Biology subjects

Meng, N.

Publications and source records attributed to Meng, N..

9 recordsLinked to original sources

bioRxiv2025

A novel and evolutionarily distinct flavoprotein monooxygenase drives skatole degradation in Rhodococcus

Skatole (3-methylindole), a persistent and toxic N-heterocyclic aromatic pollutant, poses significant bioremediation challenges due to poorly defined biodegradation processes. This study systematically investigated the genetic determinants and metabolic mechanisms of skatole degradation in the environmentally versatile Gram-positive genus Rhodococcus. We demonstrated that skatole degradation was a broadly conserved trait across diverse Rhodococcus species, including R. aetherivorans, R. pyridinivorans, R. ruber and R. qingshengii. Genomic and functional analyses uncovered a novel flavoprotein monooxygenase (FPMO) SkaA as the initial catalyst for skatole catabolism. Heterologous expression in Escherichia coli confirmed SkaAs essential catalytic role, with high-resolution liquid chromatography-tandem mass spectrometry identifying the key product 3-methyloxindole and 3-hydroxy-3-methyloxindole. Distribution studies revealed SkaA homologs predominantly in Actinobacteria, particularly Nocardia and Rhodococcus, indicating conserved metabolic capacity for skatole transformation within this phylum. Notably, Rhodococcus strains lacking the skaA gene also retained skatole degradation activity, implying the existence of alternative genetic determinants. Crucially, phylogenetic analysis positioned SkaA as a distinct subclass within Group E FPMOs, exhibiting [≤]40% sequence identity to reported styrene/indole oxygenases. The phylogenetic segregation of skatole, indole, and styrene monooxygenases provides a predictive framework for functional annotation of Group E FPMOs. Our findings elucidate a novel skatole transformation pathway, establish Rhodococcus as environmentally versatile biocatalysts, and provide new insights into the study of Group E FPMOs. IMPORTANCESkatole is a notorious, foul-smelling compound generated by the anaerobic breakdown of tryptophan, presenting substantial risks to both the environment and public health. Despite its prevalence, the enzymatic machinery initiating its biodegradation remains poorly characterized. Our study resolves this critical gap by identifying SkaA as a functionally validated enzyme catalyzing skatoles initial oxidation step. Distribution analyses revealed SkaA homologs predominantly in Actinobacteria, particularly Rhodococcus and Nocardia. Phylogenetic analysis positioned SkaA as a novel third subclass within Group E flavoprotein monooxygenases. These breakthroughs provide molecular tools for engineering targeted bioremediation solutions and new insights into the study of Group E FPMOs.

microbiology↗

bioRxiv2025

Automated and parallelized microfluidic generation of large and precisely-defined lipid nanoparticle libraries

Building on the success of lipid nanoparticles (LNPs) in vaccines, LNPs are being developed for a broad set of therapeutic applications by changing both the structures of the lipids used to formulate each LNP and their relative proportions. Because lipid synthesis and in vivo screening have been parallelized using combinatorial chemistry and LNP barcoding respectively, the manual and sequential microfluidic formulation of LNPs has become the rate-limiting step in the discovery process. In this work, we present a high-throughput, automated microfluidic platform capable of generating large, precisely-defined LNP libraries in parallel at a rate of one distinct formulation every three seconds. Each formulation is defined by varying the reagent flow ratios into one of eight microscale mixers using litho-graphically encoded fluidic resistors and dynamically controlled external pressure supplies. The microfluidic chip is integrated with custom frobotic plate handling for the rapid collection of each distinct formulation. Using this platform, we produce a library of 96 formulations, which we profile physicochemically and evaluate in terms of both in vitro and in vivo transfection.

bioengineering↗

bioRxiv2025

Transcriptome-wide Identification of GlycoRNAs by Clier-seq Pipeline

RNA molecules can undergo modification by N-glycans and be displayed on the cell surface. However, recent studies have focused primarily on N-glycan modifications on small RNAs less than 200 nt in length; the transcriptome-wide subtypes of glycosylated RNAs (glycoRNAs) remain poorly characterized. A validation system, which represents only a fraction of the total transcripts, has yet to be established to analyze glycoRNAs accurately. In this study, we aimed to comprehensively characterize transcriptome-wide global glycoRNAs and novel glycoRNA subtypes in both epithelial cells and B cells. Using metabolic labeling and density gradient centrifugation methods, we identified glycoRNAs predominantly below 2000 nt in both epithelial cells and B cells. We then developed the Clier-seq (Click chemistry-based enrichment of glycoRNAs sequencing) method to maximize the coverage of glycoRNAs (ranging from 50 to 2000 nt) and utilized the HISAT-StringTie-Ballgown pipeline to predict novel glycoRNA subtypes. We also established Clier-qPCR assays (click chemistry-based enrichment of glycoRNAs RT-qPCR) to validate the specificity of the candidate glycoRNAs. We demonstrated that transfer RNAs (tRNAs), particularly tRNAs (Ser), tRNAs (Thr), tRNAs (Val), and tRNAs (lys), are the primary targets of glycosylation. Additionally, we found that vault RNAs (vtRNAs), specifically vtRNA2-1, are glycosylated. Furthermore, we discovered several novel glycosylated long noncoding RNAs ranging from 200-400 nt in length. Herein, we propose a standardized bioinformatics pipeline for glycoRNA research, enabling accurate and comprehensive identification of glycoRNAs throughout the transcriptome.

molecular biology↗

bioRxiv2024

Diverse quorum sensing systems regulate microbial communication and biogeochemical processes in deep-sea cold seeps

Quorum sensing is a fundamental chemical communication mechanism that enables microorganisms to coordinate behavior and adapt to environmental conditions. In deep-sea cold seep ecosystems, where diverse microbial communities thrive, quorum sensing is likely a key mechanism driving microbial interactions. However, the distribution, mechanisms, and ecological roles of quorum sensing in cold seeps remain poorly understood. To address this, we analyzed 173 metagenomes, 33 metatranscriptomes, and 18 metabolomes from 17 global cold seep sites. We identified 299,355 quorum sensing genes from the seep non-redundant gene catalog, representing 34 gene types across six quorum sensing systems, with distribution patterns influenced by sediment depth and seep type. These quorum sensing genes were present in 3,576 metagenome-assembled genomes from 12 archaeal and 108 bacterial phyla, revealing a complex network of intraspecies and interspecies communication. Microbial groups involved in key metabolic processes, such as sulfate-reducing bacteria, anaerobic methanotrophic archaea, diazotrophs and organohalide reducers, were extensively regulated by quorum sensing, influencing biogeochemical cycles of carbon, nitrogen, and sulfur. Phylogenetic analysis and protein domain identification highlighted the functional roles of key quorum sensing-related proteins (e.g., PDEs, CahR, RpfC/G and LuxR) in modulating microbial behaviors, such as motility and chemotaxis. Heterologous expression and metabolomic profiling further confirmed the activity of representative LuxI-R pairs and identified inhibitors of quorum sensing in cold seep sediments. Overall, these findings highlight complexity and significance of quorum sensing in microbial interactions, ecological adaptation, and biogeochemical cycling within cold seep ecosystems, advancing our understanding of microbial communication in the deep biosphere.

microbiology↗

bioRxiv2023

Mutualist-Provided Resources Increase Susceptibility to Parasites

Background and AimsO_LIMutualistic microbes often increase their hosts susceptibility to parasite infection, but the mechanism underlying this pattern remains unknown. C_LIO_LIWe tested two competing hypotheses to identify the cause of this phenomenon. C_LIO_LIFirst, mutualist-provided resources could attract antagonists by making hosts more resource-rich. C_LIO_LISecond, mutualism establishment itself might increase host vulnerability to antagonists by enabling antagonists to exploit host defenses or immune systems, which are often downregulated during mutualism establishment. C_LI MethodsO_LITo test which mechanism underlies increased parasite susceptibility, we experimentally decoupled mutualism establishment and mutualist-provided resources in the legume-rhizobia mutualism. C_LIO_LIWe measured parasite load on Medicago truncatula plants infected with root-knot nematodes in a full-factorial design, in which we independently manipulated rhizobia inoculation (mutualism establishment) and nitrogen availability (mutualist-provided resources). C_LI Key ResultsO_LIIncreased nitrogen availability increased parasite infection. However, its effect was non-linear and was not explained by nitrogen assimilation into plant tissues, indicating that this effect is not driven by parasite attraction to resource-rich hosts. C_LIO_LIWe found little effect of mutualism establishment on susceptibility, as neither rhizobia inoculation nor nodulation without nitrogen fixation significantly increased parasite infection. C_LI ConclusionsO_LIOur results suggest that mutualist-provided resources are an important driver of indirect ecological costs of mutualism, although the mechanism linking mutualist-provided resources and susceptibility to infection remains unknown. C_LI

ecology↗

bioRxiv2023

Spatiotemporal transcriptome atlas of human embryos after gastrulation

The spatial and temporal atlas of gene expression in the human embryo at early gestation is critical in understanding embryo development, organogenesis, and disease origins. We obtained the spatiotemporal transcriptome from 90 sagittal sections of 16 whole human embryos from 3 to 8 post-conception weeks by Stereo-seq with high resolution and ultra-large field, establishing the development trajectory/regulatory profiling of 49 organs. We uncovered the organ-specific regulons as potential lineage-determining factors and identified the new regulatory networks during heart and brain development. The atlas refines the key organs/cell types vulnerable to virus infection and genetic disorders, and, reveals the dynamics of allelic gene expression in specific organs at different stages. These results present the first comprehensive delineation of the spatiotemporal transcriptomic dynamics of human organogenesis. One Sentence SummaryThe spatiotemporal transcriptome atlas presents a comprehensive delineation of human embryogenesis after gastrulation.

developmental biology↗

bioRxiv2022

Direct fluorescent labeling of NF186 and NaV1.6 in living primary neurons using bioorthogonal click chemistry

The axon initial segment (AIS) is a highly specialized neuronal compartment that regulates the generation of action potentials and maintenance of neuronal polarity. Despite its importance, live imaging of the AIS is challenging due to the limited number of suitable labeling methods. To overcome this limitation, we established a novel approach for live labeling of the AIS using unnatural amino acids (UAAs) and bioorthogonal click chemistry. The small size of the UAAs and the possibility of introducing them virtually anywhere into the target proteins make this method particularly suitable for live labeling and imaging of complex and spatially restricted proteins. With this approach, we labeled two large AIS components, the 186 kDa isoform of neurofascin (NF186) and the 260 kDa voltage-gated sodium channel (NaV1.6), and performed widefield and confocal microscopy in fixed and living neurons. Moreover, we demonstrated the applicability of this method by studying the localization of two epilepsy-causing NaV1.6 variants with a loss-of-function effect. Finally, to further improve the efficiency of the UAA incorporation, we developed adeno-associated viral (AAV) vectors for click labeling in primary neurons. The use of AAV vectors will facilitate the transfer of UAA-based click labeling technology to more complex biological systems, such as organotypic slice cultures, organoids, and animal models.

neuroscience↗

bioRxiv2021

A sexual dimorphic role of the miR-465 cluster in placental development

A sexually dimorphic role of miRNAs in placental development has never been reported. Here, we show that ablation of the miR-465 cluster caused selective degeneration of female conceptuses as early as embryonic day (E)8.5, leading to a male-biased sex ratio (60% males) among miR-465 KO mice. Given that the miR-465 cluster miRNAs were predominantly expressed in the extraembryonic tissue and ablation of these miRNAs led to dysregulation of numerous critical placental genes, our data strongly suggest the miR-465 cluster is required for full developmental potential of the female, but not the male, extraembryonic tissue/placenta.

developmental biology↗

bioRxiv2017

Biopipe: A Lightweight System Enabling Comparison of Bioinformatics Tools and Workflows

Analyzing next generation sequencing data always requires researchers to install many tools, prepare input data compliant to the required data format, and execute the tools in specific orders. Such tool installation and workflow execution process is tedious and error-prone, and becomes very challenging when researchers need to compare multiple alternative tool chains. To mitigate this problem, we developed a new lightweight and portable system, Biopipe, to simplify the creation and execution of bioinformatics tools and workflows, and to further enable the comparison between alternative tools or workflows. Biopipe allows users to create and edit workflows with user-friendly web interfaces, and automates tool installation as well as workflow synthesis by downloading and executing predefined Docker images. With Biopipe, biologists can easily experiment with and compare different bioinformatics tools and workflows without much computer science knowledge. There are mainly two parts in Biopipe: a web application and a standalone Java application. They are freely available at http://bench.cs.vt.edu:8282/Biopipe-Workflow-Editor-0.0.1/index.xhtml and https://code.vt.edu/saima5/Biopipe-Run-Workflow\n\nContactnm8247@cs.vt.edu\n\nSupplementary informationSupplementary data are available online.

bioinformatics↗