Chlamydia Effector Proteins

描述（由申请方提供）：专性细胞内细菌病原体沙眼衣原体感染眼和生殖器上皮，引起具有重要临床意义的疾病。C.沙眼衣原体通过将“效应”蛋白质跨宿主和包涵体膜转运来调节宿主细胞功能。缺乏工具来进行经典的分子遗传学分析衣原体，阻碍了识别和表征效应蛋白的进展。然而，鉴于其基因组的小尺寸，基于表达的功能方法可以被带到基因组规模，并显着加速衣原体发病机制的研究。在这里，我们建议开发功能基因组工具来识别和表征效应蛋白和衣原体感染过程中效应蛋白输出的动态。在我们的第一个目标中，我们建议产生全面的酵母和哺乳动物表达文库，以系统地调查衣原体蛋白的致病性相关的功能，并应用基于免疫学的屏幕，以确定新的分泌衣原体蛋白。这些发现将与衣原体初级体的蛋白质组数据合并，以确定和表征介导新生致病性空泡进入和生物发生的细菌因子。在我们的第二个目标中，我们扩展了功能工具的应用，以表征衣原体III型分泌（TTS）系统，传递效应蛋白的主要门户。通过双杂交技术和生物化学方法，我们建议构建一个蛋白质-蛋白质相互作用图的祖先TTS装置，以确定非保守的组件的核心易位，针复合物和推定的调节亚基。特别是，我们将把我们的研究重点放在衣原体特异性蛋白质的TSS系统的基础结构，包括一个新的TTS分子伴侣，我们已经确定和它的同源货物相互作用。 公共卫生相关性：广泛传播的病原体沙眼衣原体破坏免疫反应的程度取决于它们注入宿主细胞的毒力因子（“效应”蛋白）。拟议的研究将开发基因组工具来识别和表征这些效应蛋白。通过这种方式，我们将大大加深我们对衣原体发病机制的理解，并为疫苗设计和治疗干预产生新的靶点。

DESCRIPTION (provided by applicant): The obligate intracellular bacterial pathogen Chlamydia trachomatis infects the ocular and genital epithelium to cause diseases of significant clinical importance. C. trachomatis modulates host cellular functions by translocating "effector" proteins across host and inclusion membranes. The lack of tools to perform classical molecular genetic analysis in Chlamydia, has hampered progress in identifying and characterizing effector proteins. However, given the small size of the its genome, expression-based functional approaches can be brought to genomic scale and significantly accelerate research in Chlamydia pathogenesis. Here, we propose to develop functional genomic tools to identify and characterize effector proteins and the dynamics of effector protein export during chlamydial infection. In our first aim, we propose to generate comprehensive yeast and mammalian expression libraries to systematically survey chlamydial proteins for functions related to their pathogenicity and apply immunological-based screens to identify novel secreted chlamydial proteins. These findings will be merged with proteomic data of Chlamydia elementary bodies to identify and characterize bacterial factors mediating entry and biogenesis of the nascent pathogenic vacuole. In our second aim, we extend the application of functional tools to characterize the chlamydial Type III secretion (TTS) system, the main portal for the delivery of effector proteins. Through two-hybrid technologies and biochemical approaches we propose to construct a protein-protein interaction map of this ancestral TTS apparatus to identify non-conserved components of the core translocon, needle complex and putative regulatory subunits. In particular, we will focus our studies on Chlamydia-specific proteins that interact with the basal structure of the TSS system, including a novel TTS chaperone we have identified and its cognate cargo. PUBLIC HEALTH RELEVANCE: The extent to which the widely disseminated pathogen bacteria Chlamydia trachomatis subverts immune responses is determined by virulence factors they inject into host cells ("effector" proteins). The proposed study will develop genomic tools to identify and characterize these effector proteins. In this manner we will significantly further our understanding of chlamydial pathogenesis and generate new targets for vaccine design and therapeutic intervention.