Regulation of Chlamydia trachomatis development via the stringent response

 描述(申请人提供)：衣原体属细菌是专性的细胞内细菌寄生虫。沙眼衣原体是一种人类适应的物种，分为几个血清学上不同的变种，导致血清特异性疾病。生殖器血清型为C. 沙眼衣原体每年在全世界造成约1亿例性传播感染，眼部血清型是发展中国家可预防失明的主要原因。所有衣原体都必须在两种末端分化的细胞形式之间过渡，才能完成它们的生命周期。在感染的非复制基本体(EB)进入宿主细胞后，有机体过渡到复制的非感染性网状小体(RB)。其他在发育形式之间转换的病原菌可以通过严格反应(SR)作为分化调节因子对营养供应做出反应。典型的SR是由RNA聚合酶调节剂四磷酸鸟苷(PpGpp)(也被称为丙酮)和转录因子DksA的合成启动的。由于dks A的缺失会扰乱细菌发育的分化，所以在沙眼衣原体中的表达与sR在沙眼衣原体发育中的作用是一致的。沙眼衣原体基因组只有1.04Mb，缺乏几种预测的关键酶和完整的代谢途径，因此必须从宿主细胞获得大量的营养物质，包括氨基酸和核苷酸。拟议的研究基于这样的假设，即沙眼衣原体的发育受到通过SR获得的营养的调节，并且EB的产生与细胞内养分可获得性的下降相吻合，因为在感染和复制过程中，不断增加的细菌负荷耗尽了与细胞形态特定的角色一致的宿主营养库，细胞内复制过程中EB和RB转录活性的比较，对细胞形态特异的蛋白质组的分析，以及生化分析表明EBS和RBS在代谢上是特化的。因此，不同的营养物质可能会触发Eb-Rb和Rb-Eb之间转换的营养调节。识别这种不同的调控将需要对两种细胞形态与相同的营养线索进行直接比较。具体目标1的目的是确定衣原体SR组分(例如，dks A/dks A)是否与病原体在细胞内复制过程中的形态转变和宿主营养库的下降相一致。为了确定沙眼衣原体SR的特定营养触发因素，特定目的2的目标是在饥饿期间测量沙眼衣原体EB和RB SR的特定营养物质的分子标记。该项目解决了美国国立科学院的使命 健康，因为它将产生关于人类主要病原体的基本知识。

 DESCRIPTION (provided by applicant): Bacteria of the genus Chlamydia are obligate intracellular bacterial parasites. C. trachomatis is a human adapted species divided into several serologically different variants that cause serovar-specific diseases. While genital serovars of C. trachomatis cause approximately 100 million sexually transmitted infections worldwide annually, ocular serovars are a leading cause of preventable blindness in developing countries. All chlamydiae must transition between two terminally differentiated cell forms to complete their life cycle. Following host cell entry by the infectious, non-replicative Elementary Body (EB), the organism transitions to the replicative, non- infectious Reticulate Body (RB). Other pathogenic bacteria that transition between developmental forms can respond to nutrient availability as signaled via the stringent response (SR) as a regulator of differentiation. A typical SR is initiatd by synthesis of the RNA polymerase regulators guanosine tetraphosphate (ppGpp), also referred to as the alarmone, and the transcription factor DksA. Because deletion of dksA can disrupt differentiation of bacterial development, expression of dksA in C. trachomatis is consistent with a role for the SR in C. trachomatis development. With a genome of only 1.04 Mb, C. trachomatis lacks several predicted essential enzymes and entire metabolic pathways and must therefore obtain numerous nutrients, including amino acids and nucleotides, from the host cell. The proposed research is based on the hypothesis that C. trachomatis development is regulated by nutrient availability via the SR and that EB generation coincides with declining intracellular nutrient availability as the increasing bacterial burden depletes host nutrient pools Consistent with cell form-specific roles during infection and replication, comparison of EB and RB transcriptional activity during intracellular replication, analysis of cell form-specific proteomes, and biochemical analyses have revealed that EBs and RBs are metabolically specialized. As such, nutritional regulation of transitions between EB-RB and RB-EB may be triggered by different nutrients. Identification of such differential regulation will require direc comparison of both cell forms to the same nutritional cues. The goal for Specific Aim 1 is to determine if chlamydial SR components (e.g., dksA/DksA) are expressed coincident with pathogen morphological transitions and declining host nutrient pools during intracellular replication. In order to identify specific nutritional triggers for the C. trachomatis SR, the goalfor Specific Aim 2 is to measure molecular markers of the C. trachomatis EB and RB SR during starvation for specific nutrients. This project addresses the mission of the National Institutes of Health in that it will generate fundamental knowledge about a major human pathogen.