Roles for RCK/DDX6 in hepatitis C virus pathogenesis and hepatocellular carcinoma

RCK/DDX6 在丙型肝炎病毒发病机制和肝细胞癌中的作用

• 批准号: 7698228
• 负责人: PETER SARNOW
• 金额: $ 29.58万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-17 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7698228
• 关键词: Affect; Alcohol abuse; Antiviral Therapy; Biochemical; Biological Assay; Boxing; Cancer Etiology; Cell physiology; Cells; Cessation of life; Chronic; Cirrhosis; Cytolysis; Cytoplasmic Granules; Degradation Pathway; Detection; Disease; Economic Burden; Family; Fractionation; Gene Amplification; Gene Expression; Gene Expression Regulation; Genome; Goals; Hepatitis B; Hepatitis C; Hepatitis C virus; Hepatocyte; Immunoprecipitation; In Situ Hybridization; Individual; Infection; Interferons; Intravenous; Lead; Life; Liver; Liver diseases; Malignant neoplasm of liver; Mammalian Cell; Measures; Mediating; MicroRNAs; Monitor; Nature; Oligonucleotides; Pathway interactions; Patients; Plasmids; Primary carcinoma of the liver cells; Proteins; RNA; RNA chemical synthesis; Ribavirin; Ribosomes; Role; Staging; Stress; Structure; System; Technology; Toxic Environmental Substances; Transfection; Translation Initiation; UV induced; Up-Regulation; Viral; Viral Genome; Viral Proteins; Virus Replication; cancer cell; cellular targeting; cholesterol biosynthesis; combat; crosslink; helicase; member; new technology; novel; overexpression; protein complex; reconstitution; research study; tool; viral RNA; virus pathogenesis

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths. Although many cases of HCC are induced by chronic alcohol abuse or environmental toxins, approximately 80% are caused by infection with either hepatitis B or hepatitis C virus (HCV). The burden of HCV infection worldwide is very large, with most of the personal and economic burden yet to come, as cirrhosis and HCC take years to develop. Approximately 170 million people are infected worldwide with HCV. Most infected individuals do not clear the disease, but develop chronic infections that often lead to end-stage liver disease and HCC. Current treatment is limited to co-treatment with ribavirin and interferon , a therapy that is expensive and ineffective in 50% of infected individuals. Therefore, there is an urgent need to identify new and accessible viral and cellular targets for therapies against HCV and HCC. It has been shown recently that cellular microRNAs, especially those in the liver, can be readily inactivated by direct intravenous delivery of modified oligonucleotides. Thus, our finding that liver-specific microRNA miR-122 interacts directly with the viral 5’ noncoding region of HCV and that this interaction is essential to maintain intracellular abundance of the viral RNA, identifies a promising target for antiviral therapy. What are the host proteins required for microRNA expression in the liver? Recently, it has been found that host cell protein RCK/DDX6, a member of the DEAD box helicase family, is important for microRNA-mediated gene regulation in mammalian cells and that both miR-122 and RCK are greatly overexpressed in HCV-induced HCC in patient livers. Our preliminary results have shown that RCK is an essential positive regulator of HCV gene expression; depletion of RCK results in loss of HCV proteins. In this proposal, we propose to examine the mechanism by which RCK upregulates HCV gene expression at an apparently post-transcriptional step and by which RCK affects microRNA abundance and intracellular distribution in uninfected and HCV-infected liver cells. Using novel in situ hybridization technology, we will determine the effect of RCK expression on microRNA localization to specific cytoplasmic compartments known as P-bodies and stress granules. Finally, we propose experiments to identify targets for microRNAs whose abundance and distribution is affected by RCK, These studies will develop novel tools for HCV and HCC and are likely to point to new targets for antiviral therapy.

肝细胞癌（HCC）是癌症相关死亡的主要原因。虽然许多HCC病例是由慢性酒精滥用或环境毒素引起的，但约80%是由B型肝炎或丙型肝炎病毒（HCV）感染引起的。世界范围内HCV感染的负担非常大，大部分个人和经济负担尚未到来，因为肝硬化和HCC需要数年时间才能发展。全世界约有1.7亿人感染HCV。大多数感染者不能清除疾病，但发展为慢性感染，通常导致终末期肝病和HCC。目前的治疗仅限于与利巴韦林和干扰素α的联合治疗，这种治疗昂贵且对50%的感染者无效。因此，迫切需要确定新的和可获得的病毒和细胞靶点，用于治疗HCV和HCC。最近已经表明，细胞microRNA，特别是肝脏中的那些，可以通过直接静脉内递送修饰的寡核苷酸而容易地失活。因此，我们发现肝脏特异性microRNA miR-122直接与HCV的病毒5'非编码区相互作用，并且这种相互作用对于维持病毒RNA的细胞内丰度是必不可少的，这确定了抗病毒治疗的有希望的靶点。肝脏中microRNA表达所需的宿主蛋白质是什么？最近，已经发现宿主细胞蛋白RCK/DDX 6（DEAD box解旋酶家族的成员）对于哺乳动物细胞中microRNA介导的基因调控是重要的，并且miR-122和RCK在患者肝脏中HCV诱导的HCC中均显著过表达。我们的初步研究结果表明，RCK是一个重要的正调节HCV基因表达的RCK的缺失导致HCV蛋白的损失。在这个建议中，我们建议检查的机制，RCK上调HCV基因的表达在一个明显的转录后步骤，RCK影响microRNA丰度和细胞内分布在未感染和HCV感染的肝细胞。使用新的原位杂交技术，我们将确定RCK表达对microRNA定位到特定的细胞质隔室（称为P体和应力颗粒）的影响。最后，我们提出了实验来确定其丰度和分布受RCK影响的microRNA的靶点，这些研究将为HCV和HCC开发新的工具，并可能为抗病毒治疗指出新的靶点。