Inhibition of T Cell Responses by Bacteria

描述（由申请人提供）：我们已经开始研究CD8+ T细胞在免疫细胞内细菌病原体福氏志贺氏菌中的作用。在感染过程中，弗氏柔球菌进入细胞并逃逸到宿主细胞质中。我们预计从flexneri s.f eri分泌的蛋白质会被MHC-I加工机制降解，并且产生的肽会在表面MHC-I分子的背景下呈递给CD8+ T细胞。单核细胞增生李斯特菌（Listeria monocytogenes）是另一种逃逸到宿主细胞质中的细胞内细菌病原体，在感染李斯特菌衍生的肽与宿主MHC-I复合物时，可被CD8+ T细胞识别，并有助于产生保护性免疫。虽然CD8+ T细胞已被证明对单核增生乳杆菌的保护性免疫是必需的，但我们没有发现证据表明CD8+ T细胞在对弗氏沙门氏菌的保护性免疫中起作用。即使我们设计了flexneri，使其组成性地分泌能刺激CD8+ T细胞反应的异源表位，也没有检测到这些反应。此外，我们发现当培养细胞被表位标记的flexneri感染时，这些细胞不能被已建立的表位标签特异性T细胞克隆识别。我们之前的研究结果表明，flexneri抑制MHC-I限制性抗原向CD8+ T细胞的呈递，正是这种抗原呈递的缺乏导致CD8+ T细胞对flexneri分泌到宿主细胞质中的抗原反应失败。本实验旨在识别和表征弗氏杆菌感染期间发生的MHC-I加工和/或表现的缺陷。具体来说：1)我们将使用基因筛选来鉴定负责抑制的flexneri基因产物；2)我们将使用生化分析来确定MHC-I加工途径中哪一步或哪几个步骤在感染福氏沙门氏菌期间发生改变，从而导致宿主细胞表面抗原呈递减少。通过这些实验，我们期望识别和描述抗原加工和呈递的细菌抑制剂的活性。这种抑制剂可能代表了一类新的毒力决定因素，能够特异性地干扰保护性T细胞介导的适应性免疫的发展。了解这种抑制如何影响细菌毒力和免疫清除之间的平衡将进一步加深我们对这种细菌病原体及其哺乳动物宿主的复杂相互作用的理解。公共卫生相关性：福氏志贺氏菌是全世界严重胃肠道疾病的罪魁。我们发现这种生物有能力抑制宿主针对它的免疫反应。本应用程序中提出的工作旨在了解生物体能够避免检测的机制，并确定生物体中哪些基因负责免疫系统抑制。我们的工作可能会导致治疗策略，可能会破坏这种生物体躲避免疫系统的能力，从而允许更有效地清除细菌。

DESCRIPTION (provided by applicant): We have initiated studies to characterize the role of CD8+ T cells in immunity to the intracellular bacterial pathogen, Shigella flexneri. During infection, S. flexneri enter cells and escape into the host cell cytosol. We expected that proteins excreted from S. flexneri would be proteolytically degraded by the MHC-I processing machinery, and that the resulting peptides would be presented to CD8+ T cells in the context of surface MHC-I molecules. During infection with Listeria monocytogenes, another intracellular bacterial pathogen that escapes into the host cell cytosol, Listeria-derived peptides, in complex with host MHC-I, are recognized by CD8+ T cells and contribute to the generation of protective immunity. While CD8+ T cells have been shown to be required for protective immunity to L. monocytogenes, we have found no evidence to suggest that CD8+ T cells play a role in protective immunity to S. flexneri. Even when we engineered S. flexneri to constitutively secrete heterologous epitopes known to stimulate potent CD8+ T cell responses, those responses were not detected. Furthermore, we found that when cultured cells were infected with epitope-tagged S. flexneri, these cells were not recognized by established T cell clones specific for the epitope tag. Our results over the previous award period suggest that S. flexneri inhibit the presentation of MHC-I restricted antigens to CD8+ T cells, and it is this lack of antigen presentation that results in the failure of CD8+ T cells to respond to antigens secreted into the host cell cytosol by S. flexneri. The experiments in this proposal seek to identify and characterize the defect in MHC-I processing and/or presentation that occurs during S. flexneri infection. Specifically: 1) we will use genetic screens to identify S. flexneri gene product(s) responsible for the inhibition; and 2) we will use biochemical assays to determine the step or steps in the MHC-I processing pathway that are altered during infection with S. flexneri and that result in decreased antigen presentation on the host cell surface. Through these experiments, we expect to identify and describe the activity of a bacterial inhibitor of antigen processing and presentation. Such an inhibitor might represent a novel class of virulence determinants specifically able to interfere with development of protective T cell-mediated adaptive immunity. Understanding how this inhibition affects the balance between bacterial virulence and immune clearance will further our understanding of the complex interaction of this bacterial pathogen and its mammalian host. PUBLIC HEALTH RELEVANCE: Shigella flexneri is responsible for serious gastrointestinal disease throughout the world. We have found that this organism has the capacity to inhibit the immune response directed against it by the host. The work proposed in this application is directed at understanding the mechanisms through which the organism is able to avoid detection and identifying which genes in the organism are responsible for immune system inhibition. Our work may lead to therapeutic strategies that might undermine the ability of this organism to hide from the immune system, thereby allowing more effective clearance of the bacteria.