Regulating Influenza Polymerase Structure and Function by Phosphorylation

通过磷酸化调节流感聚合酶的结构和功能

• 批准号: 7714644
• 负责人: Andrew Mehle
• 金额: $ 9万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-07 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7714644
• 关键词: Affect; Avian Influenza; Avian Influenza A Virus; Binding; Biochemical Genetics; Biological Assay; Birds; Cell physiology; Cells; Chromosome Mapping; Complex; Coupled; DNA Sequence Rearrangement; Data; Defect; Development; Ensure; Event; Foundations; Future; Gene Expression; Genes; Genetic; Genetic Transcription; Genome; Genomics; Holoenzymes; Hour; Human; Infection; Influenza; Influenza A virus; Integration Host Factors; Libraries; Life Cycle Stages; Macromolecular Complexes; Mass Spectrum Analysis; Messenger RNA; Molecular; Molecular Structure; Nucleoproteins; Pathogenicity; Pathway interactions; Pattern; Phase; Phosphorylation; Phosphorylation Site; Phosphotransferases; Play; Polymerase; Population; Principal Investigator; Process; Production; Proteins; Public Health; RNA; Regulation; Resolution; Role; Small Interfering RNA; Staging; Stimulus; Structure; System; Techniques; Therapeutic Intervention; Training; Translations; Tropism; Viral; Viral Proteins; Virion; Virus; Virus Replication; base; design; flu transmission; functional genomics; influenza outbreak; influenzavirus; insight; loss of function; mutant; prevent; protein function; protein structure; research study; response; small hairpin RNA; transmission process; viral RNA; virus host interaction

DESCRIPTION (provided by applicant): Protein function is dynamically controlled by post-translation events. A common regulatory mechanism is the phosphorylation and subsequent conformational rearrangement of target proteins. Viral proteins are often controlled by the same pathways that target proteins encoded by the host. The replication cycle of influenza A virus is influenced by host proteins and provides a unique system in which to study these processes. A major target of regulation is the influenza virus replication machinery containing viral RNA, nucleoprotein (NP), and the trimeric polymerase composed of the proteins PB1, PB2, and PA. The replication machinery controls the ordered transition from gene expression to genome replication that is essential for a productive infection. In addition, the viral polymerase is a key determinant in the host range of influenza virus and restricts the transmission of influenza virus from avian to human populations. The processes controlling the replication machinery and the interplay between the polymerase and the host are poorly understood. We propose an integrative approach using biochemical, genetic, and structural studies to determine the host proteins and molecular mechanisms that regulate the influenza replication machinery. Specifically, Aim 1 will identify sites of phosphorylation within viral proteins and examine the functional consequences of phosphorylation on virus replication and the determination of host range. Aim 2 will use genetic mapping and high-resolution crystal structures to characterize in detail the protein interfaces of the replication complex and elucidate the conformational changes that accompany complex assembly. Finally, Aim 3 will exploit loss-of-function screens to identify cellular proteins that regulate the influenza polymerase and control transmission of influenza from birds to humans. Importantly, each Aim presents significant opportunities for training in new techniques essential for the Independent phase of the application. These studies will provide crucial insight into the viral and host factors controlling influenza replication and will provide the foundation for rational strategies to treat and prevent future influenza outbreaks in humans. Relevance: The influenza virus replication machinery is a key player in establishing infection and determining its pathogenicity. These studies will provide insight into the regulation of the influenza replication machinery, how it controls transmission of influenza virus from birds to humans, and may ultimately identify new targets for therapeutic intervention.

描述（由申请人提供）：蛋白质功能由翻译后事件动态控制。常见的调节机制是靶蛋白的磷酸化和随后的构象重排。病毒蛋白通常由靶向宿主编码的蛋白质的相同途径控制。流感的复制周期受宿主蛋白的影响，并提供了研究这些过程的独特系统。调节的主要目标是含有病毒RNA，核蛋白（NP）的流感病毒复制机制和由蛋白质PB1，PB2和PA组成的三聚聚合酶。复制机制控制着从基因表达到基因组复制的有序过渡，这对于生产性感染至关重要。此外，病毒聚合酶是流感病毒宿主范围的关键决定因素，并限制了流感病毒从鸟类到人类种群的传播。控制复制机制以及聚合酶与宿主之间的相互作用的过程对此很众所周知。我们提出了一种使用生化，遗传和结构研究的综合方法，以确定调节流感复制机制的宿主蛋白和分子机制。具体而言，AIM 1将识别病毒蛋白质中磷酸化的位点，并检查磷酸化对病毒复制和宿主范围的测定的功能后果。 AIM 2将使用遗传学映射和高分辨率晶体结构来详细表征复制复合物的蛋白质界面，并阐明伴随复合物组装的构象变化。最后，AIM 3将利用功能丧失的筛选，以鉴定调节流感聚合酶的细胞蛋白，并控制流感从鸟类到人类的传播。重要的是，每个目标都为应用程序的独立阶段至关重要的新技术提供了重要的机会。这些研究将为控制流感复制的病毒和宿主因素提供关键的见解，并为治疗和防止人类未来流感疫情的理性策略奠定了基础。 相关性：流感病毒复制机制是建立感染和确定其致病性的关键参与者。这些研究将提供有关流感复制机制调节的洞察力，如何控制流感病毒从鸟类到人类的传播，并最终可能确定用于治疗干预的新目标。