Autophagy proteins in the immune response to Mycobacterium tuberculosis infection

项目概要/摘要 结核分枝杆菌（Mtb）感染每年杀死的人比任何其他单一的传染性病原体都多。两 结核病的疾病结果和病理学是由宿主体内的免疫反应驱动的， 由参与先天免疫和适应性免疫的细胞之间的相互作用决定。由于联合国的关键作用， 由于结核病的免疫反应，人们对开发利用结核病免疫反应的免疫疗法越来越感兴趣。 以控制感染。特别地，人们对刺激自噬以控制Mtb感染感兴趣。 自噬在控制Mtb感染中的作用是基于观察到Atg 5 fl/fl-LysM-cre小鼠 单核细胞衍生细胞和中性粒细胞（多形核细胞，PMN）中缺乏Atg 5， 30天，与缺乏IFN-γ信号传导的小鼠相似的极其严重的表型。ATG 5是一个重要的 自噬蛋白和该领域的教条是，在巨噬细胞中需要Atg 5来靶向Mtb， 通过大自噬的溶酶体降解。与预期相反，我们已经证明Atg 5是 骨髓细胞中所需的控制结核分枝杆菌感染，由于自噬独立的功能，调节 PMN主导的炎症。此外，使用仅在PMN中有条件地缺失Atg 5的小鼠（Atg 5 fl/fl-1）， MRP 8-cre），我们已经表明中性粒细胞中Atg 5的缺失可以导致对Mtb感染的易感性，揭示了一种可能的机制， 在结核分枝杆菌感染过程中，中性粒细胞对Atg 5的内在作用因此，更好地理解以下方面的作用至关重要： 自噬蛋白在宿主防御中的作用，以了解干预自噬蛋白时可能发生的影响范围。 自噬流与宿主导向疗法。我们对Atg 5 fl/fl-LysM-cre小鼠中Mtb发病机制的分析表明， 揭示了感染期间Atg 5在骨髓细胞中发挥作用的多个阶段。在结核分枝杆菌感染期间，早期- 感染的Atg 5-/-细胞过度产生细胞因子和趋化因子，将PMN带入Atg 5 fl/fl-LysM-cre的肺中 小鼠的数量高于对照组小鼠。应答性Atg 5-/- PMN放大促炎信号 而Atg 5-/- PMN的一个子集没有从肺中清除，我们预测这是由于PMN的缺陷造成的。 凋亡PMN的凋亡和/或红细胞增多。凋亡的结核分枝杆菌感染的中性粒细胞的胞饮作用是重要的 用于有效的抗原呈递。事实上，清除Atg 5-/- PMN的缺陷与延迟的运输相关。 T细胞进入肺部。基于这些初步数据，我假设Atg 5在多发性骨髓瘤中发挥作用， 细胞在Mtb感染期间，包括PMN中的自噬独立功能，以及这些功能 Atg 5的表达导致对Mtb致病性的控制。为了验证这个假设，我将确定 髓系细胞中Atg 5的缺失如何导致PMN积聚并阻止保护性适应性免疫 1）分析Atg 5在调节结核杆菌前体蛋白产生中的作用， 感染过程中的炎症信号，2）确定Atg 5在调节PMN积聚和清除中的作用 在Mtb感染期间，3）确定髓样细胞中Atg 5的缺失如何影响对Mtb的适应性免疫应答。

PROJECT SUMMARY/ABSTRACT Mycobacterium tuberculosis (Mtb) infection kills more people a year than any other single infectious agent. Both the disease outcome and the pathology of TB are driven by the immune response mounted in the host, which is dictated by interactions between cells involved in innate and adaptive immunity. Because of the pivotal role of the immune response in TB, there is a growing interest in developing immunotherapies that harness the immune response to control the infection. In particular, there is interest in stimulating autophagy to control Mtb infection. The role of autophagy in controlling Mtb infection was based on the observation that Atg5fl/fl-LysM-cre mice lacking Atg5 in monocyte-derived cells and neutrophils (polymorphonuclear cells, PMN) succumb to Mtb within 30 days, an extremely severe phenotype similar to mice lacking IFN-γ signaling. Atg5 is an essential autophagy protein and the dogma in the field was that Atg5 was required in macrophages to target Mtb for lysosomal degradation through macroautophagy. Contrary to expectation, we have demonstrated that Atg5 is required in myeloid cells to control Mtb infection due to an autophagy-independent function that regulates PMN-dominated inflammation. In addition, using mice that conditionally delete Atg5 only in PMN (Atg5fl/fl- MRP8-cre), we have shown that loss of Atg5 in PMNs can result in susceptibility to Mtb infection, revealing a PMN intrinsic role for Atg5 during Mtb infection. Therefore, it will be important to better understand the roles for autophagy proteins in host defense to understand the scope of effects that may occur while intervening with autophagic flux with host-directed therapies. Our dissection of Mtb pathogenesis in Atg5fl/fl-LysM-cre mice has revealed multiple stages where Atg5 functions in myeloid cells during infection. During Mtb infection, an early- infected Atg5-/- cell overproduces cytokines and chemokines that bring PMN into the lungs of Atg5fl/fl-LysM-cre mice in higher numbers than in control mice. The responding Atg5-/- PMN amplify the pro-inflammatory signals and a subset of Atg5-/- PMN are not cleared from the lung, which we predict occurs due to a defect in PMN apoptosis and/or efferocytosis of the apoptotic PMN. Efferocytosis of apoptotic Mtb-infected PMN is important for efficient antigen presentation. Indeed, the defect in clearing Atg5-/- PMN correlates with delayed trafficking of T cells to the lungs. Based on these preliminary data, I hypothesize that Atg5 plays roles in multiple myeloid cells during Mtb infection, including an autophagy-independent function in PMN, and together these functions of Atg5 lead to control of Mtb pathogenesis. To test this hypothesis, I will determine the mechanistic basis for how loss of Atg5 in myeloid cells leads to PMN accumulation and precludes a protective adaptive immune response to Mtb by pursuing the following aims: 1) Dissect the roles for Atg5 in regulating production of pro- inflammatory signals during infection, 2) Define the roles for Atg5 in regulating PMN accumulation and clearance during Mtb infection, 3) Determine how loss of Atg5 in myeloid cells affects adaptive immune responses to Mtb.