THE ROLE OF NOVEL SURFACE PROTEINS IN YERSINIA PESTIS VIRULENCE

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 鼠疫耶尔森氏菌是鼠疫的病原体，在感染过程中可以结合在表面，也可以入侵哺乳动物的宿主细胞。这种活性是一个活跃的领域或研究，充分了解导致巨噬细胞和淋巴细胞持续存在的感染序列对于剖析这种重要的革兰氏阴性细菌的致病性至关重要。为此，我们建立了一个完整的TnPhoA文库来鉴定鼠疫杆菌的所有表面蛋白。通过对该报告融合文库的分析，通过DNA序列分析鉴定出多个侵袭素样蛋白。在候选基因上，鼠疫耶尔森菌可能的内毒素蛋白(由鼠疫菌CO92中的YPO3944基因编码)是一个由LysM基序组成的3013个残基的大蛋白，保守于革兰氏阴性细菌入侵蛋白的一个子集，以及细菌Ig样域的串联重复序列。虽然这个假定的鼠疫入侵蛋白在所有已测序的鼠疫生物型的基因组中都是保守的，但该蛋白在鼠疫致病中的确切作用还没有被研究过。我们对这个项目的具体目标是： 1.利用PhoA报告系统和实时荧光定量聚合酶链式反应检测该基因在不同环境条件下的调控。 2.通过定点突变产生缺失突变，并在细胞培养侵袭实验和病毒致病模型中比较该突变株与野生型亲本株的毒力。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Yersina pestis, the etiologic agent of plague, can bind to the surface, as well as invade, mammalian host cells during the infection process. This activity is an active area or research and a full understanding of the infection sequence leading to persistence in macrophages and lymphatic cells is essential in dissecting the pathogenicity of this important Gram negative bacterium. To this end, we generated a comprehensive TnphoA library to identify all surface proteins of Y. pestis. Analysis of this reporter fusion library identified a number of invasin like proteins by DNA sequence analysis. On candidate gene, the Yersinia pestis putative invasin protein (encoded by gene YPO3944 in Y. pestis CO92) is a large 3013 residue protein composed of a LysM motif, conserved in a subset of Gram negative bacterial invasion proteins, as well as tandem repeats of a bacterial Ig-like domain. Although this putative Y. pestis invasin protein is conserved in the genome of all sequenced Y. pestis biotypes, the exact role of this protein in the pathogenesis of Y.pestis has not been examined. Our specific goals for this project are to: 1. Determine the regulation of this gene under various environmental condtions using the phoA reporter system and real time PCR. 2. Generate a deletion mutation by site-directed mutagenesis and compare the virulence of this mutant with the wild-type parental strain in cell culture invasion assays and the mosue pathogenesis model.