Subversion of hepatocyte phosphoinositide metabolism by hepatitis C virus

丙型肝炎病毒破坏肝细胞磷酸肌醇代谢

• 批准号: 8634774
• 负责人: Andrew W. Tai
• 金额: $ 33.82万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-03-13 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8634774
• 关键词: 1-Phosphatidylinositol 4-Kinase; Accounting; Adverse drug effect; Affect; Antiviral Agents; Area; Binding; Binding Proteins; Biochemical; Cells; Cellular Membrane; Cholesterol; Chronic Hepatitis C; Clinical; DNA Sequence Rearrangement; Drug Targeting; Family Picornaviridae; Generations; Goals; Health Benefit; Hepatitis C; Hepatitis C virus; Hepatitis C-Like Viruses; Hepatocyte; Human; Imaging Techniques; Individual; Integration Host Factors; Internet; Lead; Life; Life Cycle Stages; Lipids; Liver Failure; Liver diseases; Membrane; Membrane Proteins; Metabolism; Modeling; Molecular; Names; PIK4CB gene; Peripheral; Pharmaceutical Preparations; Phosphatidylinositols; Phosphotransferases; Physiological; Population; Primary carcinoma of the liver cells; Proteins; Public Health; RNA Interference; RNA Viruses; RNA replication; Recruitment Activity; Regulation; Research; Role; Serine Protease; Structure; Testing; Therapeutic Agents; United States National Institutes of Health; Viral; Viral Proteins; Virus; Virus Replication; base; cofactor; design; innovation; inorganic phosphate; insight; liver transplantation; membrane activity; novel; novel strategies; novel therapeutics; oxysterol binding protein; particle; phosphatidylinositol 4-phosphate; public health relevance; therapeutic development; treatment response; viral resistance

DESCRIPTION (provided by applicant): Chronic hepatitis C virus (HCV) infection develops in 70-80% of all exposed individuals and affects 3% of the world's population. While new drugs that directly target the viral NS3-4A serine protease have been approved for clinical use this year, there remains a need for more effective and better-tolerated HCV therapies. Our long- term goals are to define host-HCV molecular interactions and in so doing, to identify novel therapeutic agents for HCV therapy. HCV, like all other positive-sense single-stranded RNA viruses studied to date, induces the formation of specific membranous structures (called membranous webs) within infected cells for its own replication. Multiple RNAi screens have identified a critical role for a host protein phosphatidylinositol 4-kinase (PI 4-kinase) named PI4KA in HCV replication, most likely at the level of membranous web formation. Intriguingly, three other positive-sense single-stranded RNA viruses subvert the related PI 4-kinase PI4KB for their own replication. The objective of this proposal is to understand how the host hepatocyte protein PI4KA and its product PI 4-phosphate (PI(4)P) support HCV replication. Our central hypothesis is that HCV replication requires the local generation of PI(4)P, which in turn is necessary for the recruitment of effector proteins to the nascent membranous web. The specific aims of the project are to (1) Define the functional domains of PI4KA and its regulation; (2) Identify the phosphoinositide-dependent steps in the HCV life cycle; (3) Identify and characterize downstream effectors of PI4KA and PI(4)P in the HCV life cycle. We will use a variety of biochemical, cell-based, and imaging techniques to accomplish the proposed research. The proposed research is innovative as (1) the model of web assembly presented in this proposal is novel; (2) the mechanisms by which PI4KA is regulated in the host cell and in the context of HCV replication have not been studied (Aim 1); (3) The mechanisms by which PI(4)P and other phosphoinositides support HCV replication has not been previously pursued (Aims 2 and 3); (4) The use of inducible and selective depletion of phosphoinositides (Aim 2) has not been previously used to study viral replication. We expect that the proposed research may have direct public health benefits, as a better understanding of the mechanisms by which PI4KA and host phosphoinositides support viral replication may lead to novel approaches to the treatment of chronic HCV infection. A better understanding of the HCV-host relationship is a stated goal of the NIH Action Plan for Liver Disease Research (Goal B2a). Moreover, completion of the proposed aims will also lead to new insights into the physiological functions and regulation of PI4KA, areas that are currently poorly understood.

描述(由申请人提供)：慢性丙型肝炎病毒(丙型肝炎病毒)感染发生在所有暴露的个人中的70%-80%，影响世界人口的3%。虽然直接针对病毒NS3-4A丝氨酸蛋白酶的新药今年已获准临床使用，但仍然需要更有效和更耐受性更好的丙型肝炎病毒疗法。我们的长期目标是定义宿主-丙型肝炎病毒的分子相互作用，并通过这样做，为丙型肝炎病毒治疗寻找新的治疗剂。像迄今为止研究过的所有其他正向单链RNA病毒一样，丙型肝炎病毒会在感染细胞内诱导形成特定的膜结构(称为膜网)，以进行自身复制。多个RNAi筛选发现，一种名为PI4KA的宿主蛋白磷脂酰肌醇4-激酶(PI 4-Kinase)在丙型肝炎病毒复制中发挥关键作用，最有可能的是在膜网形成水平上。有趣的是，另外三种正向单链RNA病毒颠覆了相关的PI 4-激酶PI4KB进行自身复制。这项建议的目的是了解宿主肝细胞蛋白PI4KA及其产物PI(4)P如何支持丙型肝炎病毒复制。我们的中心假设是，丙型肝炎病毒复制需要局部产生PI(4)P，而PI(4)P又是效应蛋白招募到新生膜网所必需的。该项目的具体目标是：(1)确定PI4KA及其调控的功能域；(2)确定丙型肝炎病毒生命周期中依赖肌醇磷脂的步骤；(3)识别和表征丙型肝炎病毒生命周期中PI4KA和PI(4)P的下游效应分子。我们将使用各种生化技术、基于细胞的技术和成像技术来完成拟议的研究。拟议的研究具有创新性，因为(1)本提案中提出的网络组装模型是新颖的；(2)PI4KA在宿主细胞中和在丙型肝炎病毒复制的背景下被调控的机制尚未被研究(目标1)；(3)PI(4)P和其他磷脂酰肌醇支持丙型肝炎病毒复制的机制以前从未被研究过(目标2和3)；(4)以前从未使用可诱导的和选择性地去除磷脂酰肌醇(目标2)来研究病毒复制。我们预计，这项拟议的研究可能会对公众健康产生直接的好处，因为更好地了解PI4KA和宿主磷脂酰肌醇支持病毒复制的机制可能会导致治疗慢性丙型肝炎病毒感染的新方法。更好地了解丙型肝炎病毒与宿主的关系是《美国国立卫生研究院肝病研究行动计划》(目标B2a)的明确目标。此外，完成拟议的目标还将导致对PI4KA的生理功能和调节的新见解，这是目前知之甚少的领域。