The role of a bifunctional mucinase in modulating personalized gut microbiota-Vibrio cholerae interactions during infection

双功能粘蛋白酶在感染期间调节个性化肠道微生物群-霍乱弧菌相互作用中的作用

• 批准号: 10749595
• 负责人: Ansel Hsiao
• 金额: $ 80.24万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10749595
• 关键词: Adherence; Affect; Area; Automobile Driving; Bangladesh; Carbohydrates; Carbon; Cell model; Cells; Characteristics; Cholera; Communities; Complex; Developing Countries; Diarrhea; Disease; Disease Outcome; Enzymes; Epithelial Attachment; Epithelial Cells; Epithelium; Etiology; Fibronectins; Genetic Determinism; Genetic Techniques; Genetic Transcription; Glycoproteins; Gnotobiotic; Goals; Goblet Cells; Growth; Human; Human Characteristics; In Vitro; Individual; Infection; Intervention; Intestines; Malnutrition; Mediating; Metabolic; Metabolism; Metalloproteases; Microbe; Modeling; Molecular; Morbidity - disease rate; Muc 2 protein; Mucin-2 Staining Method; Mucins; Mucolytics; Mucous Membrane; Mucous body substance; Mus; Outcome; Pathogenesis; Penetration; Play; Population; Population Heterogeneity; Predisposition; Process; Production; Proliferating; Proteins; Recurrence; Resistance; Resistance to infection; Resolution; Role; Series; Shapes; Site; Small Intestines; Source; Specificity; Structure; Surface; Testing; Variant; Vibrio cholerae; Vibrio cholerae infection; Virulence; Virulence Factors; cell motility; cohort; diarrheal disease; disease phenotype; dysbiosis; enteric infection; fecal microbiota; fitness; gut microbes; gut microbiome; gut microbiota; host-microbe interactions; in vivo; member; microbial community; microbiome; microbiota; microorganism; microorganism interaction; mortality; mucinase; mutant; novel therapeutics; pathogen; prophylactic; response; tissue culture

PROJECT SUMMARY Vibrio cholerae (Vc), the causative agent of cholera, colonizes the mucosal surface of the small intestine. Infec- tion is mediated via virulence factors to penetrate the mucus layer, attach to epithelial cells, and proliferate, all the while modulating interactions with both host cells and the gut microbiota. The human gut microbiota is highly diverse, and interpersonal variation in the structure and function of the microbiota drives dramatic differ- ences in Vc colonization. At the center of microbe-microbe and microbe-host interactions lies the host mucus layer, comprised of secreted mucin glycoproteins including the dominant mucin MUC-2. Mucus provides at- tachment sites and carbon sources for both pathogens and commensal members of the gut microbiota at the interface of the epithelium and the gut lumen, as provides a key physical barrier to infection. However, the role of mucin metabolism in metabolic interchanges between Vc and specific configurations of the gut microbiota, and the resulting impact on personalized Vc infection outcomes, has not been well studied. Here, we show that TagA, a secreted metalloprotease upregulated by the Vc virulence master regulator ToxT, promotes Vc growth in mucin, and that TagA is a bifunctional protein, acting as both mucinase and a mucus secretagogue that in- duces host mucin production. Using a combination of ex vivo tissue culture and in vivo gnotobiotic mouse colonization models combined with TagA mutants lacking either proteolytic activity or MUC2-inducing activity, we found that TagA’s two activities have different effects on Vc fitness during infection depending on the pres- ence of specific human gut microbes. We have generated model gut microbiota characteristic of human gut microbiota states: one model microbial community similar to that of healthy individuals, which promotes Vc in- fection resistance, and another model microbiota characteristic of the dysbiotic state found in cholera endemic areas associated with high susceptibility to Vc colonization. TagA mucolytic activity is important for Vc infection resistance within the colonization-resistant microbiota, while the mucin-inducing activity of TagA FN3 leads to increased Vc infection within dysbiotic communities. Therefore, we hypothesize that Tag drives Vc metabolic interactions with specific gut microbiota leading to community-specific attachment, growth, and overall infection outcomes. We will test aspects of this core hypothesis in four specific aims. Aim 1 will elucidate mechanisms driving mucin-dependent interactions of Vc with commensal gut microbes in epithelial attachment and growth. Aim 2 will examine how personalized gut microbiota structure in cholera endemic areas modulates mucin- and TagA-dependent disease phenotypes. Aim 3 will determine how TagA-microbiota interactions drives produc- tion and metabolism of host mucins. Finally, Aim 4 will elucidate the role of proximity and spatial specificity in driving microbiota-dependent Vc disease outcomes. The ultimate goal of this application is to shed light on the role of pathogen-mediated mucin metabolism in microbial interactions during enteric infection.

项目概要 霍乱弧菌 (Vc) 是霍乱的病原体，定植于小肠粘膜表面。 化是通过毒力因子介导穿透粘液层，附着于上皮细胞并增殖，所有 同时调节与宿主细胞和肠道微生物群的相互作用。 微生物群结构和功能的高度多样化和人际差异导致显着差异 Vc定植过程中微生物-微生物和微生物-宿主相互作用的核心是宿主粘液。 层，由分泌的粘蛋白糖蛋白组成，包括主要的粘蛋白 MUC-2，提供 at-。 病原体和肠道微生物群共生成员的附着位点和碳源 上皮和肠腔的界面，为感染提供了关键的物理屏障。 Vc 和肠道微生物群特定结构之间代谢交换中的粘蛋白代谢， 以及由此产生的对个性化 Vc 感染结果的影响尚未得到充分研究。 TagA 是一种分泌型金属蛋白酶，受 Vc 毒力主调节因子 ToxT 上调，可促进 Vc 生长 TagA 是一种双功能蛋白，既充当粘蛋白酶又充当粘液分泌促进剂， 使用离体组织培养和体内无菌小鼠的组合来诱导宿主粘蛋白的产生。 定植模型与缺乏蛋白水解活性或MUC2诱导活性的TagA突变体相结合， 我们发现，根据不同的情况，TagA 的两种活性在感染期间对 Vc 适应度有不同的影响。 我们已经生成了人类肠道特征的模型肠道微生物群。 微生物群指出：一种与健康个体相似的模型微生物群落，可促进 Vc- 感染抵抗力，以及霍乱地方病中发现的生态失调状态的另一种模型微生物群特征 与 Vc 定植高度易感性相关的区域，TagA 粘液溶解活性对于 Vc 感染很重要。 定植抗性微生物群中的抗性，而 TagA FN3 的粘蛋白诱导活性导致 生态失调群落中的 Vc 感染增加，因此，我们帮助 Tag 驱动了 Vc 代谢。 与特定肠道微生物群的相互作用，导致群落特异性附着、生长和整体感染 我们将在四个具体目标中测试这一核心假设的各个方面，以阐明机制。 在上皮附着和生长中驱动 Vc 与共生肠道微生物的粘蛋白依赖性相互作用。 目标 2 将研究霍乱流行地区的个性化肠道微生物群结构如何调节粘蛋白和 TagA 依赖性疾病表型将决定 TagA-微生物群相互作用如何驱动生产。 最后，目标 4 将阐明邻近性和空间特异性在其中的作用。 该应用的最终目标是阐明微生物群依赖性 Vc 疾病的结果。 病原体介导的粘蛋白代谢在肠道感染期间微生物相互作用中的作用。