AN INTERBACTERIAL ADAPTIVE IMMUNE SYSTEM ENCODED BY BACTEROIDALES

拟杆菌编码的细菌间适应性免疫系统

• 批准号: 10045595
• 负责人: Benjamin Ross
• 金额: $ 24.81万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-10 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10045595
• 关键词: Adaptive Immune System; Amino Acids; Animals; Anti-Bacterial Agents; Antibiotic Resistance; Bacteria; Bacteroides; Bacteroides fragilis; Binding; Biochemistry; Biogenesis; Bioinformatics; Biological Assay; Biology; Biosensor; Cells; Cellular biology; Communities; Core Facility; DNA cassette; Development; Disease; Ecosystem; Engineered Probiotics; Engraftment; Escherichia coli; Excision; Exhibits; Foundations; Frequencies; Future; Gene Frequency; Generations; Genes; Genetic; Genome; Gnotobiotic; Growth; Health; Human; Immunity; In Situ; In Vitro; Integrase; Integrons; Knowledge; Mediating; Methods; Molecular; Mus; Mutation; Nature; Orphan; Pathway interactions; Probiotics; Production; Proteins; Proteobacteria; Research; Role; Sampling; Specificity; Suggestion; System; Testing; Time; Toxin; Training; Tyrosine; Universities; Washington; Work; bacterial community; bacterial genetics; design; experience; experimental study; fitness; gene function; gut bacteria; gut microbiome; gut microbiota; improved; in vitro Assay; in vivo; insight; member; microbiota; pressure; prevent; probiotic therapy; programs; recombinase; recruit; residence; tool

PROJECT SUMMARY / ABSTRACT The composition of bacterial communities is paramount to their function. Intense competitive interactions between bacteria can influence community composition by altering assembly or stability through the production of anti-bacterial molecules. One such ecosystem rife with competitive interactions is the human gastrointestinal microbiota, which harbors an abundance of bacteria from the order Bacteroidales. These bacteria encode the type VI secretion system (T6SS), a contact-dependent toxin delivery pathway shown to mediate potent inter- species competition. Bacteria that encode the T6SS also encode immunity proteins which bind to and specifically inactivate toxins. I have found that Bacteroidales genomes possess arrays of “orphan immunity” genes in the absence of the T6SS or corresponding toxin. These polyimmunity arrays are found associated with xerD-like (PAX) recombinase genes that suggest an active mechanism of recruitment. I hypothesize that genes within PAX clusters represent selective pressure from past episodes of direct interbacterial antagonism. Therefore, characterization of the function, activity, and biogenesis of PAX clusters offers an avenue for the identification of direct bacterial interactions in vivo and an understanding of their ecological consequences. This proposal aims to augment my interdisciplinary background in cell biology, evolutionary biology, and genetics with new training in germfree mouse experimentation to investigate this new mechanism of adaptation to T6SS-mediated interbacterial antagonism within Bacteroidales and understand its impact on gut microbiome assembly and dynamics. In Aim 1, I propose to characterize the function of genes within PAX clusters through in vitro expression experiments in E. coli and growth competition experiments in vitro and in vivo under conditions that promote contact-dependent interbacterial antagonism. In Aim 2, I will test the hypothesis that the PAX recombinase mediates gene acquisition and excision from PAX clusters via tyrosine recombinase activity in a manner analogous to integrons. In Aim 3, I will extend my characterization of PAX clusters to their use as a new tool for deciphering interbacterial interactions by sequencing new gene insertions after exposure of a PAX-containing strain to different bacterial communities in vivo. I will train with Dr. Lynn Hajjar at the University of Washington Gnotobiotic Animal Core facility to gain experience in using germfree mice to study the role of PAX clusters in the gut microbiota in vivo. Dr. Andrew Goodman at Yale University will collaborate and provide advice and intellectual support. The proposed experiments will lay the foundation for my independent research program, in which bioinformatics, bacterial genetics, and in vivo experimentation will be combined to yield insight into direct interactions within the gut microbiota and the implications of these interactions for human health and disease.

项目摘要 /摘要 细菌合作社的组成对于它们的功能至关重要 细菌之间可以通过改变组装或稳定性来影响共同的综合。 抗菌分子的生态系统与竞争性相互作用 这些细菌编码这些细菌编码。 VI型分泌系统（T6SS），一种接触依赖性的毒素递送途径途径有效的间 物种竞争。 特别是毒素。 在没有T6S或相关毒素的基因中。 用Xerd样（PAX）重组酶基因提出了一种主动募集机制 PAX簇中的基因代表了直接细菌间拮抗作用的选择性压力。 因此，PAX簇的功能，活性和生物发生的表征为它们提供了途径 在体内识别直接的双作用相互作用，并识别其生态依赖性的识别。 该建议旨在增加我在细胞生物学，进化生物学和 遗传学和新培训在无用的鼠标经验中进行了研究，以研究这种适应的新机制 到T6SS介导的双脚类动物间拮抗作用，并了解其对肠道微生物组的影响 组装和动力学。 大肠杆菌和生长竞争的体外表达实验在体外和体内实验 在AIM 2中促进接触依赖性拮抗作用的条件。 PAX重组酶通过酪氨酸重组酶从簇中介导基因的获取和切除 以类似于Intimones的方式进行活动。 通过对新基因铭文进行测序，用作解密间相互作用的新工具 在体内对不同细菌群落的pax菌株。 华盛顿大学gnotobiotic Animal核心设施，以获得使用无用的老鼠的经验 PAX簇在体内肠道微生物群中的作用。 并提供智力支持。 独立研究计划，其中生物信息学，细菌遗传学和体内经验将是 结合屈服于肠道微生物群中的直接相互作用及其含义） 人类健康和疾病的相互作用。