Role of ISG15 in Inhibition of Ebola Virus Budding

描述（由申请人提供）：埃博拉病毒是a类病毒，是一种新出现的人类病原体，可引起毁灭性的出血热暴发。组装和出芽是埃博拉病毒复制周期中重要的后期事件。虽然对病毒复制的这些动态和多方面的阶段已经了解了很多，但组装和出芽的分子机制仍然不清楚。本课题以埃博拉病毒VP40基质蛋白为研究对象。埃博拉VP40蛋白含有出芽晚期结构域（l结构域），该结构域通过介导与宿主蛋白（如tsg101）的相互作用，在病毒输出中发挥关键作用。我们假设宿主先天免疫反应通过靶向VP40 l结构域活性来抑制埃博拉病毒出芽。初步数据提示干扰素刺激基因15 （ISG15）抑制埃博拉病毒VP40 VLP出芽。为了验证我们的假设，我们提出了3个具体目的：1)确定ISG15的表达是否以l结构域依赖的方式抑制埃博拉病毒VP40 VLPs的释放；2)确定ISG15的表达是否抑制埃博拉病毒或表达埃博拉病毒l结构域的VSV重组体的释放；3)确定ISG15的表达是否抑制埃博拉病毒VP40/宿主tsg101的相互作用。这些研究结果将为了解先天免疫反应的特定成分如何抑制埃博拉病毒出芽提供基础知识，并为进一步阐明这些先天免疫反应对埃博拉病毒感染的机制和生物学意义提供基础。

DESCRIPTION (provided by applicant): Ebola virus is a Category A virus and an emerging human pathogen that causes devastating outbreaks of hemorrhagic fever. Assembly and budding are important late events in the replication cycle of Ebola virus. While much has been learned regarding these dynamic and multifaceted stages of virus replication, the molecular mechanisms of assembly and budding remain unclear. This proposal focuses on the VP40 matrix protein of Ebola virus. Ebola VP40 protein contains late budding domains (L-domain) that play a key role in virus egress by mediating interactions with host proteins, such as tsg101. We hypothesize that host innate immune responses inhibit Ebola virus budding by targeting VP40 L-domain activity. Preliminary data implicate interferon stimulated gene 15 (ISG15) in inhibiting Ebola VP40 VLP budding. To test our hypothesis, we propose 3 Specific Aims: 1) To determine whether expression of ISG15 inhibits release of Ebola VP40 VLPs in an L-domain dependent manner, 2) To determine whether expression of ISG15 inhibits release of Ebola virus or VSV recombinants expressing Ebola virus L-domains, and 3) To determine whether expression of ISG15 inhibits Ebola VP40/host tsg101 interactions. Results from these studies should provide fundamental knowledge of how specific components of the innate immune response may inhibit Ebola virus budding, and provide the foundation for future studies to elucidate further the mechanisms and biological significance of these innate immune responses to Ebola virus infection.