Hepatitis C virus NS3 protein can activate the Notch-signaling pathway through binding to a transcription factor, SRCAP.

Persistent infections of hepatitis C virus (HCV) are known to be a major risk factor for causing hepatocellular carcinomas. Nonstructural protein 3 (NS3) of HCV has serine protease and RNA helicase domains, and is essential for the viral replication. Further, NS3 is also considered to be involved in the development of HCV-induced hepatocellular carcinomas. In this report, we focus on the function of NS3 protein, and propose a novel possible molecular mechanism which is thought to be related to the tumorigenesis caused by the persistent infection of HCV. We identified SRCAP (Snf2-related CBP activator protein) as a NS3 binding protein using yeast two-hybrid screening, and a co-immunoprecipitation assay demonstrated that NS3 can bind to SRCAP in mammalian cells. The results of a reporter gene assay using Hes-1 promoter which is known to be a target gene activated by Notch, indicate that NS3 and SRCAP cooperatively activate the Hes-1 promoter in Hep3B cells. In addition, we show in this report that also p400, which is known as a protein closely resembling SRCAP, would be targeted by NS3. NS3 exhibited binding activity also to the 1449–1808 region of p400 by a co-immunoprecipitation assay, and further the activation of the Notch-mediated transcription of Hes-1 promoter by NS3 decreased significantly by the combined silencing of SRCAP and p400 mRNA using short hairpin RNA. These results suggest that the HCV NS3 protein is involved in the activation of the Notch-signaling pathway through the targeting to both SRCAP and p400.

已知丙型肝炎病毒（HCV）的持续性感染是导致肝细胞癌的一个主要危险因素。HCV的非结构蛋白3（NS3）具有丝氨酸蛋白酶和RNA解旋酶结构域，对病毒复制至关重要。此外，NS3也被认为参与了HCV诱导的肝细胞癌的发生发展。在本报告中，我们聚焦于NS3蛋白的功能，并提出一种新的可能的分子机制，该机制被认为与HCV持续性感染导致的肿瘤发生有关。我们利用酵母双杂交筛选鉴定出SRCAP（Snf2相关的CBP激活蛋白）是一种与NS3结合的蛋白，并且免疫共沉淀实验表明NS3在哺乳动物细胞中能够与SRCAP结合。利用已知是由Notch激活的靶基因Hes - 1启动子进行的报告基因实验结果表明，NS3和SRCAP在Hep3B细胞中协同激活Hes - 1启动子。此外，我们在本报告中还表明，p400（一种已知与SRCAP非常相似的蛋白）也会成为NS3的作用靶点。通过免疫共沉淀实验，NS3对p400的1449 - 1808区域也表现出结合活性，并且通过使用短发夹RNA对SRCAP和p400的mRNA进行联合沉默，NS3对Notch介导的Hes - 1启动子转录的激活作用显著降低。这些结果表明，HCV NS3蛋白通过作用于SRCAP和p400参与了Notch信号通路的激活。