Hepatitis C and Tumor Suppressors in Hepatocellular

本项目旨在确定临床相关性和在肝癌发生中的潜在作用， 宿主细胞和丙型肝炎病毒（HCV）蛋白之间的相互作用已经在以前的细胞中鉴定， 基于文化的实验视网膜母细胞瘤易感蛋白（pRb）和细胞死亡盒RNA 解旋酶DDXS在调节细胞周期中起关键作用。我们以前的工作表明，HCV RNA 聚合酶NS 5 B与pRb形成复合物，靶向它进行泛素化和蛋白酶体依赖性的蛋白酶体修饰。 在一些实施方案中，所述方法包括降解、激活E2 F应答性启动子和刺激细胞增殖。同样，HCV cofe 蛋白质表达改变DDX 3的细胞丰度和定位。我们的假设是， 与宿主蛋白（p53和DDX 5）的其他相互作用促进肝细胞增殖并损伤DNA 损伤反应，从而促进HCV致癌。在目标1中，我们将开发和验证 荧光量子点探针能够灵敏地检测在病毒感染的 细胞我们还将确定HCV蛋白表达是否与肿瘤抑制基因的改变有关， 在转基因小鼠中表达。目的2将利用这些新的探针在激光扫描共焦和多 慢性丙型肝炎患者的肝组织的光子显微镜研究，并询问是否 pRb、p53、DDXS或DDXS的丰度和细胞定位在感染的肝细胞中改变。在目标3中， 我们将在单细胞基础上确定HCV感染是否与 增殖标志物Ki-67和PCNA。我们还将确定是否显示细胞的比例 在标准护理聚乙二醇-IFN/利巴韦林治疗后增殖标记物减少。这些结果将 与寡核苷酸芯片和定量RT-PCR检测相关，以确定肝脏是否 具有高比例HCV感染细胞的标本显示转录模式， 肝细胞增殖和/或E2 F转录因子激活。与公众健康的关系： 肝癌是美国增长最快的癌症类型之一，这反映了一个 慢性丙型肝炎患者肝癌的患病率和风险增加。该项目旨在更好地 对HCV蛋白与细胞增殖的重要调节因子（pRb，p53， DDXS和DDXS）和（这种相互作用在HCV感染引起的肝癌中的作用。 演出现场

This project seeks to determine the clinical relevance and potential role in hepatic carcinogenesis of interactions between host cell and hepatitis C virus (HCV) proteins that have been identified in previous cell culture-based experiments. The retinoblastoma susceptibility protein (pRb) and the cellular DEAD-box RNA helicase DDXS play critical roles in regulating the cell cycle. Our previous work indicates that the HCV RNA polymerase, NS5B, fc^nns a complex with pRb, targeting it for ubiquitination and proteasome-dependent degradation, activating E2F-responsive promoters, and stimulating cell proliferation. Similarly, HCV cofe protein expression alters the cellular abundance and localization of DDX3. Our hypothesis is that these and other interactions with host proteins (p53 and DDX5) promote proliferation of hepatocytes and impair DNA damage responses, tljiereby contributing to HCV carcinogenesis. In Aim 1, we will develop and validate fluorescent quantum dot probes capable of sensitive detection of HCV proteins expressed in virus-infected cells. We will also determine whether HCV protein expression conrelates with altered tumor suppressor expression in transgenic mice. Aim 2 will utilize these novel probes in laser scanning confocal and multi photon microscopy studies of liver tissue from patients with chronic hepatitis C, and ask whether the abundance and cellular localization of pRb, p53, DDXS, or DDXS is altered in infected hepatocytes. In Aim 3, we will determine whether HCV infection is associated on a single-cell basis with increased expression of the proliferation markers Ki-67 and PCNA. We will also determine whether the proportion of cells displaying proliferation markers is reduced after standard-of-care peg-IFN/ribavirin therapy. These results will be correlated with oligonucleotide microarray and quantitative RT-PCR assays to determine whether liver specimens with a high proportion of HCV-infected cells display transcriptional patterns indicative of hepatocellular proliferation and/or E2F transcription factor activation. RELEVANCE TO PUBLIC HEATH:,' Liver cancer is one qf the most rapidly increasing types of cancers in the United States, reflecting an increased prevalence ^nd risk of liver cancer in persons with chronic hepatitis C. This project seeks a better understanding of the interaction of HCV proteins with important regulators of cell proliferation (pRb, p53, DDXS, and DDXS) anc( the role of such interactions in liver cancer caused by HCV infection. PERFORMANCE SITE