Modulation of Host Vesicle Trafficking by Legionella pneumophila

描述（由申请人提供）：嗜肺军团菌，军团病的病原是一种细胞内病原体。其致病性的关键是Dot/Icm IV型分泌系统，该系统将大量细菌效应物转运到宿主细胞中。这些效应物调节多种宿主细胞过程，创造一个内质网（ER）衍生的细胞器，允许军团菌在细胞内生长。我们最近发现，其中一种被称为SidJ的效应物在复制性含军团菌液泡（LCVs）的生物发生中起着重要作用，而SidJ的缺失会导致ER蛋白向LCVs募集的时间延迟。进一步的研究表明，SidJ是一种蛋白磷酸酶，可与copii包被囊泡的外壳复合物的主要亚基Sec31相互作用并使其去磷酸化。我们的工作假设是，SidJ通过与其他效应物协调，将copii包被囊泡的运输重定向到细菌吞噬体。本项目的目的是利用遗传学、生物化学和细胞生物学的方法来表征军团菌感染中SidJ及其蛋白磷酸酶的活性。我们建议：1)分析SidJ与Sec31的相互作用以及SidJ对宿主分泌途径的影响。2)研究SidJ-Sec31相互作用对COPII包被囊泡形成的功能影响。3)与SidJ协同作用的效应物的鉴定与表征。这项研究的结果将有助于新兴的细胞微生物学领域和细胞生物学领域，特别是COPII成分的蛋白磷酸化在调节囊泡运输中的作用。这些研究也将扩大我们的研究能力，从而使我们能够建立一个具有更广泛项目的研究计划。获得K02奖可以让我投入更多的精力来发展我的研究事业，通过探索和获取在这个多学科领域取得成功所必需的背景知识和新技术。

DESCRIPTION (provided by applicant): Legionella pneumophila, the etiological agent of Legionnaires' disease is an intracellular pathogen. Essential to its pathogenicity is the Dot/Icm type IV secretion system that translocates a large number of bacterial effectors into host cell. These effectors modulate multiple host cell processes to create an endoplasmic reticulum (ER) derived organelle permissive for Legionella intracellular growth. We recently found that one of such effectors called SidJ plays an important role in the biogenesis of replicative Legionella-containing vacuoles (LCVs) and loss of SidJ results in a temporal delay in the recruitment of ER proteins to the LCVs. Further studies indicate that SidJ is a protein phosphatase that interacts with and dephosphorylates Sec31, the major subunit of the outer shell complexes of the COPII-coated vesicles. Our working hypothesis is that SidJ, by coordinating with other effectors, redirects the trafficking of COPII-coated vesicles to the bacterial phagosomes. The goal of this project is to characterize SidJ and its protein phosphatase activity in Legionella infection using genetic, biochemical and cell biological approaches. We propose to: 1) Analyzing the interactions between SidJ and Sec31 and the effect of SidJ on host secretory pathways. 2) Studying the functional consequence of SidJ-Sec31 interaction on COPII coated vesicle formation. 3) Identification and characterization of effectors that function synergistically with SidJ. Results from this study will contribute to the emerging field of cellular microbiology and to the field of cell biology, particularly the roles of protein phosphorylation of COPII components in the regulation of vesicle trafficking. These studies will also expand our research capacity, thus allowing us to establish a research program with a broader spectrum of projects. A K02 award would allow me to invest more effort to develop my research career by exploring and acquiring background knowledge and new techniques necessary for future success in this multidisciplinary field. Relevance: Because of its potential fatality, infection by Legionella pneumophila is a concern of public health. Our study can provide novel strategies for its prevention and/or treatment. Furthermore, understanding of mammalian cell processes affected by this bacterium could lead to clues in our study of other diseases associated with abnormality of these processes. Project Narrative: Infection by Legionella pneumophila often leads to the development of a fatal form of pneumonia. In addition, study on this bacterium could share light on our understanding of other intracellular pathogens because L. pneumophila and these infectious agents share many pathogenic mechanisms.