Role of Lipid Droplets in Hepatitis C Virus Infection

脂滴在丙型肝炎病毒感染中的作用

• 批准号: 9265409
• 负责人: Melanie Maria Ott
• 金额: $ 47.75万
• 依托单位: J. DAVID GLADSTONE INSTITUTES
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9265409
• 关键词: Acylation; Acyltransferase; Address; Adipocytes; Adipose tissue; Affect; Antiviral Agents; Binding; Cell Culture System; Cells; Chemicals; Cholesterol Esters; Chronic; Chronic Hepatitis C; Complex; Core Protein; Cytoplasm; Dengue Infection; Development; Diglycerides; Endoplasmic Reticulum; Energy-Generating Resources; Enzymes; Epidemic; Gastroenterology; Genotype; Hepatitis C; Hepatocyte; Hepatology; Howard Temin Award; Human; Impairment; Individual; Infectious hepatitides; Integration Host Factors; Life Cycle Stages; Link; Lipase; Lipids; Lipolysis; Liver; Liver Fibrosis; Malignant neoplasm of liver; Manuscripts; Medicine; Membrane; Modeling; Molecular; Morbidity - disease rate; Mus; National Institute of Allergy and Infectious Disease; Nature; Nonstructural Protein; Nucleocapsid; Organelles; Paper; Patients; Phospholipids; Play; Process; Production; Property; Proteins; Publishing; RNA replication; Recruitment Activity; Risk; Risk Factors; Role; Science; Surface; Symptoms; Testing; Therapeutic Intervention; Transgenic Mice; Triglycerides; Viral; Viral Proteins; Virion; Virus; Virus Assembly; Virus Diseases; common symptom; crosslink; diacylglycerol O-acyltransferase; editorial; experimental study; global health; hepatitis C virus nucleocapsid protein; hepatoma cell; inhibitor/antagonist; innovation; insight; liver development; monolayer; mortality; mutant; new therapeutic target; non-alcoholic fatty liver; novel; novel therapeutics; particle; protein acyltransferase; public health relevance; replicase; small molecule inhibitor; tandem mass spectrometry; viral RNA; virus pathogenesis; web site

DESCRIPTION (provided by applicant): The hepatitis C virus (HCV) epidemic is a global health problem and affects ~170 million people worldwide. In ~80% of cases, viral infection becomes chronic, rendering HCV a leading cause of liver-related morbidity and mortality. A common symptom of chronic HCV infection is steatosis, the abnormal accumulation of lipid droplets in the liver. Steatosis is observed in ~55% of chronically HCV-infected patients and represents an important risk factor for the development of liver fibrosis and cancer. The viral nucleocapsid core expressed in livers of transgenic mice recapitulates this condition; core itself also localizes to the surface of lipid droplets (LDs), a process critical for the assembly of progeny virions at membranes in close proximity to LDs. We recently identified the triglyceride-synthesizing enzyme DGAT1 as a novel host factor for HCV assembly (Herker et al, Nat. Med. 2010). In cells lacking DGAT1 or treated with a DGAT1 inhibitor, core cannot localize to LDs and cannot recruit viral RNA to neighboring endoplasmic reticulum membranes for encapsidation. As a consequence, HCV particle production is severely impaired. We recently published three additional studies that demonstrate 1) that a second HCV protein, NS5A, interacts with DGAT1 and requires DGAT1 for LD localization (Camus et al, J. Biol. Chem. 2013), 2) that core at the surface of LDs decreases the lipolysis of these LDs, thereby causing steatosis (Harris, Herker et al, J. Biol. Chem. 2012), and 3) that LDs play a larger role in HCV infection with a new involvement in HCV RNA replication via a new interaction between the LD- associated protein TIP47 and NS5A (Vogt et al, PLoS Pathog. 2013). We seek to study in molecular detail the role of LDs as critical host organelles in the HCV lifecycle. We propose three experiments. 1) To define the role of DGAT1 in recruiting HCV proteins to LDs. We will test the hypothesis that a tripartite complex of DGAT1, NS5A and core has evolved to co-recruit core and NS5A to DGAT1-generated LDs. We will also determine if NS5A and core are new protein targets for the acyltransferase activity of DGAT1. 2) To determine how the HCV core protein inhibits lipolysis. We will test whether core at the surface of LDs interferes with the recruitment or activity of the triglyceride lipases ATGL/PNPLA2 and PNPLA3, thus inhibiting lipolysis. Because patients infected with different HCV genotypes are at different risks to develop steatosis, we will compare anti-lipolytic activities of core proteins from different viral genotypes. 3) To explore how the NS5A-TIP47 interaction regulates HCV RNA replication. We will determine if LDs serve a dual role in HCV infection: one as membrane and energy sources for HCV RNA replication and another as assembly platforms. We will determine whether binding of NS5A to TIP47 plays an important role in this process. Collectively, our proposed studies will bring new molecular insight into the role of LDs in the HCV lifecycle and may uncover potential novel therapeutic strategies to treat chronic HCV disease.

描述（由申请人提供）：丙型肝炎病毒（HCV）流行是一个全球性的健康问题，影响着全世界约1.7亿人。在约80%的病例中，病毒感染变为慢性感染，使HCV成为肝脏相关疾病和死亡的主要原因。慢性丙型肝炎病毒感染的一个常见症状是脂肪变性，即肝脏中脂滴的异常积聚。约55%的慢性丙型肝炎感染患者存在脂肪变性，是肝纤维化和肝癌发展的重要危险因素。转基因小鼠肝脏中表达的病毒核衣壳核心反映了这种情况；核本身也定位于脂滴（LDs）的表面，这一过程对于后代病毒粒子在靠近脂滴的膜上组装至关重要。我们最近发现甘油三酯合成酶DGAT1是HCV组装的一种新的宿主因子（Herker等人，Nat. Med. 2010）。在缺乏DGAT1或用DGAT1抑制剂处理的细胞中，core不能定位到ld，也不能将病毒RNA招募到邻近的内质网膜进行封装。因此，丙型肝炎病毒颗粒的产生受到严重损害。我们最近发表了另外三项研究，证明1)第二种HCV蛋白NS5A与DGAT1相互作用，并且需要DGAT1来定位LD （Camus等，J. Biol）。Chem. 2013), 2) ld表面的核减少了这些ld的脂肪分解，从而导致脂肪变性（Harris， Herker等人，J. Biol.）。3) LD在HCV感染中发挥更大作用，通过LD相关蛋白TIP47和NS5A之间的新相互作用参与HCV RNA复制（Vogt等人，PLoS Pathog. 2013）。我们试图从分子细节上研究ld在HCV生命周期中作为关键宿主细胞器的作用。我们提出三个实验。1)明确DGAT1在募集HCV蛋白到LDs中的作用。我们将验证DGAT1、NS5A和核心的三方复合体已经进化到共同招募核心和NS5A到DGAT1产生的ld的假设。我们还将确定NS5A和core是否是DGAT1酰基转移酶活性的新蛋白靶点。2)确定HCV核心蛋白如何抑制脂肪分解。我们将测试ld表面的核心是否会干扰甘油三酯脂肪酶ATGL/PNPLA2和PNPLA3的募集或活性，从而抑制脂肪分解。由于感染不同HCV基因型的患者发生脂肪变性的风险不同，我们将比较不同病毒基因型核心蛋白的抗脂溶活性。3)探讨NS5A-TIP47相互作用调控HCV RNA复制的机制。我们将确定lld是否在HCV感染中发挥双重作用：一个是HCV RNA复制的膜和能量来源，另一个是组装平台。我们将确定NS5A与TIP47的结合是否在这一过程中发挥重要作用。总的来说，我们提出的研究将为lld在HCV生命周期中的作用提供新的分子视角，并可能揭示治疗慢性HCV疾病的潜在新治疗策略。