TYPE II SECRETION AND LEGIONELLA PNEUMOPHILA INFECTION

描述(摘自申请者摘要)：军团菌 嗜肺性肺炎是引起退伍军人病的主要原因之一。 既有社区获得性肺炎，也有医院获得性肺炎 占肺炎总数的30%。这种细菌是一种胞内寄生虫。 人类肺泡巨噬细胞和水生原生动物。我们有 发现了一个在嗜肺乳杆菌中起主要作用的基因座(PilBCD) 生理学和致病力。最重要的是，这个基因座编码一种蛋白质 (PilD)，类似于IV型菌毛中涉及的多肽酶 革兰氏阴性菌的生物发生和蛋白质(II型)分泌。 PilD的一个突变使菌毛的产生消失，表明L. 嗜肺性肺炎表现为IV型菌毛。更重要的是，PilD 突变型，但不是菌毛素缺乏菌株，在 其感染阿米巴和巨噬细胞的能力，表明II型 分泌是细胞内生长所必需的。在学习L. 在嗜肺性肺炎菌中表达，我们发现其转录 PilBCD基因座受温度调控，其表达受温度调控 军团菌IV型菌毛受温度和二氧化碳的共同调节。至 进一步了解II型分泌物和细胞内 寄生，我们将i)确定温度、二氧化碳和生长阶段 并分离出相应的调节因子 元素，ii)使用遗传、免疫学和生化筛查， 确定依赖PilD的分泌蛋白促进细胞内 感染，以及iii)阐明细胞内的贩运事件 是由PilD和II型分泌蛋白协调的。结果是 这次调查不仅将增进我们对 军团菌病，但将作为研究其他 细胞内的病原体。

DESCRIPTION (Adapted from the applicants Abstract): Legionella pneumophila, the agent of Legionnaire=s disease, is a major cause of both community-acquired and nosocomial pneumonia and can account for up to 30% of all pneumonias. This bacterium is an intracellular parasite of both human alveolar macrophages and aquatic protozoa. We have discovered a locus (pilBCD) that plays a major role in L. pneumophila physiology and virulence. Most importantly, this locus encodes a protein (PilD) that is analogous to peptidases involved in type-IV pilus biogenesis and protein (type II) secretion in gram-negative bacteria. A mutation in pilD abolished pilus production, indicating that L. pneumophila expresses type-IV pili. Of greater significance, the pilD mutant, but not a pilin deficient strain, was dramatically impaired in its ability to infect amoebae and macrophages, indicating that type II secretion is required for intracellular growth. While studying L. pneumophila pilBCD expression, we discovered that the transcription of the pilBCD locus is regulated by temperature and that the expression of Legionella type-IV pili is modulated by both temperature and CO2. To further our understanding of both type II secretion and intracellular parasitism, we will I) determine how temperature, CO2, and growth phase modulated pilD expression and isolate the corresponding regulatory elements, ii) using genetic, immunologic , and biochemical screens, identify the PilD-dependent secreted proteins that promote intracellular infection, and iii) elucidate the intracellular trafficking events that are orchestrated by PilD and type II secreted proteins. The results of this investigation will not only enhance our understanding of legionellosis but will serve as a model for the study of other intracellular pathogens.