Autophagy in Antiviral Immunity

描述（由申请人提供）：自噬是一种古老的进化保守途径，旨在通过降解细胞质中的长寿命蛋白质和细胞器来维持细胞稳态。它也被用作饥饿条件下的生存机制。最近的研究表明，自噬是先天免疫系统和适应性免疫系统的细胞用来对抗病毒感染的。病毒的先天识别通过两种不同的途径发生。在专业的病毒传感器浆细胞样树突状细胞（pDC）中，通过toll样受体（TLR） 7和9在核内体中识别病毒。我们最近的工作表明，自噬在通过TLR7识别pDCs中病毒感染的特征中起着关键作用。与pDCs相反，大多数其他类型的细胞通过RIG-I样受体（RLR）家族利用细胞质传感器进行病毒复制。参与自噬的分子已被证明可以阻断RLR信号。此外，最近的报道表明，自噬将内源性病毒抗原传递到MHC II类装载室，从而激活CD4 T细胞。然而，这些途径在体内病毒感染中的相关性尚不清楚。在本研究中，我们提供了初步数据，揭示了在单纯性疱疹病毒（HSV； TLR9激动剂）感染后，通过TLR9信号转导导致pDCs中I型IFN基因激活的过程中，Atg5是自噬体形成所需的关键分子。此外，我们发现自噬负调控非浆细胞样树突状细胞在水泡性口炎病毒感染（VSV； RIG-I激动剂）时的RLR通路。最后，我们证明了自噬在HSV-1感染时树突状细胞MHC II类上各种形式抗原的加工和呈递中的关键体内作用。在这些初步研究的基础上，我们建议使用各种分子和细胞生物学技术并使用已建立的病毒感染小鼠模型来检查自噬在先天和适应性免疫反应中的重要性。在第一个目标中，我们将通过使用分子和细胞生物学技术确定Atg5和/或自噬通过TLR9介导HSV感染pDCs的信号传导机制。在第二个目标中，我们建议通过蛋白质组学和生化方法确定自噬如何调节VSV感染时RNA传感器的激活。在最后一篇论文中，我们将探讨树突状细胞如何在体外和体内利用自噬来加工和呈递细胞外病毒抗原，在树突状细胞群中选择性地缺乏自噬。通过对自噬如何协调产生针对病毒感染的先天免疫和适应性免疫的基本了解，这些研究将有助于为设计针对各种病毒病原体的疫苗和抗感染措施奠定重要基础

DESCRIPTION (provided by applicant): Autophagy is an ancient evolutionarily conserved pathway designed to maintain cellular homeostasis by degrading long-lived proteins and organelles in the cytosol. It is also used as a survival mechanism under starvation conditions. Recent studies demonstrated that autophagy is utilized by the cells of the innate and adaptive immune systems to combat viral infections. Innate recognition of viruses occurs via two distinct pathways. In professional viral sensors, the plasmacytoid dendritic cells (pDC), recognition of viruses occurs in the endosomes via Toll-like receptors (TLR) 7 and 9. Our recent work has demonstrated that autophagy plays a key role in recognizing signatures of viral infection in pDCs through TLR7. In contrast to pDCs, most other cell types of the body utilize cytosolic sensors of viral replication via RIG-I like receptor (RLR) family. Molecules involved in autophagy have been shown to block RLR signaling. In addition, recent reports indicate that autophagy delivers endogenous viral antigens to the MHC class II loading compartment, allowing activation of CD4 T cells. However, the relevance of such pathways during in vivo virus infection is unknown. In this application, we present preliminary data that reveal the requirement for Atg5, a key molecule required for formation of autophagosomes, in the transduction of signaling through TLR9 leading to the activation of type I IFN genes in pDCs upon herpes simplex virus (HSV; TLR9 agonist) infection. In addition, we show that autophagy negatively regulates RLR pathway in non-plasmacytoid dendritic cells upon vesicular stomatitis virus infection (VSV; RIG-I agonist). Finally, we demonstrate a key in vivo role for autophagy in the processing and presentation of various forms of antigens on MHC class II in dendritic cells upon HSV-1 infection. Building on these preliminary studies, we propose to examine the importance of autophagy in both innate and adaptive immune responses using a variety of molecular and cell biological techniques and using established mouse models of virus infection. In the first Aim, we will determine the mechanism by which Atg5 and/or autophagy mediates signaling through TLR9 upon HSV infection in pDCs through the use of molecular and cellular biological techniques. In the second Aim, we propose to determine how autophagy regulates RNA sensor activation upon VSV infection through proteomics and biochemical approaches. In the final Aim, we will interrogate how dendritic cells utilize autophagy for processing and presentation of extracellular viral antigens in vitro and in vivo in mice selectively deficient in autophagy within the dendritic cell populations. By providing basic understanding of how autophagy orchestrates the generation of innate and adaptive immunity against virus infections, these studies will help to establish important foundation with which to design vaccines and anti-infective measures against a variety of viral pathogens PUBLIC HEALTH RELEVANCE: While autophagy is an ancient evolutionarily conserved pathway designed to maintain cellular homeostasis by degrading long-lived proteins and organelles in the cytosol, recent studies from our group and others have revealed the role of autophagy in the immune system. In this application, we propose to examine the importance of autophagy in both innate and adaptive antiviral immune responses using well-established genetic, biochemical and cell biological tools as well as in vivo animal models of both RNA and DNA virus infections. The understanding gained from the proposed studies will not only provide scientific advances in how autophagy is utilized by the immune system, but also to help establish critical foundation with which to design immunological interventions and preventative measures against a wide variety of viral diseases.