↗

bioRxiv2025

Establishing a continuum of cell types in the visual cortex

The mammalian cerebral cortex is composed of neurons whose properties vary in a continuous fashion rather than falling into discrete cell types. In the mouse visual cortex, excitatory neurons in layer 2 and 3 (L2/3) form such a continuum along cortical depth, patterned by the graded expression of hundreds of genes. Here we sought to understand how this continuum develops and contributes to cortical wiring. Using single-nucleus multiomics (RNA- and ATAC-Seq) and spatial transcriptomics, we show that the L2/3 continuum is established in two phases. During the first postnatal week, a genetically hardwired program establishes a primitive continuum of cell identities spanning the depth of L2/3. The second program, promoted by visual experience, is later superimposed upon the preexisting continuum. This second phase is driven by activity-regulated transcription factors that drive the L2/3 depth-dependent expression of genes linked to synaptic function and plasticity. We show that neurons at different positions along the L2/3 continuum project preferentially to distinct higher visual areas and that visual deprivation disrupts targeting to some higher visual areas while sparing others. Thus, cortical continua emerge through a stepwise process in which genetic programs and sensory experience specify neuronal identity and sculpt intracortical wiring specificity.

neuroscience↗

bioRxiv2025

FOXJ1 mediates taxane resistance through regulation of microtubule dynamics

Docetaxel is the first-line chemotherapy for metastatic castration-resistant prostate cancer (PC), but clinically meaningful mechanisms of resistance remain to be established. We generated an in vivo model of docetaxel resistance using castration-resistant patient-derived xenografts and found increased expression of genes that drive development of multiciliated cells, including FOXJ1 and its effector genes, many of which regulate ciliary microtubules (MTs). Mechanistically, FOXJ1 overexpression conferred docetaxel resistance in vitro and in vivo, which was associated with decreased docetaxel-mediated MT bundling. Overexpression of a MT-associated FOXJ1-regulated gene (TPPP3) had similar effects. Conversely, FOXJ1 knockdown impaired basal MT function, enhanced taxane binding to MTs, and increased docetaxel sensitivity. These results establish mechanistic causality between the FOXJ1 signaling axis, MT biology, and taxane resistance. Clinically, FOXJ1 gene amplification was increased in taxane-treated PC patients. Moreover, in the CHAARTED clinical trial of docetaxel combined with androgen deprivation for metastatic PC, higher baseline FOXJ1 was predictive of decreased survival in PC patients treated with docetaxel, further supporting clinical relevance. Together these findings identify a previously unrecognized clinically impactful mechanism of taxane resistance whose exploitation could stratify patients that will not benefit from taxane treatment. Statement of SignificanceThe early use of docetaxel in combination with androgen receptor targeted therapy can enhance survival in a subset of PC. This study indicates that increased FOXJ1 is predictive of tumors that will not benefit from docetaxel, and that agents targeting FOXJ1 or downstream effectors may enhance taxane responses and improve patient outcomes.

cancer biology↗

bioRxiv2025

Population-specific brain charts reveal Chinese-Western differences in neurodevelopmental trajectories

Human brain charts provide unprecedented opportunities for decoding neurodevelopmental milestones and establishing clinical benchmarks for precision brain medicine 1-7. However, current lifespan brain charts are primarily derived from European and North American cohorts, with Asian populations severely underrepresented. Here, we present the first population-specific brain charts for China, developed through the Chinese Lifespan Brain Mapping Consortium (Phase I) using neuroimaging data from 43,037 participants (aged 0-100 years) across 384 sites nationwide. We establish the lifespan normative trajectories for 296 structural brain phenotypes, encompassing global, subcortical, and cortical measures. Cross-population comparisons with Western brain charts (based on data from 56,339 participants aged 0-100 years) reveal distinct neurodevelopmental patterns in the Chinese population, including prolonged cortical and subcortical maturation, accelerated cerebellar growth, and earlier development of sensorimotor regions relative to paralimbic regions. Crucially, these Chinese-specific charts outperform Western-derived models in predicting healthy brain phenotypes and detecting pathological deviations in Chinese clinical cohorts. These findings highlight the urgent need for diverse, population-representative brain charts to advance equitable precision neuroscience and improve clinical validity across populations.

neuroscience↗

bioRxiv2025

Hydrogen-Induced Calcium Influx via the TRPC4-TRPC4AP Axis

BackgroundCalcium ions (Ca{superscript 2}) serve as universal intracellular messengers regulating diverse physiological processes, while dysregulated Ca{superscript 2} homeostasis triggers cytotoxicity. Molecular hydrogen (H2) exhibits protective effects against oxidative stress-related pathologies, but its mechanism of action remains incompletely understood. Transient receptor potential canonical 4 (TRPC4) channels and their associated protein TRPC4AP are critical mediators of Ca{superscript 2} influx ( [Ca{superscript 2}]i), yet their role in H2-mediated calcium signaling is unexplored. This study investigates the molecular mechanism by which H2 modulates Ca{superscript 2} dynamics through the TRPC4-TRPC4AP axis, aiming to establish its therapeutic potential for calcium-related disorders. MethodsThe study employed heterogeneous cellular models (e.g., mesenchymal stem cells, neurons, fibroblasts) and in vivo two-photon calcium imaging in C57BL/6J mice. Techniques included CRISPR-Cas9 knockout, siRNA-mediated gene silencing, molecular docking (AlphaFold 3), and protein-protein interaction analysis. Calcium flux was quantified via fluorescence imaging, while mitochondrial integrity and cytoskeletal dynamics were assessed using JC-1 staining, ATPase activity assays, and live-cell imaging. Structural validation of TRPC4-TRPC4AP binding sites utilized mutagenesis and complementation experiments. ResultsH2 selectively enhanced extracellular Ca{superscript 2} influx via TRPC4-TRPC4AP, with no cytotoxicity or mitochondrial dysfunction observed. Key arginine residues (730Arg-731Arg) in the TRPC4 CIRB domain formed hydrogen-bond networks essential for channel activation. In vivo, H2 increased neuronal Ca{superscript 2} transient frequency and amplitude in the primary motor cortex. TRPC4AP knockout abolished H2-induced Ca{superscript 2} influx, while mutagenesis of 730Arg/731Arg disrupted channel activity. H2 also promoted cytoskeletal remodeling and cell motility, dependent on TRPC4AP-mediated Ca{superscript 2} signaling. ConclusionsThis study identifies H2 as a novel calcium agonist that activates the TRPC4-TRPC4AP axis to regulate extracellular Ca{superscript 2} influx. The 730Arg-731Arg motif in TRPC4 serves as a critical H2-sensitive site, enabling dynamic calcium homeostasis without overload. These findings provide a mechanistic basis for H2-based therapies targeting calcium dysregulation in neurodegenerative, inflammatory, and metabolic diseases, while highlighting TRPC4AP as a pivotal molecular switch for gasotransmitter signaling.

biochemistry↗

bioRxiv2024

Spatial Single-Cell Mapping of Transcriptional Differences Across Genetic Backgrounds in Mouse Brains

Genetic variation can alter organ structure and, in turn, function. Comparative statistical analysis of organs across genetic backgrounds requires spatial, single-cell, atlas-scale data in replicates, which current technologies do not provide at scale. We introduce Atlas-scale Transcriptome Localization using Aggregate Signatures (ATLAS), a scalable tissue mapping method. ATLAS learns transcriptional signatures from scRNAseq data, encodes them in situ with tens of thousands of oligonucleotide probes, and decodes them to infer cell types and imputed transcriptomes. We validated ATLAS in the mouse brain by comparing its cell type inferences with direct MERFISH measurements of marker genes and quantitative comparisons to four other technologies. Using ATLAS, we mapped the central brains of five male and five female C57BL/6J (B6) mice and five male BTBR T+ tf/J (BTBR) mice, an idiopathic model of autism, collectively profiling over 40 million cells across over 400 coronal sections. Our analysis revealed over 40 significant differences in cell type distributions and identified 16 regional composition changes across male-female and B6-BTBR comparisons. ATLAS thus enables systematic comparative studies, facilitating organ-level structure-function analysis of disease models.

genomics↗

bioRxiv2024

Gradients of Recognition Molecules Shape Synaptic Specificity of a Visuomotor Transformation

Converting sensory information into motor commands is fundamental to most of our actions. In Drosophila, visuomotor transformations are mediated by Visual Projection Neurons (VPNs). These neurons convert object location and motion into directional behaviors downstream through a synaptic gradient mechanism. However, the molecular origins of such graded connectivity remain unknown. We addressed this question in a VPN cell type called LPLC2, which integrates looming motion and transforms it into an escape response through two parallel dorsoventral synaptic gradients at its inputs and outputs. We identified two corresponding dorsoventral expression gradients of cell recognition molecules within the LPLC2 population that regulate this synaptic connectivity. Dpr13 determines synaptic outputs of LPLC2 axons by interacting with its binding partner DIP-{varepsilon} expressed in the Giant Fiber, a neuron that mediates escape. Similarly, beat-VI regulates synaptic inputs onto LPLC2 dendrites by interacting with Side-II expressed in upstream motion-detecting neurons. Behavioral, physiological, and molecular experiments demonstrate that these coordinated molecular gradients control differential synaptic connectivity, enabling the accurate transformation of visual features into motor commands. As within-neuronal-type variation in gene expression is also observed in the mammalian brain, graded expression of cell recognition molecules may represent a common mechanism underlying synaptic specificity.

neuroscience↗

bioRxiv2024

Androgen deprivation-mediated activation of AKT is enhanced in prostate cancer with TMPRSS2:ERG fusion

TMPRSS2:ERG gene fusion (T:E fusion) in prostate adenocarcinoma (PCa) puts ERG under androgen receptor (AR) regulated TMPRSS2 expression. T:E fusion is associated with PTEN loss, and is highly associated with decreased INPP4B expression, which together may compensate for ERG-mediated suppression of AKT signaling. We confirmed in PCa cells and a mouse PCa model that ERG suppresses IRS2 and AKT activation. In contrast, ERG downregulation did not increase INPP4B, suggesting its decrease is indirect and reflects selective pressure to suppress INPP4B function. Notably, INPP4B expression is decreased in PTEN-intact and PTEN-deficient T:E fusion tumors, suggesting selection for a nonredundant function. As ERG in T:E fusion tumors is AR regulated, we further assessed whether AR inhibition increases AKT activity in T:E fusion tumors. A T:E fusion positive PDX had increased AKT activity in vivo and response to AKT inhibition in vitro after androgen deprivation. Moreover, two clinical trials of neoadjuvant AR inhibition prior to radical prostatectomy showed greater increases in AKT activation in the T:E fusion positive versus negative tumors. These findings indicate that AKT activation may mitigate the efficacy of AR targeted therapy in T:E fusion PCa, and that these patients may most benefit from combination therapy targeting AR and AKT.

cancer biology↗

bioRxiv2024

Proteomics Landscape of Triple-Negative Apocrine Breast Carcinoma Reveals Molecular Mechanisms of Tumorigenesis and Characteristics of Chemo-insensitivity

Triple-negative apocrine carcinoma (TNAC) is an exceptionally rare, chemotherapy-insensitive subtype of triple-negative breast cancer (TNBC) characterized by androgen receptor (AR) positivity and low proliferative activity. Despite poor chemotherapy response, TNAC exhibits favorable long-term survival, suggesting distinct resistance mechanisms. Here, we integrate mass spectrometry-based proteomics and whole-exome sequencing (WES) to characterize TNACs molecular features. We identify progressive overexpression of PI3K/AKT and AR signaling proteins, along with ECM remodeling and upregulation of GTPase-related proteins, highlighting TNACs invasive potential. WES reveals somatic mutations in ECM-related genes (COL18A1) and immune-associated genes (C3, NRDC), implicating the tumor microenvironment. Unlike TNBC, post-chemotherapy TNAC proteomics shows mixed ribosomal regulation and heightened inflammation, rather than reliance on proliferation, suggesting a need for targeted metabolic and immune therapies. Our findings support PI3K/AKT inhibitors and AR antagonists, potentially alongside GTPase inhibitors, metabolic disruptors, and immune therapy, as alternative strategies to standard chemotherapy in TNAC.

cancer biology↗

bioRxiv2024

Identification of Multi-landscape and Cell Interactions in the Tumor Microenvironment through High-Coverage Single-Cell Sequencing

Single-cell RNA sequencing (scRNA-seq) is a widely used method for classifying cell types and states and revealing disease mechanisms. However, most contemporary scRNA-seq platforms fail to explore the multilandscape of RNA. Here, we designed a microfluidic chip that combines oligo-dT primers and Random Bridging Co-labeling (RBCL) RNA sequencing to develop an innovative Chigene scRNA-seq technology that can identify gene expression, mutations, and RNA splicing landscapes at the single-cell level. The Chigene scRNA-seq platform demonstrated exceptional performance, with minimal doublet rates of 0.94% (Chigene V1) and 1.93% (Chigene V2). Both versions exhibit high sensitivity, with Chigene V2 achieving nearly 100% RNA coverage and detecting over 1800 genes per cell on average. Targeted capture of single-cell gene mutations enhances mutation detection sensitivity. Moreover, this Chigene V2 platform has been validated in clinical samples for its ability to detect mutations, gene fusions and alternative splicing. The reliability of the platform was further corroborated via known functional gene mutation (CDKN1A) and fusion (FGFR3-TACC). To validate this methods potential for discovering novel gene mutations in clinical samples, our investigation revealed an intriguing cell subpopulation carrying an ARHGAP5 mutation in urothelial carcinoma. These cells exhibited high-frequency mRNA splicing and exhibited specific crosstalk with T cells, distinguishing them from the subpopulation with the ARHGAP5 wild-type phenotype. Overall, this method provides a robust scRNA-seq platform suitable for comprehensive analyses of clinical specimens at different genetic information levels, thereby offering significant potential in the discovery of novel genes and interactions at the single-cell level.

genomics↗

bioRxiv2024

Biochemical characterization of extended-spectrum β-lactamases from Akkermansia genus

Akkermansia muciniphila, a member of the Verrucomicrobiota phylum, is recognized as a key gut microbe and has emerged as a potential next-generation probiotic. Assessment of antibiotic resistance in probiotics is a prerequisite for their therapeutic application, while very few is known in Akkermansia species. Firstly, we screened eight representative class A {beta}-lactamases (36.90%-41.30% identity with known {beta}-lactamases) from the Akkermansia species, which could increase the minimum inhibitory concentration (MIC) of Escherichia coli to{beta}-lactams. Secondly, four{beta}-lactamases were purified and identified as extended-spectrum {beta}-lactamase because they exhibited hydrolase activity against 19 {beta}-lactam antibiotics from penicillin, cephalosporins, and monobactam classes. Based on sequence alignment, three-dimensional structure, and binding pocket information, we hypothesized and validated that serine at 51 position was catalytic amino acid. Thirdly, the genomic context analysis revealed the absence of mobile genetic elements or other antibiotic resistance genes surrounding {beta}-lactamase genes, suggesting that the {beta}-lactamases from Akkermansia species may not be transferable. The finding and biochemical characterization of {beta}-lactamase from Akkermansia species provide a foundational basis for the safety evaluation of Akkermansia species as probiotics.

microbiology↗

bioRxiv2024

RinRK1 Promotes Accumulation of Nod Factor Receptors in Nanodomain-like structures at the tip of infected root hairs

Legume-rhizobia root-nodule symbioses involve the recognition of rhizobial Nod factor (NF) signals by NF receptors, triggering both nodule organogenesis and rhizobial infection. RinRK1 is induced by NF signaling and is essential for infection thread (IT) formation in Lotus japonicus. However, the precise mechanism underlying this process remains unknown. Here, we show that RinRK1 interacts with the extracellular domains of NF receptors NFR1 and NFR5 to promote their accumulation at root hair tips in response to rhizobia or NFs. Furthermore, Flotillin 1 (Flot1), a nanodomain-organizing protein, associates with the kinase domains of NFR1, NFR5 and RinRK1. RinRK1 promotes the interactions between Flot1 and NFR1 or NFR5 and both RinRK1 and Flot1 are necessary for the accumulation of NFR1 and NFR5 at root hair tips upon NF stimulation. Our study show that RinRK1 and Flot1 play a crucial role in NF receptor complex assembly within localized plasma membrane signaling centers to promote symbiotic infection.

plant biology↗