Project 1: Listeria metabolites and innate immunity

项目摘要/摘要(项目1，Portnoy) 这项提案是P01更新中的项目1，题为“先天免疫和获得性免疫的交集” 细胞内的病原体。“我们要解决的一个中心问题是细胞内病原体是如何被识别的 通过宿主的先天免疫系统以及多种信号是如何整合来诱导适当的 反之，病原体如何避免和/或操纵寄主反应以促进其 发病机制。在项目1中，我们选择通过继续详细分析 单核细胞增多性李斯特菌，一种兼性的细胞内、食源性病原体，已被研究了数十年 作为一个模型系统，用来剖析感染和免疫的基本方面。之前，我们发现 单核细胞增多性乳杆菌分泌c-diamp，c-diamp与宿主刺痛结合并激活，导致I型 干扰素反应。在目标1中，我们开发了一种新开发的李斯特菌病口服模型，在那里我们发现 细菌分泌的c-di-AMP诱导一种依赖于刺的宿主保护反应。我们建议 分析STING如何提供保护，并检验STING也促进传播到 胎盘和大脑。在目标2和目标3中，我们开始探索c-di-amp可能只是一个 小细菌代谢物在形成宿主免疫中起重要作用的原理的例子。L. 单核细胞增多症是一种需要核黄素的微生物，我们探索了这样的假设 营养要求允许细菌避免粘膜相关不变T细胞(MAIT)的激活 细胞)，这是由核黄素生物合成的修饰中间体刺激的先天T细胞。在……里面 初步数据，我们将5个枯草芽孢杆菌基因(RiDEAHT)导入单核细胞增多性乳杆菌中进行转化 转化成一种核黄素合成菌，现在准备检验我们的假设并了解它的作用 在细菌感染和免疫期间MAIT细胞的数量。我们还建议探讨如何激活 人类占优势的伽马三角洲T细胞(Vg9Vd2 T细胞)。这些细胞被一种中间体激活 细菌类异戊二烯生物合成(HMBPP)。我们的最终目标是确定这种与生俱来的重要性 灵长类动物(人和非人)感染期间的免疫途径。在这里，我们建议构建 细菌突变体，要么不能制造HMBPP，要么制造更多HMBPP来检测HMBPP的细菌含量 在组织培养模型的感染过程中检测到。我们还建议与考克斯实验室合作 (项目2)探讨HMBPP在结核分枝杆菌感染和免疫中的重要性。

Project Summary/Abstract (Project 1, Portnoy) This proposal is Project 1 within a P01 renewal, entitled “The intersection of innate and adaptive immunity to intracellular pathogens.” A central problem that we address is how intracellular pathogens are recognized by the host innate immune system and how multiple signals are integrated to induce an appropriate response, and conversely, how pathogens avoid and/or manipulate host responses to promote their pathogenesis. In Project 1, we have chosen to approach this problem by continuing a detailed analysis of Listeria monocytogenes, a facultative intracellular, food-borne pathogen that has been studied for decades as a model system with which to dissect basic aspects of infection & immunity. Previously, we discovered that L. monocytogenes secretes c-di-AMP which binds to and activates host STING leading to a type I interferon response. In Aim 1, we exploit a newly developed oral model of listeriosis where we find that bacterially secreted c-di-AMP induces a STING-dependent host protective response. We propose to analyze how STING provides protection and test the hypothesis that STING also promotes dissemination to the placenta and brain. In Aims 2 and 3, we begin to explore the idea that c-di-AMP might be just one example of the principle that small bacterial metabolites play important roles in shaping host immunity. L. monocytogenes is a riboflavin requiring microorganism, and we explore the hypothesis that that this nutritional requirement allows the bacteria to avoid activation of mucosal-associated invariant T-cells (MAIT cells), which are innate T-cells stimulated by a modified intermediate of riboflavin biosynthesis. In preliminary data, we introduced five Bacillus subtilis genes (ribDEAHT) into L. monocytogenes converting it into a riboflavin-synthesizing bacterium and are now poised to test our hypothesis and learn about the role of MAIT cells during bacterial infection and immunity. We also propose to explore the activation of the dominant gamma delta T-cell in humans (Vg9Vd2 T-cells). These cells are activated by an intermediate of bacterial isoprenoid biosynthesis (HMBPP). Our ultimate goal is to determine the importance of this innate immune pathway during infection of primates (human and non-human). Here we propose to construct bacterial mutants that either fail to make HMBPP or make more of it to examine how bacterial HMBPP is detected during infection in tissue culture models. We further propose to collaborate with the Cox lab (Project 2) to explore the importance of HMBPP during M. tuberculosis infection and immunity.