Influenza Viral RNA Synthesis and Processing

描述(申请人提供)：项目概述：我的职业目标是在一家研究和教育机构经营一个分子病毒学实验室，最初专注于阐明流感病毒复制所需的生化机制。这项拟议的研究旨在确定流感病毒RNA合成和加工所需的病毒和细胞蛋白的作用。实验旨在探索突变病毒蛋白的功能，并确定流感病毒传播中涉及的病毒与宿主之间的相互作用。具体目标1将在体外和体内检测突变流感蛋白的RNA合成和加工活性，以确定病毒-病毒和病毒-宿主蛋白相互作用对病毒繁殖至关重要的作用。实验将探讨流感核衣壳蛋白(NP)和RNA依赖RNA聚合酶(RDRP)接触的作用，以及两种已知的病毒与宿主之间的相互作用；流感RDRP与细胞RNA聚合酶II的C末端结构域(CTD)，以及流感NP与宿主细胞剪接和输出因子UAP56。具体目标2将确定被流感病毒mRNAs劫持的宿主剪接和核输出因子。已知的细胞输出途径将在流感病毒感染期间进行功能检查。为了发现新的细胞核输出途径，将通过突变筛选确定病毒mRNAs中需要输出病毒mRNA的顺式区域。然后，发现的核糖核酸元件(S)将被用来鉴定与病毒核糖核酸特异结合的细胞蛋白质。病毒信使核糖核酸的剪接和输出是流感病毒知之甚少的领域。这项拟议的研究将提供筛选和发现抑制流感病毒感染的化合物的方法，并可能为抗病毒疗法的开发奠定基础。此外，这项研究有可能发现新的细胞RNA加工和核输出途径。相关性：流感是人类面临的一个紧迫的健康问题。对于一种快速出现的高致病性病毒，最好的防线是抗病毒药物，尽管耐药性使寻找新的抗病毒靶标成为一场持续的斗争。这项拟议的研究旨在了解流感病毒感染所需的分子机制，为开发新的抗病毒疗法奠定基础。

DESCRIPTION (provided by applicant): Project Summary: My career objective is to run a molecular virology laboratory at a research and education institute initially focused on elucidating the biochemical mechanisms required for influenza viral replication. The proposed research aims to define the roles of viral and cellular proteins required for influenza viral RNA synthesis and processing. Experiments are designed to probe function of mutant viral proteins and identify virus-host interactions involved in influenza virus propagation. Specific Aim 1 will examine the RNA synthesis and processing activities of mutant influenza proteins, both in vitro and in vivo, to define the role of viral - viral and viral - host protein interactions essential to viral propagation. Experiments will probe the role of influenza nucleocapsid protein (NP) and RNA dependent RNA polymerase (RdRP) contacts, and two identified viral - host interactions; the influenza RdRP with the C-terminal domain (CTD) of cellular RNA polymerase II, and influenza NP with host cellular splicing and export factor UAP56. Specific Aim 2 will identify host splicing and nuclear export factors hijacked by influenza viral mRNAs. Known cellular export pathways will be examined for their function during influenza virus infection. To discover novel cellular nuclear export pathways, cis regions within the viral mRNAs that are required for viral mRNA export will be defined through mutagenesis screening. The discovered RNA element(s) will then be utilized to identify cellular proteins which specifically bind the viral RNA. Viral mRNA splicing and export is an area poorly understood for influenza viruses. The proposed research will provide assays to screen and discover compounds which inhibit influenza virus infection and may lay foundation for development of antiviral therapies. Further, this research has potential to discover novel cellular RNA processing and nuclear export pathways. Relevance: Influenza is a pressing health issue for humans. The best line of defense for a rapidly emerging highly pathogenic virus is anti-viral medications, although resistance makes the search for new anti-viral targets a constant struggle. The proposed research aims to understand molecular mechanisms required for influenza viral infection with the intent to lay the foundation for development of novel anti-viral therapies.