Autophagy in Antiviral Immunity

描述（由申请人提供）：自噬是一种古老的进化上保守的途径，旨在通过降解胞质溶胶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，自噬被先天性和适应性免疫系统的细胞利用来对抗病毒感染。病毒的天然识别通过两种不同的途径发生。在专业的病毒传感器中，浆细胞样树突状细胞（pDC），通过Toll样受体（TLR）7和9在内体中识别病毒。我们最近的工作已经证明，自噬通过TLR 7在识别pDC中的病毒感染的特征中起关键作用。与pDC相反，身体的大多数其他细胞类型通过RIG-I样受体（RLR）家族利用病毒复制的胞质传感器。参与自噬的分子已经显示出阻断RLR信号传导。此外，最近的报道表明，自噬将内源性病毒抗原递送至MHC II类装载区室，从而允许活化CD 4 T细胞。然而，在体内病毒感染过程中这些途径的相关性是未知的。在本申请中，我们提出了初步的数据，揭示了需要Atg 5，一个关键的分子需要形成的自噬体，在转导信号通过TLR 9导致激活的I型IFN基因在pDC后单纯疱疹病毒（HSV; TLR 9激动剂）感染。此外，我们发现，自噬负调控非浆细胞样树突状细胞的RLR途径后，水泡性口炎病毒感染（VSV; RIG-I激动剂）。最后，我们证明了一个关键的自噬作用在体内的处理和介绍的各种形式的抗原MHC II类在树突状细胞HSV-1感染后。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研究自噬在先天性和适应性免疫应答中的重要性。在第一个目标中，我们将通过使用分子和细胞生物学技术来确定在pDC中HSV感染后Atg 5和/或自噬通过TLR 9介导信号传导的机制。在第二个目标中，我们建议通过蛋白质组学和生物化学方法来确定自噬如何调节VSV感染后RNA传感器的激活。在最后的目标，我们将询问树突状细胞如何利用自噬的处理和介绍细胞外病毒抗原在体外和体内的小鼠选择性缺乏自噬的树突状细胞群体。通过提供自噬如何协调针对病毒感染的先天性和适应性免疫的产生的基本理解，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 公共卫生相关性：虽然自噬是一种古老的进化保守途径，旨在通过降解胞质溶胶中的长寿蛋白质和细胞器来维持细胞稳态，但我们小组和其他人最近的研究揭示了自噬在免疫系统中的作用。在本申请中，我们建议使用完善的遗传，生物化学和细胞生物学工具以及RNA和DNA病毒感染的体内动物模型来研究自噬在先天性和适应性抗病毒免疫应答中的重要性。从拟议的研究中获得的理解不仅将为免疫系统如何利用自噬提供科学进展，而且还有助于建立关键基础，以设计针对各种病毒性疾病的免疫干预和预防措施。

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.