DESCRIPTION (provided by applicant): The broad, long-term goal of our program is to advance knowledge of virus-host cell interactions by characterizing cellular cofactors essential for hepatitis C virus (HCV) infection. HCV infects 170 million people worldwide and is major cause of hepatocellular carcinoma. Understanding how this virus interacts with the host cell (e.g., what makes a cell susceptible to HCV infection) is of critical importance to the development of diagnostics, antiviral drugs, and a prophylactic vaccine. Our previous studies of pluripotent stem cells and directed hepatic differentiation demonstrated a correlation between hepatic specification during the differentiation process and permissiveness to HCV infection. Characterization of the cellular transition from nonpermissive to permissive revealed candidate genes that are involved in regulating viral susceptibility. One of these cellular factors interacts with HCV glycoprotein E1 and plays an essential role in the HCV life cycle. The experiments proposed here will determine the molecular determinants of HCV permissiveness during hepatic development and identify new cellular cofactors and potential drug targets for HCV therapy. The following experiments will be performed: (1) Suppression of putative proviral factors and overexpression of putative antiviral factors in stem cells. These will be followed by hepatic differentiation and HCV infection of the differentiated human hepatocyte-like cells (DHHs). (2) Mechanistic studies of a novel proviral factor whose role has been validated by our preliminary studies. Specific steps of the HCV life cycle will be examined, and the interaction between viral and host protein will be dissected. (3) In vivo experiments designed to assess the potential of genetically modified DHHs to repopulate mouse liver and confer HCV resistance to the chimeric liver. The results of the proposed studies will not only provide significant insights into the properties of the host cells that govern HCV susceptibility but also reveal new therapeutic targets for antiviral intervention.

PUBLIC HEALTH RELEVANCE: Hepatitis C virus (HCV) infection and its associated diseases continue to impose a huge healthcare burden worldwide. No prophylactic vaccine exists to prevent HCV infection, and a better understanding of what makes a cell susceptible or resistant to infection will aid drug-development efforts. The proposed research will harness the power of stem- cell technology and a small-animal model to determine the molecular determinants of HCV permissiveness and reveal additional targets for therapeutic intervention.