Mucosal modulation by LGG and R. gnavus specific tryptophan metabolites

LGG 和 R. gnavus 特异性色氨酸代谢物对粘膜的调节

• 批准号: 9788244
• 负责人: RONALDO PARAOAN FERRARIS
• 金额: $ 38.75万
• 依托单位: RUTGERS THE STATE UNIV OF NJ NEWARK
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-20 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9788244
• 关键词: 16S ribosomal RNA sequencing; ARNT gene; Ablation; Acute; Address; Affect; Affinity; Agonist; Animal Model; Anti-inflammatory; Aryl Hydrocarbon Receptor; Bacteria; Binding; Biological; Biological Assay; Cell Compartmentation; Cell Nucleus; Cell physiology; Colitis; Collaborations; Data; Diet; Disease; Elements; Engineering; Epithelial; Epithelial Cells; Essential Amino Acids; Excision; Gene Expression; Genes; Genetic; Germ-Free; Gnotobiotic; Health; Homeostasis; Immune; Impairment; Inflammatory disease of the intestine; Innate Immune Response; Intervention; Intestinal Mucosa; Intestines; Knock-out; Knockout Mice; Lactobacillus; Lactobacillus casei rhamnosus; Lamina Propria; Ligands; Link; Literature; Mammals; Mediating; Mucolytics; Mucosal Immune Responses; Mucous Membrane; Mus; Nature; Organoids; Pathway interactions; Perfusion; Physiology; Play; Predisposition; Probiotics; Property; Protein Biosynthesis; Public Health; Receptor Activation; Receptor Signaling; Recording of previous events; Role; Signal Pathway; Signal Transduction; Source; Structure; System; Testing; Tight Junctions; Tryptamines; Tryptophan; Work; aryl hydrocarbon receptor ligand; bacterial genetics; beta catenin; cytokine; dietary manipulation; dimer; dysbiosis; experience; high risk; host-microbe interactions; innovation; metabolomics; microbial; microbial community; microbiota; mouse model; mutant; pathobiont; promoter; receptor structure function; screening; stem cell division; transcriptome sequencing

PROJECT SUMMARY: The dietary benefits of tryptophan have been classically linked to its property as an essential amino acid required by mammals for protein synthesis, but recent work in the past decade has revealed its equally essential role as an important source of metabolites for the microbial community to generate signals, mainly via the Aryl Hydrocarbon Receptor (AHR), to affect a plethora of cellular functions. AHR signaling pathway is highly promiscuous not only responding to diverse ligands with resembling structures, but also crosstalk with and is modulated by numerous cellular signaling pathways. Distinct agonists can result in ligand selective differences in AHR structure and function, ultimately leading to agonist-selective gene expression and biological effects. In the gut, AHR activation by bacterial tryptophan catabolites influences mucosal homeostasis and microbiota composition. Although bacterial tryptophan metabolites and AHR axis may contribute to microbial sensing and tolerance, it is unclear if probiotics and pathobionts produce distinct tryptophan metabolites thereby eliciting bacterial species- and agonist-specific host mucosal modulating effects. Both Lactobacillus and Ruminoccocus spp catabolize tryptophan to a variety of tryptamine and indolic products that bind at low to high affinities to the AHR. The identities and specific biological effects of tryptophan metabolites produced by the probiotic Lactobacillus rhamnosus GG (LGG) and the pathobiont R. gnavus are currently unknown. Preliminary RNA-Seq analysis of mouse intestinal mucosa perfused with live LGG revealed a robust induction of AHR signaling activity as well as anti-inflammatory cytokines. Bacterial 16s rRNA sequencing and untargeted fecal metabolomics screening of a newly engineered Lyz1 knockout mouse model revealed a dysbiosis featured by expansion of mucolytic bacterial species (e.g., R. gnavus), accompanied by a pronounced elevation of tryptophan metabolites and a changed epithelial cell composition. The hypothesis is that the probiotic LGG and the pathobiont R. gnavus metabolize dietary tryptophan into distinct agonists that elicit differential mucosal modulating effects via the host AHR signaling. Aim I will use mono-colonization of germ-free mice, dietary tryptophan depletion, unbiased metabolomics profiling, and isogenic mutant bacteria deficient in tryptophan-metabolizing to identify LGG and R. gnavus specific tryptophan metabolites and their differential mucosal modulating effects. Aim 2 will employ organoid culture, intraluminal perfusion of AHR agonists, and genetic ablation of host AHR signaling to determine the direct effects of bacterial species-specific tryptophan metabolites and AHR signaling on Lyz1-deficient mouse mucosal homeostasis and colitis susceptibility. This MPI project uses rigorous approaches, including gnotobiotics, bacterial genetics, dietary manipulation, and unbiased metabolomics, etc. to address an important mechanism involving diet-microbe-host interaction that may underlie the biological natures of probiotics and pathobionts. The two PIs have a history of productive collaboration, and have demonstrated experiences in studying intestinal physiology. This project employing innovative animal models is significant for public health and disease intervention.

项目概要： 色氨酸的饮食益处与其作为必需氨基酸的特性有关 哺乳动物的蛋白质合成，但最近的工作在过去的十年中已经揭示了它同样重要的作用， 微生物群落产生信号的代谢产物的重要来源，主要通过芳基 碳氢化合物受体（AHR），影响过多的细胞功能。AHR信号通路高度 混杂的不仅响应于具有相似结构的不同配体，而且还与 由许多细胞信号通路调节。不同的激动剂可导致配体选择性差异 在AHR结构和功能中，最终导致激动剂选择性基因表达和生物学效应。在 肠道，细菌色氨酸催化剂激活AHR影响粘膜稳态和微生物群 混合物.虽然细菌色氨酸代谢物和AHR轴可能有助于微生物传感， 耐受性，尚不清楚益生菌和致病菌是否产生不同的色氨酸代谢物，从而引起 细菌物种和激动剂特异性宿主粘膜调节作用。乳酸菌和瘤胃球菌 spp将色氨酸分解代谢为多种色胺和吲哚产物，其以低至高的亲和力结合到 AHR益生菌色氨酸代谢产物的特性及其生物学效应 鼠李糖乳杆菌（Lactobacillus rhamnosus GG，LGG）和致病菌R.目前尚不清楚。初步RNA-Seq 用活LGG灌注的小鼠肠粘膜的分析揭示了AHR信号传导活性的强诱导 以及抗炎细胞因子。细菌16 s rRNA测序和非靶向粪便代谢组学 筛选一个新的工程Lyz 1敲除小鼠模型，揭示了一种以扩增为特征的生态失调， 粘液溶解细菌物种（例如，R. gnavus），伴随着色氨酸代谢物的显著升高 以及上皮细胞组成的改变假设益生菌LGG和致病菌R.有痣的 将饮食色氨酸代谢成不同的激动剂，通过宿主引起不同的粘膜调节作用 AHR信号。目的我将使用单菌落无菌小鼠，饮食色氨酸耗竭，无偏倚 代谢组学分析，和在葡聚糖代谢缺陷的同基因突变体细菌鉴定LGG和R. gavus特异性色氨酸代谢物及其不同的粘膜调节作用。目标2将采用 类器官培养、AHR激动剂的腔内灌注和宿主AHR信号传导的基因消融，以确定 细菌种特异性色氨酸代谢产物和AHR信号对Lyz 1缺陷小鼠的直接影响 粘膜稳态和结肠炎易感性。这个MPI项目使用严格的方法，包括 gnotobiotics，细菌遗传学，饮食操纵，和公正的代谢组学等，以解决一个重要的 涉及饮食-微生物-宿主相互作用的机制，可能是益生菌生物学性质的基础， 致病生物这两个PI有着富有成效的合作历史，并在以下方面展示了经验： 研究肠道生理学该项目采用创新的动物模型对公共卫生具有重要意义 和疾病干预。