The mammalian small RNA machinery and its role in antiviral defenses

描述(由申请人提供)：了解病毒致病的分子基础是未来抗病毒疗法发展的基本要求。虽然我们知道病毒是如何识别的，以及它们是如何被抗击的，但仍有大量的细胞抗病毒免疫领域尚不清楚。这项研究建议集中在一个这样的领域。这项提议的核心问题是，小RNA或哺乳动物小RNA机制的组成部分是否有助于细胞对感染的反应。这个问题的重要性和从实验上解决这个问题的必要性可以通过大量的初步数据得到证明，这些数据表明，在哺乳动物细胞中存在一种对抗病毒感染的新方法，但还有待表征。为了了解哺乳动物小RNA途径与病毒感染反应之间的相互作用，我们发现了RNA诱导沉默复合体(RISC)抑制病毒复制的效力，分离出了第一个抑制RISC功能的哺乳动物病毒蛋白，并发现在RNA病毒感染过程中，小RNA生物发生中的关键角色DROSHA主动移位到细胞质中。综上所述，我们认为这项研究提供了令人信服的证据，证明哺乳动物中存在一种未被认可的小RNA介导的抗病毒反应。在这里，我们建议使用RISC的病毒拮抗剂(称为VP55)来定义体内感染的小RNA反应，并表征DROSHA如何参与这一抗病毒活性。这项建议的总体目标是描绘小RNA途径如何有助于我们对病毒感染的反应。

DESCRIPTION (provided by applicant): Understanding the molecular basis for why viruses cause disease is an essential requirement for future development of antiviral therapeutics. While we know a great deal about how viruses are recognized and the various ways in which they are combated, there are still vast areas of cellular antiviral immunity that remain unknown. This research proposal focuses on one such area. The central question being addressed in this proposal is whether small RNAs, or components of the mammalian small RNA machinery, contribute to the cellular response to infection. The importance of this question and the need to address it experimentally can be justified through extensive preliminary data that suggest a novel means to combat virus infection is present in mammalian cells but awaits characterization. In an effort to understand the interplay between the small RNA pathway of mammals and the response to virus infection we have discovered the potency by which the RNA induced silencing complex (RISC) can inhibit virus replication, isolated the first mammalian virus protein which inhibits RISC function, and found that Drosha, a key player in small RNA biogenesis, actively translocates to the cytoplasm during RNA virus infection. Taken together, we believe this research provides compelling evidence for an unappreciated small RNA-mediated antiviral response in mammals. Here we propose to use the virus antagonist to RISC (called VP55) to define the small RNA response to infection in vivo as well as characterize how Drosha participates in this antiviral activity. The overall objective of this proposal is to delineate howthe small RNA pathway contributes to our response to virus infection.