Probing novel innate immune detection mechanisms using an intracellular bacterial

描述(申请人提供)：免疫学中的一个基本问题是免疫细胞如何识别并对入侵的微生物做出强有力的反应。能够在免疫细胞中引发不同反应的遗传上可处理的病原体在确定负责这种反应的微生物配体或活性以及相应的宿主免疫监视机制方面非常有用。嗜肺军团菌是一种普遍存在的细菌，在环境中主要与阿米巴作为一种细胞内寄生虫有关。这种细菌还可以在哺乳动物的肺泡巨噬细胞内复制，并导致军团病，一种严重的炎症性肺炎。阿米巴宿主被认为为其获得和维持细胞内复制所需的细菌特征提供了主要或唯一的进化压力。因此，嗜肺性乳杆菌似乎没有进化出复杂的免疫逃避机制，可以作为识别宿主免疫系统感知的细菌配体或活性的极好模型。我们发现，一株环境分离的嗜肺乳杆菌(菌株LPE509)不能在A/J小鼠的骨髓来源的巨噬细胞中复制，这些巨噬细胞允许大多数(如果不是全部)临床分离的嗜肺乳杆菌临床分离株在细胞内复制。菌株LPE509在阿米巴宿主盘基网眼菌和分化的人巨噬细胞U937细胞中旺盛生长，这一过程依赖于Dot/ICM IV型分泌素系统，而Dot/ICM IV型分泌素系统是所有所研究的嗜肺性菌株毒力所必需的。此外，我们发现LPE509在需要Dot/ICM转运蛋白而不需要鞭毛蛋白或NAIP5-NLRC4炎症体的过程中诱导了A/J巨噬细胞的广泛细胞死亡。因此，我们假设菌株LPE509编码一个或多个可被宿主免疫监视系统识别的因素，导致宿主细胞死亡。在这个提议中，我们将通过追求两个具体的假设来检验这个假设 目的：首先，我们将鉴定导致A/J巨噬细胞死亡的细菌因子(S)。其次，我们将确定参与识别细菌信号的宿主途径(S)。

DESCRIPTION (provided by applicant): A fundamental question in immunology is how immune cells recognize and mount a robust response to invading microorganisms. Genetically tractable pathogens capable of triggering distinct responses in immune cells are extremely useful in determining the microbial ligands or activity responsible for such responses and the corresponding host immune surveillance mechanisms. Legionella pneumophila is a ubiquitous bacterium, which in the environment is primarily associated with amoebae as an intracellular parasite. This bacterium also can replicate within mammalian alveolar macrophages and causes Legionnaires' disease, a severe inflammatory pneumonia. The amoebae host is believed to provide the primary or exclusive evolutionary pressure for its acquisition and maintenance of bacterial traits necessary for intracellular replication. Thus, L. pneumophila appears not to have evolved sophisticated immune-evasive mechanisms and can serve as an excellent model for identifying bacterial ligands or activity sensed by the host immune system. We found that an environmental isolate of L. pneumophila (strain LPE509) was unable to replicate in bone marrow-derived macrophages from A/J mice, which are permissive for intracellular replication by most, if not all examined clinic L. pneumophila isolates. Strain LPE509 grows robustly in the amoebae host Dictyostelium discoideum and the differentiated human macrophages U937 cells in a process that is dependent upon the Dot/Icm type IV secretin system, which is required for the virulence of all examined L. pneumophila strains. Further, we found that LPE509 induced extensive cell death in A/J macrophages in a process that required the Dot/Icm transporter but not flagellin or the NAIP5-NLRC4 inflammasome. Thus, we hypothesized that strain LPE509 codes for one or more factors that can be recognized by the host immune surveillance system, leading to host cell death. In this proposal, we will test this hypothesis by pursuing two specific Aims: First, we will identify the bacterial factor(s) responsible for the induction of cell death i A/J macrophages. Second, we will determine the host pathway involved in the recognition of the bacterial signal(s).