Characterization of the Brucella abortus virB locus

描述(申请人提供)：流产布鲁氏菌为兼性 通过气溶胶途径具有高度传染性的细胞内病原体 会导致慢性衰弱疾病。感染流产杆菌的一个关键步骤是 巨噬细胞内持续感染的建立。细菌 编码毒力机制的基因是相互作用所必需的 布鲁氏菌和巨噬细胞在很大程度上仍未被发现。我们已经确定了一个 确定感染所需的流产杆菌VIRB基因座 在巨噬细胞和小鼠模型中都是如此。流产杆菌VIRB基因座为 通过序列同源性预测编码IV型分泌系统。我们的长- Range的目标是阐明VIRB基因座的调节机制 细胞内存活和持续感染。这样做的目的是 应用是研究VIRB基因的表达，并比较 野生型流产巴氏杆菌与Virb突变体在液泡中的相互作用 巨噬细胞的贩运。这个应用程序的中心假设是 VIRB基因座调节与巨噬细胞的关键相互作用 允许流产杆菌建立感染。建议的理由是 研究表明，流产杆菌毒力因子的特性介导 与巨噬细胞的特定相互作用将构成新方法的基础 治疗或预防布鲁氏菌病我们做好了独一无二的准备， 建议的研究，因为我们已经生成了研究VIRB的工具 在转录和翻译水平上都有表达。此外， 这项工作将在一个良好的研究环境中进行， 有利于它的完成。我们的部门有几个基金 致力于细胞内细菌病原体和优秀BL-3的研究人员 设施，以及可用于研究的其他共享资源 寄主/病原体的相互作用。核心假设将得到检验，而 本应用程序的目标通过追求以下两个目标来实现 具体目标：(1)确定体外和体内表达的条件 流产杆菌VIRB基因座和细菌中蛋白质产物的定位，以及 (2)确定VIRB基因座使流产杆菌 在巨噬细胞内存活和生长。我们预计， 这项工作的结果将为表达的第一个直接证据 流产杆菌VIRB蛋白以及定义环境信号 诱导该基因座的表达。此外，我们的结果将提供 定义细胞相互作用的关键信息 VIRB轨迹。这些结果将是重要的，因为他们预计将 为将要采用的预防或治疗干预提供新的目标 在非法使用这种细菌病原体的情况下。此外，它是 预计这些结果将促进我们对IV型分泌物的了解 系统，这些系统被许多不同的细菌病原体用来 颠覆宿主的防御机制。

DESCRIPTION (provided by applicant): Brucella abortus is a facultative intracellular pathogen that is highly infectious by the aerosol route and causes chronic, debilitating disease. A key step in B. abortus infection is the establishment of persistent infection within macrophages. The bacterial genes encoding virulence mechanisms required for specific interactions between Brucella and the macrophage remain largely undiscovered. We have identified a genetic locus of B. abortus, virB, that is required for establishing infection both in macrophages and In the mouse model. The B. abortus virB locus is predicted by sequence homology to encode a type IV secretion system. Our long- range goal is to elucidate the mechanism by which the virB locus mediates intracellular survival and persistent infection. The objective of this application is to study the expression of the virB genes and compare the interaction of wild type B. abortus and virB mutants with regard to vacuolar trafficking in the macrophage. The central hypothesis of this application is that the virB locus mediates a critical interaction with the macrophage that allows B. abortus to establish infection. The rationale for the proposed research is that characterization of B. abortus virulence factors mediating specific interactions with macrophages will form the basis for new approaches to treat or prevent brucellosis. We are uniquely prepared to undertake the proposed research, because we have generated tools for studying virB expression at both the transcriptional and translational level. Furthermore, the work will be performed in an excellent research environment that is conducive to its completion. Our Department contains several funded investigators working on intracellular bacterial pathogens and excellent BL-3 facilities, as well as other shared resources available for the study of host/pathogen interactions. The central hypothesis will be tested, and the objectives of this application accomplished by pursuing the following two specific aims: (1) Identify conditions for In vitro and in vivo expression of the B. abortus virB locus and localize protein products in the bacterium, and (2) Determine the mechanism by which the virB locus enables B. abortus to survive and grow intracellularly within macrophages. We expect that the results of this work will provide the first direct evidence for expression of the B. abortus virB proteins as well as define the environmental signals that induce expression of this locus. Furthermore, our results will provide information essential to defining the cellular interaction mediated by the virB locus. These results will be significant, because they are expected to provide new targets for preventive or therapeutic interventions to be employed in the case of illegitimate use of this bacterial pathogen. In addition, it is expected that these results will advance our knowledge of type IV secretion systems, which are used by a number of different bacterial pathogens to subvert the host's defense mechanisms.