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: 10064021
• 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 2021-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10064021
• 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.

项目概要/摘要 细菌群落的组成对其功能至关重要。激烈的竞争互动 细菌之间的相互作用可以通过改变生产过程中的组装或稳定性来影响群落组成 的抗菌分子。人类胃肠道就是这样一个充满竞争性相互作用的生态系统 微生物群，含有大量拟杆菌目细菌。这些细菌编码 VI 型分泌系统（T6SS），一种接触依赖性毒素传递途径，被证明可以介导有效的相互作用 物种竞争。编码 T6SS 的细菌也编码免疫蛋白，这些蛋白结合并 专门灭活毒素。我发现拟杆菌基因组具有一系列“孤儿免疫” 在没有 T6SS 或相应毒素的情况下的基因。这些多重免疫阵列被发现相关 xerD 样 (PAX) 重组酶基因表明存在主动的招募机制。我假设 PAX簇内的基因代表了来自过去细菌间直接拮抗作用的选择压力。 因此，PAX簇的功能、活性和生物发生的表征为研究提供了一条途径。 识别体内直接细菌相互作用并了解其生态后果。 该提案旨在增强我在细胞生物学、进化生物学和 遗传学通过无菌小鼠实验的新训练来研究这种新的适应机制 拟杆菌内 T6SS 介导的细菌间拮抗作用，并了解其对肠道微生物组的影响 装配和动力学。在目标 1 中，我建议通过以下方式表征 PAX 簇内基因的功能： 大肠杆菌体外表达实验和体内外生长竞争实验 促进接触依赖性细菌间拮抗作用的条件。在目标 2 中，我将检验以下假设： PAX 重组酶通过酪氨酸重组酶介导 PAX 簇中的基因获取和切除 以类似于整合子的方式活动。在目标 3 中，我将把 PAX 集群的特征扩展到它们的 用作通过暴露后对新基因插入进行测序来破译细菌间相互作用的新工具 含有 PAX 的菌株对体内不同细菌群落的影响。我将在 Lynn Hajjar 博士的指导下进行培训 华盛顿大学知生动物核心设施将获得使用无菌小鼠进行研究的经验 PAX 簇在体内肠道微生物群中的作用。耶鲁大学安德鲁·古德曼博士将合作 并提供建议和智力支持。所提出的实验将为我的研究奠定基础 独立研究计划，其中生物信息学、细菌遗传学和体内实验将 结合起来，深入了解肠道微生物群内的直接相互作用及其影响 人类健康和疾病之间的相互作用。

项目成果

Assembly of a unique membrane complex in type VI secretion systems of Bacteroidota.

• DOI: 10.1038/s41467-023-44426-1
• 发表时间: 2024-01-10
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Bongiovanni, Thibault R.;Latario, Casey J.;Le Cras, Youn;Trus, Evan;Robitaille, Sophie;Swartz, Kerry;Schmidtke, Danica;Vincent, Maxence;Kosta, Artemis;Orth, Jan;Stengel, Florian;Pellarin, Riccardo;Rocha, Eduardo P. C.;Ross, Benjamin D.;Durand, Eric
• 通讯作者: Durand, Eric