Role of Yersinia pestis Ail in Yop delivery and plague

描述(由申请人提供)：革兰氏阴性细菌有效的三型分泌物(T3s)需要与目标宿主细胞黏附。我们最近已经确定，鼠疫杆菌粘附素Ail通过与宿主纤维连接蛋白结合来促进耶尔森菌外部蛋白(YOPs)的T3S。YOP是鼠疫感染所必需的，缺乏YOP的突变株是完全无毒的。All突变体在体外的YOP传递能力很差，在体内的作用减弱(&gt；LD50增加3000倍)。因此，AIL似乎是YOP体内传递的一种重要的粘附素。这使得AIL成为鼠疫疫苗开发和抗鼠疫治疗的重要潜在目标。我们假设AIL与纤维连接蛋白一起与宿主细胞相互作用，从而实现有效的YOP传递，可能是通过刺激YOP传递所需的受体启动的细胞信号传递。我们进一步假设Fn在表达AIL的细菌和宿主细胞上的21个整合素(Fn的受体)之间起到了桥梁作用。最近研究表明，整合素信号通路在侵袭素介导的YOP传递过程中起着重要作用，而YOP与假结核杆菌密切相关。在这个提案中，我们将定义AIL与FN相互作用的区域，并解决AIL与FN结合以及随后与整合素β1结合的影响。通过通过鼠疫杆菌的一种关键粘附素了解YOP传递过程中的这一关键步骤，我们将显著扩展我们对粘附性在YOP传递中的作用的知识，特别是在一般的T3s中。这些研究将广泛适用于利用T3S机制传递毒素的其他细菌病原体。此外，通过定义鼠疫杆菌和宿主细胞之间的关键相互作用机制，我们可以确定可用于治疗鼠疫的治疗靶点，鼠疫是一种迅速致命的疾病和生物恐怖主义威胁。 这项建议的目的是： 目的1：测定宿主细胞结合和YOP传递所需的AIL残基 目的2：鉴定AIL与宿主细胞纤维连接蛋白的结合以及21种整合素在AIL介导的YOP传递中的作用 公共卫生意义：鼠疫耶尔森氏菌引起迅速致命的传染病鼠疫，使鼠疫成为生物恐怖主义的威胁。本项目旨在加深我们对鼠疫杆菌表面蛋白Ail与宿主细胞相互作用的了解。这种关键的相互作用是鼠疫致病力所必需的，确定这种相互作用的特征可能会导致开发出抗鼠疫疗法和/或有效的鼠疫疫苗。

DESCRIPTION (provided by applicant): Efficient Type Three Secretion (T3S) by Gram-negative bacteria requires adhesion to the targeted host cell. We have recently determined that the Yersinia pestis adhesin Ail facilitates T3S of Yersinia outer proteins (Yops) via binding to host fibronectin. Yop delivery is required for plague infection and mutants that lack Yops are completely avirulent. A ?ail mutant has poor Yop delivery in vitro and is attenuated in vivo (>3000-fold increase in LD50). Thus, Ail appears to be a prominent adhesin for Yop delivery in vivo. This makes Ail an important potential target for Y. pestis vaccine development and anti-plague therapies. We hypothesize that Ail in conjunction with fibronectin interacts with host cells to allow efficient Yop delivery, possibly by stimulating receptor-initiated cell signaling required for Yop delivery. We further hypothesize that Fn acts as a bridge between Ail-expressing bacteria and 21 integrins on host cells (receptors for Fn). ?1 integrin signaling has recently been shown to be important for invasin-mediated Yop delivery by the closely related pathogen, Y. pseudotuberculosis. In this proposal, we will define the regions of Ail that interact with Fn and address the effects of Ail binding to Fn and subsequent engagement of ?1 integrins. By understanding this critical step in the Yop delivery process by a key adhesin of Y. pestis, we will significantly expand our knowledge of the role of adhesion in Yop delivery in particular and T3S in general. These studies will be broadly applicable to other bacterial pathogens that utilize a T3S mechanism for toxin delivery. Furthermore by defining a crucial interaction mechanism between Y. pestis and host cells, we may identify targets for therapeutics that can be used to treat plague, a rapidly fatal disease and bioterrorism threat. The Aims of this proposal are: Aim 1: Determine the residues of Ail required for host cell binding and Yop delivery Aim 2: Characterize Ail binding to host cell fibronectin and the role of 21 integrins in Ail-mediated Yop delivery PUBLIC HEALTH RELEVANCE: Yersinia pestis causes the rapidly fatal infectious disease plague, making Y. pestis a bioterrorism threat. This project is aimed at furthering our understanding of the interaction of the Y. pestis surface protein Ail with host cells. This critical interaction is required for plague virulence and characterizing this interaction may lead to development of anti-plague therapeutics and/or an effective plague vaccine.