The roles of Anaplasma phagocytophilum surface proteins in cellular invasion

描述(申请人提供)：吞噬细胞无浆菌是一种专性的细胞内细菌病原体，它侵入中性粒细胞和内皮细胞，导致新出现的和潜在的致命感染，人类粒细胞无浆体病(HGA)。A.吞噬细胞将其宿主中性粒细胞转化为特洛伊木马，从而促进病原体的复制和传播。它下调了中性粒细胞的抗微生物反应，从而增加了对机会性感染的敏感性。因此，阻断嗜中性粒细胞的吞噬杆菌感染可能会阻止与病原体传播相关的HGA阶段，并增加机会性感染的风险。吞噬艾美耳球虫可感染心脏和肝脏的微血管内皮细胞。内皮细胞也被认为是吞噬细胞性假丝酵母菌感染的初始细胞类型。受感染的内皮细胞能够将细菌转移到中性粒细胞。因此，消除吞噬阿糖胞菌对内皮细胞的侵袭有可能防止主要器官的感染和最初的感染建立。该项目的总体目标是鉴定和鉴定促进感染的吞噬细胞性假丝酵母菌外膜蛋白。我们已经确定外膜蛋白A(OmpA)和Asp14(14 kDa AP表面蛋白)是吞噬阿糖胞菌的前两个蛋白，它们对髓系细胞和内皮细胞的侵袭都是至关重要的。在目标1中，我们将确定介导吞噬杆菌摄取的OmpA和Asp14结构域，并确定这两种蛋白是否足以入侵。在目标2中，我们将鉴定OmpA和Asp14的宿主细胞受体。在目标3中，我们将使用新型转基因吞噬细胞生物直接评估OmpA和Asp14与感染性的相关性，这些转基因生物可以被诱导表达针对OmpA或Asp14的反义RNA，从而下调OmpA或Asp14的表达。实现我们的目标将提供对吞噬细胞侵袭的强有力的理解，并识别任何无浆体科病原体的前两个侵袭素受体对。它还将为评估专性细胞内细菌的基因功能提供一个有价值的工具。最后，我们的工作可能有助于设计针对OmpA和Asp14的新型干预策略，通过阻断多个阶段的感染来预防HGA。

DESCRIPTION (provided by applicant): Anaplasma phagocytophilum is an obligate intracellular bacterial pathogen that invades neutrophils and endothelial cells to cause the emerging and potentially fatal infection, human granulocytic anaplasmosis (HGA). A. phagocytophilum converts its host neutrophil into a Trojan Horse that facilitates pathogen replication and dissemination. It downregulates the neutrophil antimicrobial response, thereby raising susceptibility to opportunistic infections. Thus, blocking A. phagocytophilum infection of neutrophils would potentially prevent the stage of HGA associated with pathogen dissemination and increased risk of opportunistic infections. A. phagocytophilum infects microvascular endothelial cells of heart and liver. Endothelial cells are also implicated as the initial cell typ that A. phagocytophilum infects following inoculation via tick feeding. Infected endothelial cells are capable of transferring the bacterium to neutrophils. Therefore, abrogating A. phagocytophilum invasion of endothelial cells would potentially prevent infection of major organs and initial establishment of infection. The overall goal of this project is to identify and functioally characterize A. phagocytophilum outer membrane proteins that facilitate infection. We have identified outer membrane protein A (OmpA) and Asp14 (14-kDa Ap surface protein) as the first two A. phagocytophilum proteins that are critical for invasion of both myeloid cells and endothelial cells. In Aim 1, we will identify the OmpA and Asp14 domains that mediate A. phagocytophilum uptake and determine if either protein is sufficient for invasion. In Aim 2, we will identify the host cell receptors of OmpA and Asp14. In Aim 3, we will directly assess the relevance of OmpA and Asp14 to infectivity using novel transgenic A. phagocytophilum organisms that can be induced to express antisense RNA against ompA or asp14 to knock down OmpA or Asp14 expression. Achieving our goals will provide a robust understanding of A. phagocytophilum cellular invasion and identify the first two invasin-receptor pairs for any Anaplasmataceae pathogen. It will also yield a valuable tool for assessing gene function in obligate intracellular bacteria. Lastly, our work will potentially aid the design of novel intervenion strategies that target OmpA and Asp14 to prevent HGA by blocking multiple stages of infection.