Mechanisms of SFG Rickettsia-Host Interactions

项目摘要/摘要 在美国，由斑点热病引起的一组新出现的、但特征不佳的壁虱传播疾病 群(SFG)立克次体。这些专有的细胞内细菌病原体会导致轻微的甚至危及生命的血管 人类的疾病，诊断和治疗干预措施也很有限。促进广泛传播 疾病，SFG立克次体物种进化出动态策略，入侵宿主细胞，逃逸到细胞质中， 从一个细胞传播到另一个细胞。我们假设SFG立克次体通过传递 分泌的细菌蛋白(即效应器)的武器库，它重新编程宿主细胞的过程。不幸的是， 这些分泌效应器的身份和宿主目标在很大程度上仍不清楚。此外，直接分析 分泌系统和假定的效应器由于在生长和生长中的挑战而受到阻碍 在实验室里对这些病原体进行基因操作。为了克服这一障碍，我们最近向前调整了 对模型SFG成员帕氏立克次体的遗传学和基于质粒的互补，允许 SFG立克次体发病机制的强大功能遗传学研究。使用这些工具，我们发现 分泌型效应子Sca4通过调控宿主细胞与细胞间的黏附，促进细胞间扩散的晚期。 此外，功能未知的分泌型效应器(RARP-1)及其组分的转座子突变 异常的4型立克次体分泌系统(VirB6e)损害帕氏杆菌感染的不同阶段。这 提案将利用这些关键进展来研究分泌系统和分泌效应器如何促进 帕氏杆菌感染生命周期的不同阶段。在这里，我们将我们的功能遗传策略和 在宿主细胞生物学方面的专业知识，以及现代生化技术，以揭示关键的、机械的见解 SFG立克次体发病机制。在目标1中，我们将研究T4SS组件的功能和分泌组 VirB6e。在目标2中，我们将确定分泌的效应物RARP-1如何促进帕氏杆菌感染。在《目标3》中， 我们将阐明Sca4是如何在帕氏杆菌细胞间的过程中特异性地靶向宿主细胞-细胞黏附复合体的。 散开。总的来说，拟议的研究将极大地提高我们对 立克次体生物学和立克次体与宿主的相互作用并揭示预防或治疗扁虱病的治疗靶点 疾病。

PROJECT SUMMARY/ABSTRACT A subset of emerging, but poorly characterized tickborne diseases in the U.S. are caused by Spotted Fever Group (SFG) Rickettsia. These obligate intracellular bacterial pathogens cause mild-to-life-threatening vascular diseases in humans and have a limited set of diagnostics and therapeutic interventions. To promote widespread disease, SFG Rickettsia species have evolved dynamic strategies to invade host cells, escape into the cytosol, and spread from cell to cell. We hypothesize that SFG Rickettsia coordinate their complex life cycle by delivering an arsenal of secreted bacterial proteins (i.e., effectors) that reprogram host cell processes. Unfortunately, the identity and host targets of these secreted effectors have remained largely unknown. Furthermore, direct analysis of the secretion systems and putative effectors have been hampered due to challenges in growing and genetically manipulating these pathogens in the lab. To overcome this barrier, we recently adapted forward genetics and plasmid-based complementation to the model SFG member Rickettsia parkeri, allowing for powerful functional-genetic studies of SFG Rickettsia pathogenesis. Using these tools, we discovered that the secreted effector Sca4 promotes a late stage of cell-to-cell spread by manipulating host cell-cell adhesion. Additionally, transposon mutagenesis of a secreted effector of unknown function (RARP-1) and a component of the anomalous Rickettsia Type 4 secretion system (VirB6e) impairs distinct stages of R. parkeri infection. This proposal will leverage these key advances to examine how a secretion system and secreted effectors promote different steps of the R. parkeri infectious life cycle. Here, we combine our functional-genetic strategies and expertise in host cell biology, with modern biochemical techniques to reveal critical, mechanistic insights into SFG Rickettsia pathogenesis. In Aim 1, we will examine the function and secretome of the T4SS component VirB6e. In Aim 2, we will determine how the secreted effector RARP-1 promotes R. parkeri infection. In Aim 3, we will elucidate how Sca4 specifically targets host cell-cell adhesion complexes during R. parkeri cell-to-cell spread. Collectively, the proposed research will dramatically improve our fundamental understanding of Rickettsia biology and Rickettsia-host interactions and reveal therapeutic targets to prevent or treat tickborne diseases.