PROTEINS IN THE CHLAMYDIAL INCLUSION MEMBRANE

描述(改编自申请人的摘要)：四种 衣原体在人类和许多人身上都会引起各种严重的疾病 动物物种。人类最严重的衣原体疾病包括 沙眼、肺炎和一系列不同的性传播疾病 条件。全世界有数百万人因这些疾病而变得虚弱。 衣原体是一种专性的细胞内细菌病原体 在受感染的宿主细胞内的未酸化的空泡(包涵体)内。 包裹体形成和发育的生物学才刚刚开始 明白了。鹦鹉热衣原体产生的蛋白质集合(印加， IncB、IncC)在衣原体感染过程中定位于包涵膜 发展。这些是已知的仅有的衣原体或衣原体蛋白 寄主来源，定位于衣原体包涵体的膜。 最近，印加被证明是被宿主细胞酶和 暴露在包涵体表面的细胞质上。这些结果 暗示印加在包含与其相互作用中可能是重要的 细胞内环境。INCA/B/C IS的功能特征 由于衣原体中缺乏遗传系统，以及 这些蛋白质与基因中的任何序列之间都没有相似性 数据库。这项研究提案的总体目标是确定 Inca/B/C在衣原体发育过程中的作用三位一体 本提案中的目标描述了为实现这一目标而设计的方法。 目标1中提出的实验旨在识别 INCA/B/C暴露于感染细胞的细胞质中。AIM 2利用 酵母双杂交系统和噬菌体展示方法的研究 INCA/B/C与宿主和衣原体蛋白的相互作用 可能存在于细胞质或包涵体内的起源。目标3 描述了两种表达系统的使用，痘苗病毒和真核细胞。 表达载体，用于检测和诱变宿主细胞内的Inca/B/C。 这些表情实验将在缺席或 存在平行的衣原体感染。这些不同的方法 将研究印加/B/C的独特生物学并探索新的 研究衣原体与哺乳动物细胞相互作用的技术。

DESCRIPTION (Adapted from the applicant's abstract): The four species of chlamydiae cause a variety of serious diseases in both humans and in many animal species. The most serious chlamydial diseases of humans include trachoma, pneumonia, and a spectrum of different sexually transmitted conditions. Millions of people are debilitated by these diseases worldwide. The chlamydiae are obligate intracellular bacterial pathogens that develop within a non-acidified vacuole (the inclusion) within infected host cells. The biology of inclusion formation and development is only beginning to be understood. A collection of proteins produced by Chlamydia psittaci (IncA, IncB, IncC) are localized to the inclusion membrane during chlamydial development. These are the only known proteins, of either chlamydial or host origin, that are localized to the membrane of the chlamydial inclusion. Recently IncA was shown to be phosphorylated by host cell enzymes and exposed to the cytoplasm at the surface of the inclusion. These results suggest IncA may be important in the interaction of the inclusion with its intracellular environment. Functional characterization of IncA/B/C is complicated by the lack of a genetic system in the chlamydiae, as well as the absence of similarity between these proteins and any sequences in the databases. The overall goal of this research proposal is to identify the functions of IncA/B/C in the chlamydial developmental process. The three Aims in this proposal describe methodologies designed to achieve that goal. The experiments proposed in Aim 1 are designed to identify domains of IncA/B/C exposed to the cytoplasm in infected cells. Aim 2 utilizes the yeast two-hybrid system and bacteriophage display methodologies to examine interactions between IncA/B/C and proteins of either host or chlamydial origin that may be present in the cytoplasm or within the inclusion. Aim 3 describes the use of two expression systems, vaccinia virus and eukaryotic expression plasmids, to examine and mutagenize incA/B/C within host cells. These expression experiments will be conducted both in the absence or the presence of a parallel chlamydial infection. These different approaches will examine the unique biology of IncA/B/C as well as explore new technologies for investigating chlamydial interactions with mammalian cells.