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: 8728531
• 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/8728531
• 关键词: 1,2-diacylglycerol; Acylation; Acyltransferase; Address; Adipocytes; Adipose tissue; Affect; Antiviral Agents; Binding; Cell Culture System; Cells; Chemicals; Cholesterol Esters; Chronic; Chronic Hepatitis C; Commit; Complex; Core Protein; Cytoplasm; Dengue Virus; Development; Diglycerides; Endoplasmic Reticulum; Energy-Generating Resources; Enzymes; Epidemic; Gastroenterology; Genotype; Hepatitis C; Hepatitis C virus; Hepatocyte; Hepatology; Howard Temin Award; Human; Individual; Infectious hepatitides; Integration Host Factors; Life Cycle Stages; Link; Lipase; Lipids; Lipolysis; Liver; Liver Fibrosis; Liver diseases; 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; Relative (related person); Risk; Risk Factors; Role; Science; Surface; Symptoms; Testing; Therapeutic Intervention; Transgenic Mice; Triglycerides; Viral; Viral Proteins; Virion; Virus Assembly; Virus Diseases; crosslink; diacylglycerol O-acyltransferase; editorial; global health; hepatitis C virus nucleocapsid protein; hepatoma cell; inhibitor/antagonist; innovation; insight; monolayer; mortality; mutant; new therapeutic target; non-alcoholic fatty liver; novel; novel therapeutics; particle; protein acyltransferase; public health relevance; replicase; research study; small molecule; 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，影响全球约1.7亿人。在约80%的病例中，病毒感染变为慢性，使HCV成为肝脏相关发病率和死亡率的主要原因。慢性丙型肝炎病毒感染的常见症状是脂肪变性，即肝脏中脂滴的异常积聚。脂肪变性在约55%的慢性HCV感染患者中观察到，并且是肝纤维化和癌症发展的重要风险因素。在转基因小鼠的肝脏中表达的病毒核衣壳核心重现了这种情况;核心本身也定位于脂滴（LD）的表面，这是子代病毒体在靠近LD的膜处组装的关键过程。我们最近鉴定了合成组氨酸的酶DGAT 1作为HCV组装的新宿主因子（Herker等，Nat.Med.2010）。在缺乏DGAT 1或用DGAT 1抑制剂处理的细胞中，核心不能定位于LD，并且不能将病毒RNA募集到邻近的内质网膜上进行胞苷化。因此，HCV颗粒的产生严重受损。我们最近发表了另外三项研究，证明1）第二种HCV蛋白NS 5A与DGAT 1相互作用，并需要DGAT 1进行LD定位（Camus等，J.Biol.Chem.2013），2）LD表面的核减少了这些LD的脂解，从而引起脂肪变性（Harris，Herker等，J.Biol.Chem.2012），和3）LD在HCV感染中发挥更大的作用，通过LD相关蛋白TIP 47和NS 5A之间的新的相互作用新参与HCV RNA复制（沃格特等，PLoS Pathog. 2013年）。我们试图在分子上详细研究LDs作为HCV生命周期中关键宿主细胞器的作用。我们提出了三个实验。1)明确DGAT 1在招募HCV蛋白至LD中的作用。我们将检验以下假设：DGAT 1、NS 5A和核心的三方复合物已经进化为共同募集核心和NS 5A至DGAT 1产生的LD。我们还将确定NS 5A和核心是否是DGAT 1酰基转移酶活性的新蛋白质靶标。2)确定HCV核心蛋白如何抑制脂解。我们将测试LD表面的核心是否干扰甘油三酯脂肪酶ATGL/PNPLA 2和PNPLA 3的募集或活性，从而抑制脂解。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肪分解活性。3)探讨NS 5A-TIP 47相互作用如何调控HCV RNA复制。我们将确定LD是否在HCV感染中起双重作用：一个作为HCV RNA复制的膜和能量来源，另一个作为组装平台。我们将确定NS 5A与TIP 47的结合是否在这一过程中起重要作用。总的来说，我们提出的研究将带来新的分子洞察LD在HCV生命周期中的作用，并可能发现潜在的新的治疗策略来治疗慢性HCV疾病。