LC3-Associated Phagocytosis

摘要 两个古老的过程，吞噬作用和巨噬细胞，出现了作为满足能量需求的方式 手机。两者也都进化成了寄主防御机制。在对这笔赠款的前一次支持期间，我们 发现了一种我们称之为“LC3相关吞噬作用”(LAP)的过程。在这个过程中，信号是 吞噬细胞吞噬颗粒时产生的物质诱导自噬机制的组件 与吞噬小体结合，促进其与溶酶体的融合(吞噬小体成熟)。当被吞没时 乳胶珠(例如)不会引起LAP，与TLR1/2、TLR2/6、TLR4、FCR或 吞噬死亡细胞的受体，导致吞噬体膜上的LC3(ATG8)的募集。喜欢 宏自噬，这种LC3关联依赖于Beclin1、PI3P生成、ATG5和ATG7，但不同于 自噬，Lc3与单噬菌体膜(而不是双膜)有关 自噬小体)。此外，与宏自噬不同的是，LAP在缺乏 自噬预启动复合体、ULK1、ATG13和FIP200。这提出了一种耐人寻味的可能性：现在是 已经证实，自噬机制的某些组件中的缺陷会促进炎症 并损害宿主对细胞内感染的防御。LAP作为一种离散现象的存在 这表明至少有一些这样的影响可能特别与LAP有关。在这里，我们建议将 LAP及其与吞噬小体成熟的关系及其在先天性免疫反应和正常发育中的作用 动态平衡。我们的中心假设，也就是这个应用程序所基于的，是取决于信号 在吞噬的同时，LAP可以促进吞噬小体的货物分选到细胞内 用于进一步信号检测、处理或降级的隔室。具体地说，我们会问：1.什么 LAP的启动与宏观自噬有何区别？在这里，我们将探索分子事件 启动和传播LAP，并评估这些与传统的巨型自噬有何不同。2. LAP如何促进吞噬小体成熟？在这里，我们将研究LAP的组件如何 极大地加速吞噬小体的成熟和每条途径中发生这种增强的点。 我们将进一步研究LAP诱导的吞噬小体成熟对巨噬细胞介导的后果 主场防守。3.LAP对炎症和动态平衡有何影响？在这里，我们将使用体外和 在活体系统中，在体外和体内，询问LAP在死亡细胞的炎症反应中的作用。 虽然凋亡细胞被认为是“免疫沉默的”，但我们的证据表明，至少 部分原因是由于LAP抑制吞噬细胞中的炎性细胞因子反应。我们将对此进行测试 令人兴奋的想法，并探索LAP缺陷对巨噬细胞和 其他车厢。总体而言，我们的项目试图描述LAP是如何作为两个古老的 途径，影响先天免疫，为理解炎症性疾病提供了新的途径。

Abstract Two ancient processes, phagocytosis and macroautophagy, arose as ways to meet the energy demands of the cell. Both also evolved into mechanisms of host defense. During the previous support period for this grant, we discovered a process we term "LC3-Associated Phagocytosis" (LAP). In this process, signals that are generated upon engulfment of particles by phagocytic cells induce components of the autophagy machinery to associate with the phagosome, promoting its fusion to lysosomes (phagosome maturation). While engulfment of latex beads (for example) does not induce LAP, particles that engage TLR1/2, TLR2/6, TLR4, FcR, or receptors for engulfment of dying cells, cause recruitment of LC3 (ATG8) to the phagosome membrane. Like macroautophagy, this LC3 association depends on Beclin1, PI3P generation, ATG5, and ATG7, but unlike autophagy, LC3 associates with the single phagosome membrane (rather then the double membrane of autophagosomes). Further, unlike macroautophagy, LAP proceeds in the absence of elements of the autophagic pre-initiation complex, ULK1, ATG13, and FIP200. This raises an intriguing possibility: It is now well established that defects in some components of the autophagy machinery promote inflammatory disease and compromise host defense to intracellular infections. The existence of LAP as a discrete phenomenon suggests that at least some such effects may specifically relate to LAP. Here, we propose to characterize LAP, its relationship to phagosome maturation, and its roles in innate immune responses and normal homeostasis. Our central hypothesis, upon which this application is based, is that depending on signaling that accompany phagocytosis, LAP can be engaged to promote the sorting of the phagosome cargo to intracellular compartments for further signal detection, processing, or degradation. Specifically, we will ask: 1. What distinguishes the initiation of LAP versus macro-autophagy? Here we will explore the molecular events that initiate and propagate LAP and evaluate how these differ from those of conventional macroautophagy. 2. How does LAP promote phagosome maturation? Here we will investigate how the components of LAP greatly accelerate phagosome maturation and the points in each pathway where this enhancement occurs. We will further examine the consequences of LAP-induced phagosome maturation for macrophage-mediated host defense. 3. How does LAP impact on inflammation and homeostasis? Here we will use in vitro and in vivo systems to interrogate the roles of LAP in the inflammatory response to dying cells, in vitro and in vivo. While apoptotic cells are thought to be "immunologically silent" our evidence suggests that this may be, at least in part, due to suppression of the inflammatory cytokine response by LAP in phagocytes. We will test this exciting idea, and explore the long-term inflammatory consequences of defective LAP in macrophages and other compartments. Overall, our project seeks to characterize how LAP, as the conjunction of two ancient pathways, impacts innate immunity, offering new avenues for understanding inflammatory disease.