Genetic determinants of susceptibility to mycobacterial infections

描述（由申请人提供）：传染病是一个主要的，重新出现的健康问题，占世界死亡率的33%（1700万/年）。新的致命变种的出现、对抗生素的广泛耐药性以及世界范围内旅行的增加只是增加问题严重性的几个因素，这些因素不仅在发展中国家，而且对前往这些地区的人以及免疫功能降低的人也是如此。分枝杆菌感染仍然是一个严重的健康威胁，在其他健康的个体中引起结核病，并在免疫抑制的患者中引起播散性感染。宿主对这些感染的防御机制，包括巨噬细胞的效应功能和保护性免疫的参数，需要更好地理解。这种宿主防御机制可以表现为人类群体和相应动物模型中对感染的先天抗性或易感性的遗传决定因素。在小鼠中使用遗传方法，我们已经检测到7号染色体上的一个主要位点，命名为Trl 3，它控制小鼠对肺结核的易感性。在本申请的第一个具体目标中，我们提出鉴定控制这种差异易感性的基因和蛋白质。这将通过创建和表征一系列同源和亚同源小鼠品系来完成，其中Trl 3已通过选择性育种分离。将仔细检查最小遗传间隔是否存在位置候选物，然后在体内转基因小鼠和体外巨噬细胞中系统地检测位置候选物。该基因的蛋白产物的功能及其在巨噬细胞和树突状细胞防御结核病中的作用将被研究。独立地，我们已经发现转录因子IRF 8的突变导致小鼠对肺结核的极端易感性，并且已经在患有孟德尔易感性分枝杆菌病（MSMD）的患者中鉴定了纯合子功能丧失IRF 8突变。在本申请的第二个目的中，我们提出实施全基因组方法，包括用微阵列和染色质免疫沉淀（ChIP）和基因组DNA阵列（ChIP on chip）进行转录谱分析，以系统地鉴定由IRF 8调控的基因，并且其可能编码巨噬细胞对分枝杆菌的关键防御机制。还提出了一系列体外和体内实验来验证IRF 8对所鉴定的靶标的调节，并评估它们在巨噬细胞防御分枝杆菌中的作用。我们还建议通过监测300例MSMD患者队列中这些靶点的完整性来评估这些靶点在人类分枝杆菌感染中的作用。 公共卫生相关性：传染病是一个主要的、重新出现的健康问题，占世界死亡率的33%（每年1700万人）。分枝杆菌感染仍然是一个严重的健康威胁，在其他健康的个体中引起结核病，并在免疫抑制的患者中引起播散性感染。宿主对这些感染的防御机制，包括巨噬细胞的抗分枝杆菌效应功能和保护性免疫的参数，需要更好地理解。这种宿主防御机制可以表现为人类群体和相应动物模型中对感染的先天抗性或易感性的遗传决定因素。所涉及的基因的鉴定和表征不仅能够更好地理解感染的发病机制和宿主对其的反应，而且还可以确定药物发现和药理学干预这些感染的新靶点。

DESCRIPTION (provided by applicant): Infectious diseases are a major, re-emerging health problem, accounting for 33% of the world mortality (17 million/year). The emergence of new virulent variants, widespread resistance to antibiotics, and increased worldwide travel are but a few factors that have increased the severity of the problem not only in developing countries, but also for people traveling to these regions, and for individuals with reduced immune functions. Mycobacterial infections continue to be a serious health threat, causing tuberculosis in otherwise healthy individuals, and causing disseminated infections in immuno-suppressed patients. The host defense mechanisms against these infections, including the effector functions of macrophages and parameters of protective immunity, need to be better understood. Such host defense mechanisms can manifest themselves as genetic determinants of innate resistance or susceptibility to infections in human populations, and in corresponding animal models. Using a genetic approach in mice, we have detected a major locus on chromosome 7, designated Trl3, which controls susceptibility to pulmonary tuberculosis in mice. In the first specific aim of this application, we propose to identify the gene and protein that control this differential susceptibility. This will be done by creating and characterizing a series of congenic and subcongenic mouse strains, in which Trl3 has been isolated by selective breeding. The minimal genetic interval will be carefully scrutinized for the presence of positional candidates that will then be systematically tested in transgenic mice in vivo and in macrophages in vitro. The function of the protein product of the gene and its role in macrophage and dendritic cell defenses against tuberculosis will be investigated. Independently, we have uncovered that a mutation in the transcription factor IRF8 causes extreme susceptibility to pulmonary tuberculosis in mice, and have identified a homozygote loss-of-function IRF8 mutation in a patient suffering from Mendelian Susceptibility to Mycobacterial Disease (MSMD). In the second aim of this application we propose to implement genome-wide methods, including transcript profiling with microarrays and chromatin immunoprecipitation (ChIP) and genomic DNA arrays (ChIP on chip) to systematically identify genes regulated by IRF8, and that may code for key defense mechanisms of macrophages against mycobacteria. A series of in vitro and in vivo experiments are also proposed to validate IRF8 regulation of the identified targets, and to evaluate their role in macrophage defenses against mycobacteria. We also propose to assess the role of these targets in human mycobacterial infections, by monitoring their integrity in a cohort of 300 patients suffering from MSMD. PUBLIC HEALTH RELEVANCE: Infectious diseases are a major, re-emerging health problem, accounting for 33% of the world mortality (17 million/year). Mycobacterial infections continue to be a serious health threat, causing tuberculosis in otherwise healthy individuals, and causing disseminated infections in immuno-suppressed patients. The host defense mechanisms against these infections, including the anti-mycobacterial effector functions of macrophages and parameters of protective immunity, need to be better understood. Such host defense mechanisms can manifest themselves as genetic determinants of innate resistance or susceptibility to infections in human populations, and in corresponding animal models. The identification and characterization of the genes involved will not only enable a better understanding of infection pathogenesis and host response to it, but may also identify novel targets for drug discovery and pharmacological interventions in these infections.