Acid pH suppression of mycobacterial type-7 ESX-1 protein secretion

酸性 pH 值抑制分枝杆菌 7 型 ESX-1 蛋白分泌

• 批准号: RGPIN-2016-05730
• 负责人: Chen, Jeffrey
• 金额: $ 2.26万
• 依托单位: University of Saskatchewan
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=615872
• 关键词: Acid; pH; suppression; mycobacterial; type

Mycobacterium tuberculosis and Mycobacterium bovis, the agents of human and bovine tuberculosis (TB) respectively have evolved to be exquisitely successful pathogens. Infections caused by these two organisms starts with an acute phase and is followed by a latent phase that can persist for years or decades. Host immunodeficiency and the absence of timely treatment will often lead to symptomatic and fatal disease states. Whilst it is recognized that the establishment of acute and latent TB infection hinges on the pathogen’s ability to sense and respond to changing conditions in the host, there are significant gaps in our knowledge of how this occurs that my research program will seek to address. TB bacteria secrete virulence proteins through its ESX-1 secretion system to thwart the host's immune system and promote infection - mutants unable to do so are significantly attenuated. However, an M. tuberculosis mutant that was found to display increased ESX-1 secretion induced hyper-immune responses in the host and was attenuated during long-term infection. This suggests ESX-1-mediated secretion may be modulated by TB bacteria while it is in the host, likely because some secreted ESX-1 proteins are potent antigens as well as virulence factors. As such, the central hypothesis of this proposal is that TB bacteria senses external cues during infection, and modulates the ESX-1 system to balance its virulence and immune-activating functions. However, the nature and effect of these cues remain poorly defined and the molecular mechanisms underlying them are unknown. In unpublished on-going studies, we discovered that acid pH, an external cue encountered by TB bacteria during infection suppresses the secretion of EsxA, a major ESX-1 substrate, by M. tuberculosis in vitro. Furthermore, a screen of small molecule inhibitors identified potassium clavulanate to be an antagonist of this phenotype, suggesting that its molecular target the beta-lactamase BlaC of TB bacteria may be involved in acid pH suppression of EsxA secretion. We will pursue two main objectives in this proposal: 1) Define the parameters and ESX-1 targets of acid pH-mediated suppression of secretion. 2) Elucidate the molecular mechanism(s) underlying acid pH suppression of M. tuberculosis EsxA secretion through the study of chemical antagonists and genetic mutants. The proposed work will generate new insights into the regulation of TB virulence and provide novel research tools to study TB infections. Ultimately, our findings will be harnessed to develop better tools to combat both human and animal TB infections, thereby benefiting Canadians and the global community. Importantly, my research program will provide excellent training value for microbiology trainees at all levels and will endow them with highly marketable biomedical research skills in the life sciences, whether in academia, the government or the private sector.

结核分枝杆菌（Mycobacterium tuberculosis）和牛分枝杆菌（Mycobacterium bovis）分别是人类和牛结核病（TB）的病原体，它们已经进化成为非常成功的病原体。由这两种微生物引起的感染从急性期开始，然后是潜伏期，可持续数年或数十年。宿主免疫缺陷和缺乏及时治疗往往会导致症状和致命的疾病状态。虽然人们认识到，急性和潜伏性结核病感染的建立取决于病原体感知和应对宿主不断变化的条件的能力，但我们对这种情况如何发生的知识存在重大差距，我的研究计划将寻求解决。 TB细菌通过其ESX-1分泌系统分泌毒力蛋白，以阻碍宿主的免疫系统并促进感染-不能这样做的突变体显着减弱。然而，M。发现显示增加的ESX-1分泌的结核病突变体在宿主中诱导超免疫应答，并且在长期感染期间减弱。这表明ESX-1介导的分泌可能在宿主中受到TB细菌的调节，这可能是因为一些分泌的ESX-1蛋白是有效的抗原以及毒力因子。因此，这一提议的中心假设是，TB细菌在感染期间感知外部线索，并调节ESX-1系统以平衡其毒力和免疫激活功能。然而，这些线索的性质和作用仍然不清楚，其分子机制也不清楚。 在未发表的正在进行的研究中，我们发现，酸性pH值，结核菌感染期间遇到的外部线索，抑制分泌EsxA，一个主要的ESX-1底物，由M。体外培养结核病此外，小分子抑制剂的筛选鉴定出克拉维酸钾是该表型的拮抗剂，表明其分子靶点TB细菌的β-内酰胺酶BlaC可能参与EsxA分泌的酸性pH抑制。我们在这项建议中将追求两个主要目标： 1)定义酸性pH介导的分泌抑制的参数和ESX-1靶标。 2)阐明了M.通过研究结核EsxA分泌的化学拮抗剂和遗传突变体。 拟议的工作将产生对结核病毒力调控的新见解，并为研究结核病感染提供新的研究工具。最终，我们的研究结果将用于开发更好的工具，以对抗人类和动物结核病感染，从而造福加拿大人和全球社会。重要的是，我的研究计划将为各级微生物学学员提供出色的培训价值，并将赋予他们在生命科学领域具有高度市场化的生物医学研究技能，无论是在学术界，政府还是私营部门。