Hepatitis C virus (HCV) often causes persistent infection, and an association between hepatocellular carcinoma (HCC) and HCV infection has been reported. The only approved therapy for chronic HCV infection is IFN-a with or without ribavirin that offers limited benefit depending on the genotype of the infecting virus. Therefore, we need additional modalities that alone or in combination with existing therapy will generate improve responses. Understanding the molecular basis of HCV mediated disease progression is a major challenge. Our long-term goal is to understand the HCV mediated pathogenesis at the molecular level, which is likely to be mediated by both viral and host factors. Studies on HCV is challenging because of its poor replication in cell cultures and the lack of a convenient animal model. Despite these limitations, our work from the last grant period, together with that of other laboratories, suggested that HCV NS5A protein have many intriguing properties. These include inhibition of TNF-alpha mediated apoptosis from in vitro and in vivo studies, a regulatory role on important cellular genes and promotion of cell growth. Given a significant progress in this area, several questions related to the contribution of HCV NS5A in disease progression remains unanswered. This proposal is designed to extend our research findings in the following areas: (1) Does functional activity of p53 alter upon association with NS5A? (2) Analyze the effect of NS5A on interferon regulatory pathways and induction of inflammatory cytokines in transgenic mice, and (3) Investigate the silencing of HCV gene expression using RNA interference. Results from this study will identify the molecular events specifically regulated by NS5A for improving our understanding of host-virus interaction. This study may also lead to discovery of new therapeutic modalities against persistent HCV infection that affects over 170 million people worldwide.