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(申请人提供)：蛋白质功能由翻译后事件动态控制。一种常见的调控机制是靶蛋白的磷酸化和随后的构象重排。病毒蛋白通常由与宿主编码的目标蛋白相同的途径控制。甲型流感病毒的复制周期受宿主蛋白的影响，并提供了一个独特的系统来研究这些过程。一个主要的调控目标是包含病毒RNA、核蛋白(NP)和由蛋白质PB1、PB2和PA组成的三聚体聚合酶的流感病毒复制机制。复制机制控制着从基因表达到基因组复制的有序过渡，这对生产性感染是必不可少的。此外，病毒聚合酶是流感病毒宿主范围内的关键决定因素，并限制流感病毒从禽类传播到人类。人们对控制复制机制的过程以及聚合酶和宿主之间的相互作用知之甚少。我们提出了一种综合的方法，利用生化、遗传和结构研究来确定调节流感复制机制的宿主蛋白和分子机制。具体地说，AIM 1将确定病毒蛋白中的磷酸化位置，并检查磷酸化对病毒复制和宿主范围确定的功能后果。AIM 2将使用遗传作图和高分辨率晶体结构来详细表征复制复合体的蛋白质界面，并阐明伴随着复合体组装的构象变化。最后，Aim 3将利用功能丧失筛查来识别调节流感聚合酶并控制流感从禽类传播到人类的细胞蛋白。重要的是，每个目标都提供了培训新技术的重要机会，这些新技术对应用程序的独立阶段至关重要。这些研究将为控制流感复制的病毒和宿主因素提供重要的洞察，并将为合理的战略提供基础，以治疗和预防未来在人类中爆发的流感。 相关性：流感病毒复制机制是确定感染和确定其致病性的关键因素。这些研究将深入了解流感复制机制的调控，以及它如何控制流感病毒从禽类传播到人类，并最终可能确定新的治疗干预目标。