Innate Immune Defenses Against Filoviruses.

描述（由申请人提供）：负义RNA病毒包括丝状病毒、横纹肌病毒和副粘病毒是世界范围内人类多种严重疾病的病因。组装和出芽是这些病毒复制周期中重要的后期事件。虽然关于这些动态和多方面的复制阶段已经了解了很多，但调节病毒组装/出芽的宿主途径仍在继续被确定。丝状病毒和病毒样颗粒（vlp）的组装/出芽主要由基质蛋白VP40驱动。我们和其他人已经证明VP40内的晚出芽结构域（l结构域）对有效出芽很重要，因为它们介导与宿主蛋白（例如Nedd4泛素连接酶）的相互作用，以促进病毒与细胞的分离。值得注意的是，越来越清楚的是，宿主先天免疫反应导致诱导无数调节病毒复制、出芽和发病机制的蛋白质。先天免疫反应是抵御病毒病原体的第一道关键防线，重要的是，埃博拉病毒和马尔堡病毒靶向这种反应的关键调节因子，包括巨噬细胞/单核细胞和树突状细胞。更好地了解宿主先天免疫相互作用和对丝状病毒的反应对于开发治疗和预防感染的疗法和疫苗至关重要。为此，我们最近证明干扰素刺激基因ISG15的表达导致VP40泛素化和VP40 VLPs的抑制。虽然鉴定ISG15抗埃博拉病毒的抗病毒活性机制是重要的第一步，但我们对ISG15和相关isg酰化因子如何抑制埃博拉病毒和可能的马尔堡病毒出芽的理解仍然不完整。本研究的总体目标是确定埃博拉病毒VLPs是否可以通过tlr4依赖途径诱导ifn调控的Herc5 E3连接酶的表达，Herc5 E3连接酶是否有助于靶蛋白（宿主或病毒）的isg酰化，从而抑制埃博拉病毒的出芽，以及宿主泛素化和isg酰化途径是否在调节马尔堡病毒VLPs的出芽中发挥竞争作用。我们相信这些研究将对我们理解丝状病毒蛋白和先天免疫系统成分之间的相互作用产生重大影响，这些相互作用导致病毒输出和可能的疾病进展和严重程度的调节。此外，我们的发现还将影响我们对操纵宿主免疫反应介质如何具有治疗益处的理解，并可能有助于优化对潜在疫苗（例如埃博拉病毒或马尔堡病毒VLPs）和/或疫苗成分（例如埃博拉病毒或马尔堡病毒GP和VP40）的免疫反应。

DESCRIPTION (provided by applicant): Negative-sense RNA viruses including filoviruses, rhabdoviruses and paramyxoviruses are the cause of a multitude of serious diseases in humans worldwide. Assembly and budding are important late events in the replication cycles of these viruses. While much has been learned regarding these dynamic and multifaceted stages of replication, host pathways that modulate virus assembly/budding continue to be identified. Assembly/budding of filoviruses and virus-like particles (VLPs) is driven largely by the matrix protein, VP40. We and others have shown that Late budding domains (L-domains) within VP40 are important for efficient budding, as they mediate interactions with host proteins (e.g. Nedd4 ubiquitin ligase) to facilitate virus-cell separation. Notably, it is becoming clear that host innate immune responses result in the induction of a myriad of proteins that modulate virus replication, budding, and pathogenesis. The innate immune response represents the first critical line of defense against viral pathogens, and importantly, ebolavirus and marburgvirus target key regulators of this response including macrophages/monocytes and dendritic cells. A better understanding of host innate immune interactions and responses to filoviruses will be crucial for developing therapeutics and vaccines to treat and prevent infection. Toward this end, we demonstrated recently that expression of interferon stimulated gene ISG15 resulted in inhibition of VP40 ubiquitination and subsequent egress of VP40 VLPs. Although identification of this previously undescribed mechanism of antiviral activity for ISG15 against ebolavirus was an important first step, our understanding of how ISG15 and associated ISGylation factors inhibit budding of ebolavirus and possibly marburgvirus remains incomplete. The overarching goals of this proposal are to determine whether Ebola VLPs can induce expression of IFN-regulated Herc5 E3 ligase through a TLR4-dependent pathway, whether Herc5 E3 ligase contributes to ISGylation of target proteins (host or viral) resulting in inhibition of ebolavirus budding, and whether host ubiquitination and ISGylation pathways play a competing role in modulating budding of marburgvirus VLPs. We believe that these studies will have a major impact on our understanding of the interplay between filovirus proteins and components of the innate immune system leading to modulation of virus egress and likely disease progression and severity. In addition, our findings will also impact our understanding of how manipulation of host immune response mediators may have therapeutic benefit, and may help to optimize immunological responses to potential vaccines (e.g. ebolavirus or marburgvirus VLPs) and/or vaccine components (e.g. ebolavirus or marburgvirus GP and VP40). PUBLIC HEALTH RELEVANCE: The filoviruses are deadly human pathogens and potential agents of bioterrorism for which no vaccines, nor antivirals exist. The innate immune system represents a first critical line of defense against these viral pathogens. A better understanding of how the host innate immune system recognizes, responds, and defends against filovirus infection will be critical for developing effective therapeutics and vaccines.