Innate Immune Defenses Against Filoviruses.

描述(申请人提供)：负义RNA病毒包括丝状病毒、横纹状病毒和副粘病毒，是世界各地人类多种严重疾病的原因。组装和萌芽是这些病毒复制周期中的重要后期事件。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/萌发的宿主途径仍在继续被识别。丝状病毒和病毒样颗粒(VLP)的组装/萌发主要由基质蛋白VP40驱动。我们和其他人已经证明，VP40中的晚期萌芽结构域(L结构域)对于有效的萌发是重要的，因为它们介导了与宿主蛋白(例如Nedd4泛素连接酶)的相互作用，从而促进病毒与细胞的分离。值得注意的是，宿主的先天免疫反应导致大量蛋白质的诱导，这些蛋白质调节病毒的复制、萌发和致病。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针对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。更好地了解宿主与生俱来的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，我们最近证实，干扰素刺激基因ISG15的表达导致VP40泛素化和随后VP40 VLP的外流受到抑制。尽管鉴定ISG15抗埃博拉病毒的这一先前未被描述的抗病毒活性机制是重要的第一步，但我们对ISG15及其相关ISG化因子如何抑制埃博拉病毒和可能的马尔堡病毒萌发的了解仍然不完整。该建议的主要目标是确定埃博拉病毒VLP是否可以通过依赖TLR4的途径诱导干扰素调节的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化从而抑制埃博拉病毒的出芽，以及宿主泛素化和ISG化途径是否在调节MarburgVirus VLP的出芽过程中发挥竞争作用。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组件之间的相互作用产生重大影响，这些相互作用导致病毒出口的调节以及可能的疾病进展和严重程度。此外，我们的发现还将影响我们对操纵宿主免疫反应介体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VLP)和/或疫苗组件(例如埃博拉病毒或马尔堡病毒GP和VP40)的免疫学反应。 与公共卫生相关：丝状病毒是致命的人类病原体，是生物恐怖主义的潜在病原体，目前既没有疫苗，也没有抗病毒药物。先天免疫系统是抵御这些病毒病原体的第一道关键防线。更好地了解宿主先天免疫系统是如何识别、反应和防御丝状病毒感染的，对于开发有效的治疗方法和疫苗至关重要。

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.