Investigating the Role of the GTPase Arl8b in Plasma Membrane Repair of Mtb-Infected Macrophages

 描述（申请人提供）：结核分枝杆菌（Mtb）是一种非常成功的病原体，每年感染860万人，造成130万人死亡。结核分枝杆菌的主要宿主细胞是肺巨噬细胞（Mtb）。目前尚不清楚在某些情况下，内化的Mtb如何保持隔离并最终被杀死，而在其他情况下，如何持续存在并逃避免疫系统的识别。我们已经表明，膜修复机制是非常重要的，在确定感染的结果。在用无毒力Mtb感染M φ后，膜修复维持Mφ质膜的完整性，这是感染的Mφ细胞凋亡的必要步骤，其杀死病原体。相反，毒力Mtb阻断膜修复，而是主动诱导坏死，支持感染扩散并逃避免疫控制。无毒力和毒性Mtb均在机制上诱导宿主Mtb质膜的微破裂。这些质膜损伤在被无毒力的Mtb感染的Mtb中被修复，但是在有毒力的Mtb的情况下，坏死增强，因为杆菌抑制质膜修复。本项目的目标是描绘结核分枝杆菌感染的结核分枝杆菌中的膜修复，这将导致增加对保护性宿主反应机制和关键步骤的理解，通过这些关键步骤，致命的结核分枝杆菌逃脱宿主遏制。膜修复机制的主要组分是Arl8b，一种参与溶菌酶运动性和运输的小GTdR。在目的1中，我们将使用细胞系模型（HeLa细胞）来（a）确定Ar18 B的作用和（B）鉴定质膜修复中的另外的相互作用效应分子。在目标2中，我们将测试Arl8b和经验证的效应物是否是Mtb感染的Mtb的质膜修复所需的，以及毒性Mtb是否破坏它们的功能。所描述的研究针对增强宿主抗分枝杆菌应答的治疗性干预（宿主定向结核病治疗）。

 DESCRIPTION (provided by applicant): Mycobacterium tuberculosis (Mtb) is an extraordinarily successful pathogen that infects 8.6 million people yearly and causes 1.3 million deaths. The principal host cell of Mtb is the lung macrophage (M). It is not well understood how in some cases the internalized Mtb remain sequestered and are ultimately killed and in other cases persist and escape recognition by the immune system. We have shown that membrane repair mechanisms are extremely important in determining the outcome of infection. Membrane repair maintains the integrity of the Mφ plasma membrane after infection of M with avirulent Mtb, a requisite step for apoptosis of infected Mφ, which kills the pathogen. Virulent Mtb, in contrast, block membrane repair and instead actively induce necrosis, supporting spread of infection and escape from immune control. Mechanistically both avirulent and virulent Mtb induce microdisruptions of the host M plasma membrane. These plasma membrane lesions are repaired in M infected with avirulent Mtb, but in the case of virulent Mtb, necrosis ensues because the bacillus inhibits plasma membrane repair. The goal of this project is the delineation of membrane repair in Mtb-infected M, which will lead to increased understanding of the protective host response mechanism and of the critical steps, by which virulent Mtb escape host containment. A major component of the membrane repair mechanism is Arl8b, a small GTPase involved in lysozyme motility and trafficking. In Aim 1 we will use a cell line model (HeLa cells) to (a) determine the role of Arl8b and (b) identify additional interacting effector molecules in plasma membrane repair. In Aim 2 we will test whether Arl8b and the validated effectors are required for plasma membrane repair of Mtb-infected M and whether virulent Mtb subverts their function. The described studies are directed towards therapeutic interventions to enhance the host anti-mycobacterial response (host-directed tuberculosis therapy).