Mechanisms of Rickettsia invasion, intracellular survival, and actin-based motility

立克次体侵袭、细胞内存活和基于肌动蛋白的运动的机制

• 批准号: 10238082
• 负责人: Matthew D Welch
• 金额: $ 38.01万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10238082
• 关键词: Actins; Address; Automobile Driving; Autophagocytosis; Bacteria; Bacterial Physiology; Cell Surface Receptors; Cells; Cellular Structures; Cellular biology; Cytoskeleton; Cytosol; Diagnosis; Diagnostic; Disease; Endothelium; Funding; Genes; Genome; Growth; Human; In Vitro; Infection; Invaded; Knowledge; Lead; Mediating; Membrane; Membrane Proteins; Molecular; Mus; Mutagenesis; Mutation; Nutrient; Pathogenesis; Phagosomes; Phase; Phenotype; Phospholipase; Play; Polymers; Process; Proteins; Publishing; Rickettsia; Rickettsia Infections; Rickettsia parkeri; Role; Study models; Surface Antigens; Testing; Typhus; Virulence; Work; bacterial genetics; base; cell motility; human disease; improved; in vivo; innovation; macrophage; mouse model; mutant; novel strategies; pathogen; pathogenic bacteria; polymerization; receptor; spotted fever; ubiquitin ligase; unpublished works

PROJECT SUMMARY/ABSTRACT The Rickettsiae are obligate intracellular bacterial pathogens that cause serious diseases, such as spotted fever and typhus. We study the model spotted fever group (SFG) species Rickettsia parkeri, which causes an eschar-associated human rickettsiosis, is experimentally tractable, and has emerging mouse models of pathogenesis, making it ideal for revealing molecular mechanisms of SFG Rickettsia infection and virulence. During infection, SFG Rickettsia invade host cells by mobilizing the actin cytoskeleton, escape from the phagosome into the cytosol, replicate while avoiding degradation by autophagy, and harness actin polymerization to promote intracellular motility and cell-cell spread. However, there are fundamental gaps in our knowledge of the molecular mechanisms by which SFG Rickettsia exploit or disrupt host cell components to promote their infection cycle. To bridge these gaps, in the current funding period we have pioneered an innovative combination of bacterial genetics and host cell biology to identify key Rickettsia factors that manipulate host cells. In unpublished work, we discovered that outer membrane protein OmpB is crucial for both invasion and avoidance of autophagy. We also observed that patatin-like phospholipase Pat1 plays a role in phagosome escape and/or autophagy evasion. Additionally, in published work, we demonstrated that Rickettsia use two actin-polymerizing surface proteins to direct sequential phases of motility – with RickA driving early motility and surface cell antigen Sca2 driving late motility. However, key outstanding questions remain, including: How do Rickettsia engage host receptors to promote invasion? How do Rickettsia degrade membranes during phagosome escape or inhibit membrane engulfment to avoid autophagy? And how do Rickettsia coordinate and use two actin assembly factors in distinct phases of motility? Our preliminary and published findings suggest the overall hypothesis that OmpB, Pat1, RickA, and Sca2 are multifunctional proteins that mobilize or disrupt host cell components and play a crucial role in infection in vivo. This hypothesis will be tested in three Aims focused on uncovering the mechanisms through which OmpB, Pat1, RickA, and Sca2 influence invasion, intracellular survival, and motility. The Aims are to: (1) define the role of Rickettsia surface protein OmpB in invasion and intracellular survival; (2) investigate the role of Pat1 phospholipase in phagosome escape and intracellular survival; and (3) determine how and why Rickettsia use two distinct actin-based motility mechanisms. The proposed studies will advance the field by revealing crucial molecular mechanisms used by Rickettsia and other pathogens to manipulate host cells and the importance of these mechanisms to infectivity. Our studies may also lead to improved diagnostics and treatments for rickettsial and other infections.

项目概要/摘要 立克次体是专性细胞内细菌病原体，可引起严重疾病，例如斑点病 我们研究了模型斑疹热组 (SFG) 种帕克立克次体，它会引起发烧和斑疹伤寒。 与焦痂相关的人类立克次体病，在实验上是易于处理的，并且已经出现了以下小鼠模型： 发病机制，使其成为揭示 SFG 立克次体感染和毒力分子机制的理想选择。 在感染过程中，SFG立克次体通过动员肌动蛋白细胞骨架侵入宿主细胞，逃离宿主细胞。 吞噬体进入细胞质，进行复制，同时避免自噬降解，并利用肌动蛋白 然而，在这方面存在根本性的差距。 我们对 SFG 立克次体利用或破坏宿主细胞成分的分子机制的了解 为了促进他们的感染周期，为了弥补这些差距，在当前的资助期间，我们开创了一个项目。 细菌遗传学和宿主细胞生物学的创新组合，以确定关键的立克次体因子 在未发表的工作中，我们发现外膜蛋白 OmpB 对于操纵宿主细胞至关重要。 我们还观察到patatin样磷脂酶Pat1在侵袭和避免自噬中发挥作用。 此外，在已发表的作品中，我们证明了这一点。 立克次体使用两种肌动蛋白聚合表面蛋白来指导运动的连续阶段 – 与 RickA 驱动早期运动和表面细胞抗原 Sca2 驱动晚期运动 然而，关键的悬而未决的问题。 仍然存在，包括：立克次体如何与宿主受体结合以促进入侵？立克次体如何降解？ 吞噬体逃逸或抑制膜吞噬以避免自噬的过程如何？ 立克次体在运动的不同阶段协调和使用两种肌动蛋白组装因子？ 已发表的研究结果表明总体假设：OmpB、Pat1、RickA 和 Sca2 是多功能的 动员或破坏宿主细胞成分并在体内感染中发挥关键作用的蛋白质。 假设将在三个目标中进行检验，重点是揭示 OmpB、Pat1、 RickA 和 Sca2 影响侵袭、细胞内存活和运动。目的是：(1) 定义 RickA 和 Sca2 的作用。 立克次体表面蛋白OmpB在侵袭和细胞内存活中的作用（2）探讨Pat1的作用； 磷脂酶在吞噬体逃逸和细胞内存活中的作用；以及（3）确定立克次体使用的方式和原因； 两种不同的基于肌动蛋白的运动机制将通过揭示关键的机制来推进该领域的发展。 立克次体和其他病原体操纵宿主细胞的分子机制以及其重要性 这些传染性机制也可能导致改进的诊断和治疗。 立克次体和其他感染。