Identification of human host factors involved in Chlamydia development

说明(申请人提供)：衣原体是一种革兰氏阴性细菌病原体，可感染多种宿主并导致各种疾病，包括发展中国家可预防的失明、性传播疾病和肺炎。衣原体在一个膜结合的隔间中进行细胞内复制，该隔间可能是免疫监视的保护罩，但也是与宿主细胞交换因子的“平台”。尽管衣原体作为一种人类病原体非常重要，但人们对感染过程中涉及的细菌和宿主因素知之甚少。这种知识的匮乏主要是由于衣原体不是一种遗传上易驯化的生物，以及在哺乳动物宿主中进行遗传方法的困难。我们建议进行全基因组siRNA筛选，以识别和表征与沙眼衣原体感染入侵后事件有关的人类宿主因素。我们已经证明了该方法在基因组子集上的可行性，并建议将该方法扩展到整个人类基因组(目标1)。由于RNAi治疗已被证明与显示与目标基因有限序列同源性的基因的意外沉默有关，我们将对候选基因进行系统的验证程序，以明确地建立目标基因的敲除与观察到的表型之间的功能关系(目标#2)。最后，我们将进行二次检测，以确定和表征沙眼衣原体入侵后细胞内发育事件中涉及的衣原体特定候选者(目标#3)。总体而言，这一发展建议可能为衣原体致病机理提供新的见解，并为未来与沙眼衣原体感染相关的已知和/或新途径的表征相关的资金申请(S)产生所需的初步结果。 与公共卫生相关：衣原体是导致沙眼、性传播疾病和肺炎等各种疾病的专性细胞内细菌病原体。我们研究衣原体如何劫持细胞成分，以便在宿主内成功复制和传播。我们通过确定衣原体发育所需的宿主因素，重点剖析了感染过程中涉及的细胞机制(S)。识别这些因素可能有助于设计新的治疗方法。

DESCRIPTION (provided by applicant): Chlamydia are Gram-negative bacterial pathogens that infect a wide range of hosts and cause various diseases, including preventable blindness in developing countries, sexually transmitted disease and pneumonia. Chlamydia replicate intracellularly in a membrane-bound compartment that potentially serves as a protector shield against immune surveillance but also acts as a "platform" to exchange factors with the host cell. Despite the primary importance of Chlamydia as a human pathogen, little is known about the bacterial and host factors involved in the infection process. This paucity of knowledge is mainly due to the fact that Chlamydia is not a genetically tractable organism and to the difficulty of conducting genetic approaches in the mammalian host. We propose to conduct a genome-wide siRNA screen in order to identify and characterize human host factors involved in post-invasion events of Chlamydia trachomatis infection. We have demonstrated the feasibility of the approach on a subset of the genome and we propose to extend the approach to the entire human genome (Aim#1). Since RNAi treatment has been shown to be associated with the unintended silencing of genes displaying limited sequence homology with the targeted gene, we will perform a systematic validation procedure of the candidates to unambiguously establish a functional relationship between the knock-down of the targeted gene and the observed phenotype (Aim#2). Finally, we will conduct secondary assays to identify and characterize Chlamydia specific candidates involved in post-invasion events of C. trachomatis intracellular development (Aim#3). Overall, this developmental proposal is likely to provide new insights in Chlamydia pathogenesis and to generate the required preliminary results for future funding application(s) related to the characterization of known and/or novel pathway(s) involved in C. trachomatis infection. PUBLIC HEALTH RELEVANCE: Chlamydia are obligate intracellular bacterial pathogens responsible for various diseases such as trachoma, sexually transmitted disease and pneumoniae. We study how Chlamydia hijack cellular components to successfully replicate and disseminate inside the host. We focus on dissecting the cellular mechanism(s) invloved in the infection process by identifying the host factors required for Chlamydia development. The identification of these factors may help design new therapeutic treatments.