Immune Evasion of Autophagy in Gamma-Herpesviruses infection

描述(申请人提供)：3-疱疹病毒(3-HV)已经发展出一种与宿主相互作用的独特模式，以建立一种终身持续感染，这经常与各种恶性肿瘤的发生有关。参与3-HV持久性和致癌性的一个关键毒力因子是所有3-HV编码的Bcl2蛋白(称为vBcl2)的病毒同源物。除了已知的抗细胞凋亡活性外，我们的初步研究还证实，3-HV家族的vBcl2通过直接靶向关键的自噬效应蛋白Beclin1，有效地抑制了抗病毒自噬途径(自噬、溶酶体依赖的降解和细胞内成分对应激的循环)。此外，与宿主相比，vBcl2已经进化出更强的抗自噬活性。基于这些发现，我们推测vBcl2对自噬的抑制构成了3-HV逃避宿主免疫并导致持续感染和致病的新机制。为了验证这一假设，我们将主要关注3HV68的vBcl2，使用成熟的体外和体内系统。3HV68与EBV和KSHV具有广泛的遗传同源性和生物学相似性。3HV-68感染小鼠为研究3HV的慢性感染提供了一种在体遗传学上易处理的模型。值得注意的是，vBcl-2的丢失并不影响3HV68的裂解复制。相反，它严重削弱了3HV68在小鼠身上建立慢性感染的能力。在第一个目标中，我们将剖析vBcl2拮抗Beclin1依赖的自噬的分子机制。我们已经成功地鉴定了区分vBcl2介导的抑制自噬和vBcl2介导的拮抗凋亡的特定突变。在第二个目标中，我们将研究vBcl2介导的抗自噬在体内病毒毒力中的具体作用。从这项研究中获得的见解将揭示vBcl2在3Hs感染中的作用的新范式，并建立自噬作为抵抗病毒感染的基本宿主防御机制。公共卫生相关性：越来越多的人认识到自噬在宿主抗病毒防御中是必不可少的，但它在3-疱疹病毒感染中的作用在很大程度上仍不清楚。本研究旨在了解vBcl2介导的抑制自噬的分子机制及其在3-疱疹病毒持续感染和/或发病机制中的作用。从这项研究中获得的见解将确立自噬在病毒毒力控制中的直接作用，并为急需的抗病毒疗法提供战略建议。

DESCRIPTION (provided by applicant): 3-Herpesviruses (3-HVs) have developed a unique mode of interaction with their hosts to establish a life-long persistent infection, which frequently associates with the onset of various malignancies. One critical virulence factor involved in 3-HVs persistence and oncogenicity are the viral homologs of the Bcl-2 protein (referred to as vBcl-2) encoded by all 3-HVs. Alongside its well-characterized anti-apoptotic activity, our preliminary studies have established that the vBcl-2 of the 3-HV family effectively suppresses the anti-viral autophagy pathway ('self-eating', lysosome-dependent degradation and recycling of the intracellular components in response to stress), by directly targeting a key autophagy effector protein, Beclin1. Moreover, vBcl-2 has evolved enhanced anti-autophagic activity when compared to the host counterpart. Based on these findings, we hypothesize that the inhibition of autophagy by vBcl-2 constitutes a novel mechanism by which 3- HVs evade host immunity and confer persistent infection and pathogenesis. To test this hypothesis, we will focus primarily on the vBcl-2 of 3HV68 using well-established in vitro and in vivo systems. 3HV68 shares extensive genetic homology and biological similarity with EBV and KSHV. Infection of mice with 3HV68 provides a genetically tractable in vivo model for characterizing the chronic infection of 3HVs. Notably, the loss of vBcl-2 does not affect the lytic replication of 3HV68. Instead, it severely impairs the ability of 3HV68 to establish chronic infection in mice. In the first aim, we will dissect the molecular mechanism by which vBcl-2 antagonizes Beclin1-dependent autophagy. We have successfully identified the specific mutations that distinguish vBcl-2-mediated inhibition of autophagy from vBcl-2-mediated antagonism of apoptosis. In the second aim, we will investigate the specific roles of vBcl-2-mediated anti-autophagy in viral virulence in vivo. Insights gained from this study will reveal a novel paradigm for the roles of vBcl-2 in 3HVs infection, and establish autophagy as a fundamental host defensive mechanism against viral infections. PUBLIC HEALTH RELEVANCE: Autophagy has been increasingly recognized essential in host anti-viral defense, but its role in 3- herpesviruses infection remains largely unknown. The proposed study is targeted to understand the molecular mechanism of vBcl-2-mediated inhibition of autophagy, and its contribution to the persistent infection and/or pathogenesis of 3-herpesviruses. Insights gained from this study will establish a direct role for autophagy in viral virulence control and suggest strategy for much- needed anti-viral therapeutics.