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.

项目摘要 霍乱的病因霍乱弧菌（VC），将小肠的粘膜表面定居。英语 通过病毒因子介导，以穿透粘液层，附着于上皮细胞，并增殖，所有这些 同时调节与宿主细胞和肠道菌群的相互作用。人类肠道微生物群是 高度多样化，以及微生物群的结构和功能的人际变化驱动着戏剧性的不同 - 在VC殖民化中加密。在微生物微生物和微生物 - 宿主相互作用的中心是宿主粘液 层由分泌的粘蛋白糖蛋白（包括显性粘蛋白MUC-2）组成。粘液提供 - 病原体和肠道菌群的锥度和碳源 上皮和肠腔的界面，为感染提供了关键的物理障碍。但是，角色 VC与肠道菌群的特定构型之间的粘蛋白代谢互换的 并且对个性化VC感染结果的影响并不是很好。在这里，我们表明 Taga是VC病毒主调节器TOXT更新的分泌金属蛋白酶，促进VC增长 在粘蛋白中，taga是一种双功能蛋白 DUCES宿主粘蛋白产生。使用离体组织培养和体内gnotobiotic小鼠的组合 结合模型与缺乏蛋白水解活性或MUC2诱导的活性的TAGA突变体相结合， 我们发现，塔加（Taga）的两项活动在感染过程中对VC适应性产生不同的影响 特定人类肠道微生物的影响。我们已经生成了人类肠道的肠道菌群特征 微生物群指出：一种模型的微生物群落，类似于健康个体，它促进了风险投资 派系耐药性，以及在霍乱内构中发现的不植物状态的另一个模型菌群特征 与VC定植的高敏感性相关的区域。 taga粘液溶解活性对风险感染很重要 抗性抗菌菌群内的抗性，而粘蛋白诱导的TAGA FN3活性导致 增生性群落中VC感染的增加。因此，我们假设标签驱动VC代谢 与特定肠道菌群的相互作用，导致特定于社区的依恋，增长和整体感染 结果。我们将以四个特定目标来检验该核心假设的各个方面。 AIM 1将阐明机制 在上皮附着和生长中，VC与共生肠道微生物的依赖粘蛋白依赖性相互作用。 AIM 2将检查霍乱内部分子区域中个性化的肠道微生物群结构如何调节粘蛋白和 TAGA依赖性疾病表型。 AIM 3将决定TAGA-MICROBIOTA相互作用如何推动生产 - 宿主粘蛋白的作用和代谢。最后，AIM 4将阐明邻近性和空间特异性的作用 驱动依赖微生物群的VC疾病结果。此应用的最终目标是阐明 病原体介导的粘蛋白代谢在肠感染过程中微生物相互作用中的作用。