The interaction of Chlamydia with the host cytoskeleton

描述(由申请人提供)：专性细胞内沙眼衣原体是世界上导致可预防的失明的主要原因，也是人类最常见的性传播细菌病原体。在美国，每年新发的衣原体生殖道感染病例约为400万例，导致盆腔炎等严重疾病导致输卵管性不孕。衣原体复制发生在宿主细胞内称为包涵体的独特的膜结合室中。包涵体成熟的最早步骤之一是向心迁移到核周区域，这一过程在所有衣原体物种中都是保守的，表明在发病机制中发挥了重要作用。这种迁移是由衣原体驱动的，因为衣原体蛋白的合成是微管运动蛋白Dynein募集所必需的。衣原体包涵体运输到细胞的微管组织中心(MTOC)需要动力蛋白的募集和激活，这最终导致包涵体与宿主中心体之间的密切联系。成熟的衣原体包涵体仍然与中心体有关，导致中心体数量缺陷、纺锤体缺陷和染色体不稳定。我们假设，在衣原体感染过程中，动力蛋白的募集和激活是发病的一个中心机制。这项建议的重点是确定参与动力蛋白激活的衣原体蛋白，并研究动力蛋白和中心体在细胞间扩散和细胞转化中的作用。目的1利用动力蛋白免疫沉淀、中心体纯化和微管分离技术鉴定与动力蛋白结合的衣原体蛋白。中心体是动态细胞器，在细胞分裂过程中活跃地分裂成子细胞，因此目标2将研究动力蛋白和中心体在持续和非持续生长条件下衣原体在细胞间传播中的作用。中心体功能是维持染色体保真度的关键。衣原体和包涵体之间独特的衣原体驱动的相互作用可能是衣原体感染和宫颈癌之间联系的一个重要因素。目的3确定衣原体引起中心体缺陷的机制，并评价这些缺陷在细胞转化中的作用。此外，宫颈癌和衣原体之间的联系可能涉及与人类乳头瘤病毒(HPV)感染的相互作用。因此，我们将评估衣原体引起的中心体缺陷和高危HPV16E6和E7癌基因表达之间的相互作用在细胞转化中的作用。公共卫生相关性：据估计，美国每年有400万人感染衣原体。感染这种细菌会导致盆腔炎、妇女不孕不育，并增加患宫颈癌的几率。确定衣原体在细胞内发育的机制及其对宿主细胞复制机制的影响将有助于对衣原体疾病的了解，并提供潜在的新的治疗靶点。

DESCRIPTION (provided by applicant): The obligate intracellular bacterium Chlamydia trachomatis is the world's leading cause of preventable blindness and the most common sexually transmitted bacterial pathogen of humans. In the US, the incidence of new cases of chlamydial genital tract infection is approximately 4 million annually, causing severe illness such as pelvic inflammatory disease leading to tubal infertility. Chlamydia replication takes place in a unique membrane bound compartment within the host cell termed the inclusion. One of the earliest steps in inclusion maturation is centripetal migration to the perinuclear region, a process that is conserved among all chlamydial species suggesting an important role in pathogenesis. This migration is chlamydial driven, as chlamydial protein synthesis is required for the recruitment of the microtubule motor protein dynein. Dynein recruitment and activation is required for chlamydial inclusion trafficking to the microtubule organizing center (MTOC) of the cell, which ultimately leads to an intimate association between the inclusion and the host centrosome. The mature chlamydial inclusion remains associated with the centrosomes causing centrosome number defects, spindle defects, and chromosome instability. We hypothesize that the recruitment and activation of dynein is a central mechanism of pathogenesis during chlamydial infection. The focus of this proposal is to identify the chlamydial proteins involved in dynein activation, and investigate the role of dynein and centrosomes in cell to cell spread and cellular transformation. Aim 1 will identify chlamydial proteins that bind to dynein using dynein immunoprecipitation, centrosome purification and microtubule isolation techniques. Centrosomes are dynamic organelles and are actively partitioned into daughter cells during cell division, therefore Aim 2 will investigate the role of dynein and the centrosome in cell-to-cell spread of Chlamydia under persistent and non-persistent growth conditions. Centrosome function is crucial in the maintenance of chromosome fidelity. The unique chlamydial driven interaction between dynein and the inclusion is likely an important factor in the link between chlamydial infection and cervical cancer. Aim 3 will determine the mechanism of chlamydial induced centrosome defects and evaluate these defects in cellular transformation. Additionally, the link between cervical cancer and Chlamydia likely involves an interaction with human papilloma virus (HPV) infection. We will therefore assess the role of the interaction between chlamydial induced centrosome defects and expression of the high risk HPV16 E6 and E7 oncogenes on cellular transformation. PUBLIC HEALTH RELEVANCE: The bacteria Chlamydia infects an estimated 4 million people annually in the United States. Infection with this organism leads to pelvic inflammatory disease, infertility in women and raises the chance of cervical cancer development. Determining the mechanisms of chlamydial intracellular development and its impact on host cell replication machinery will lead to advances in understanding chlamydial disease and offer potential novel therapeutic targets.