Host actin cytoskeleton rearrangements induced by Salmonella

描述（由申请人提供）：入侵非吞噬性宿主细胞的能力对沙门氏菌的毒力至关重要。SipA, SipC, SopB， SopE和SopE2是已知的通过直接或间接调节宿主肌动蛋白动力学促进沙门氏菌入侵的五种III型效应物。SipA和SipC通过模拟宿主肌动蛋白结合蛋白的功能直接调节肌动蛋白动力学。SopB、SopE和SopE2间接破坏宿主细胞信号转导通路，诱导肌动蛋白细胞骨架重排和膜褶皱。这些细菌效应蛋白的协同作用最终导致细菌的有效吸收。SipA, SipC， SopB是多功能效应器，也参与效应器易位和诱导宿主炎症反应。尽管对这些效应物及其在沙门氏菌诱导的肌动蛋白细胞骨架重排中的作用有了更好的了解，但对它们的多功能活性的分离以及这些活性如何在功能上相互作用以促进沙门氏菌入侵的研究却很少。在这项研究中，我们建议研究它们的肌动蛋白调节活性的个体贡献以及它们在细菌入侵过程中的功能协调。我们的初步数据表明SipC通过瞬时二聚体形成发挥其肌动蛋白成核活性，并且SipA和SipC可能在功能上协同促进细菌进入。我们还证明了sopb依赖性VAMP8在沙门氏菌诱导的膜褶中募集，并且VAMP8是有效细菌入侵所必需的。我们进一步确定SipA和SipC可能介导SopB和sope独立的沙门氏菌进入。我们假设SipA、SipC、SopB和SopE共同或独立地调节宿主肌动蛋白动力学以促进沙门氏菌的进入。为了验证这一假设，我们将：(i)表征SipA， SipC肌动蛋白调节活性在沙门氏菌入侵中的作用；（ii）明确SopB促进肌动蛋白细胞骨架重排和沙门氏菌入侵的分子机制；（iii）研究SipA、SipC、SopB和SopE如何协调其动作蛋白调节活动促进沙门氏菌入侵。该研究结果将为SipA、SipC和SopB如何单独或协同利用宿主肌动蛋白细胞骨架促进沙门氏菌进入提供分子基础。

DESCRIPTION (provided by applicant): The ability to invade non-phagocytic host cells is essential for Salmonella virulence. SipA, SipC, SopB, SopE, and SopE2 are five type III effectors that are known to promote Salmonella invasion by modulating host actin dynamics both directly and indirectly. SipA and SipC regulate actin dynamics directly by mimicking functions of host actin-binding proteins. Indirectly, SopB, SopE, and SopE2 subvert host cell signal transduction pathways to induce actin cytoskeletal rearrangements and membrane ruffling. The concerted actions of these bacterial effector proteins eventually lead to the efficient uptake of the bacterium. SipA, SipC, SopB are multifunctional effectors also involved in effector translocation and in inducing host inflammatory responses. Despite improved understanding of these effectors and their individual roles in Salmonella-induced actin cytoskeleton rearrangements, little has been done to separate their multifunctional activities and to examine how these activities functionally interact to promote Salmonella invasion. In this study, we propose to investigate the individual contributions of their actin-modulating activities and how they are functionally coordinated during bacterial invasion. Our preliminary data showed that SipC exerts its actin-nucleation activity through transient dimer formation and that SipA and SipC may functionally cooperate to promote bacterial entry. We also demonstrated SopB-dependent VAMP8 recruitment to Salmonella-induced membrane ruffles and that VAMP8 is required for efficient bacterial invasion. We further determined that SipA and SipC may mediate SopB- and SopE-independent Salmonella entry. We hypothesize that SipA, SipC, SopB, and SopE function both cooperatively and independently to regulate host actin dynamics to promote Salmonella entry. To test this hypothesis, we will: (i) characterize the contributions of SipA, SipC actin-modulating activities in Salmonella invasion; (ii) define the molecular mechanism by which SopB promotes actin cytoskeleton rearrangements and Salmonella invasion; (iii) study how SipA, SipC, SopB, and SopE coordinate their actin-modulating activities to promote Salmonella invasion. Results from this study will provide a molecular basis for how SipA, SipC and SopB exploit the host actin cytoskeleton both independently and cooperatively to promote Salmonella entry. Project Narrative: Despite the understanding gained by our long-standing interest in studying Salmonella, salmonellosis continues to pose worldwide medical concerns and remains the number one cause of food-borne diseases even in developed countries. The identification of host cellular targets of Salmonella virulence proteins will aid clinical therapeutic drug designs and treatment of multidrug-resistant bacteria. Results from this study will not only broaden our understanding of Salmonella pathogenesis but will also provide vital insights into basic host cell biology.