An estimated 170 million people worldwide are infected with the hepatitis C virus (HCV), a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. Recently an infectious HCV cell culture (HCVcc) system that recapitulates the complete virus life cycle has been described that allows virus morphogenesis to be studied for the first time. The HCV p7 protein is a small, integral membrane protein that oligomerizes in vitro to form amantadine sensitive ion channels. p7 is essential for infectivity of HCV in chimpanzees and contains genotype specific sequences that are important for this function, suggesting that p7 interacts with other virus proteins. The role of p7 during the virus life cycle, however, is unknown. In preliminary studies I have demonstrated that p7 is required for infectious HCVcc production and that it contains essential genotype specific sequences. In addition, I have genetically identified putative interactions between p7 and viral nonstructural proteins including NS2. In this application we will identify the stage of virus morphogenesis at which p7 acts as well as the importance of p7 processing intermediates for these functions. I will use mutagenesis to precisely define the essential genotype specific sequences in p7 and isolate second-site revertants to define the viral proteins that interact with p7. I will then examine these putative interactions using confocal fluorescence microscopy to first localize p7 in infected cells and then to determine whether p7 co-localizes with other viral proteins. The interactions of p7 with other viral proteins will be further examined by coimmunoprecipitation and FRET-based techniques. The experiments described in this application will provide much needed detail into the role(s) of p7 in infectious virus production and the participation of other viral proteins in this process. HCV is a major human pathogen for which there is no vaccine and only limited treatment options. The identification of novel targets is necessary for the development of more effective and satisfactory antiviral strategies. These studies of the HCV p7 protein and its essential role in the enigmatic process of infectious virus production may provide such targets. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32DK081193-02
Application #
7501961
Study Section
Special Emphasis Panel (ZRG1-F13-P (20))
Program Officer
Podskalny, Judith M,
Project Start
2007-09-03
Project End
2010-09-02
Budget Start
2008-09-03
Budget End
2009-09-02
Support Year
2
Fiscal Year
2008
Total Cost
$52,898
Indirect Cost
Name
Rockefeller University
Department
Microbiology/Immun/Virology
Type
Other Domestic Higher Education
DUNS #
071037113
City
New York
State
NY
Country
United States
Zip Code
10065
Scull, Margaret A; Schneider, William M; Flatley, Brenna R et al. (2015) The N-terminal Helical Region of the Hepatitis C Virus p7 Ion Channel Protein Is Critical for Infectious Virus Production. PLoS Pathog 11:e1005297
Meuleman, Philip; Catanese, Maria Teresa; Verhoye, Lieven et al. (2012) A human monoclonal antibody targeting scavenger receptor class B type I precludes hepatitis C virus infection and viral spread in vitro and in vivo. Hepatology 55:364-72
Schoggins, John W; Wilson, Sam J; Panis, Maryline et al. (2011) A diverse range of gene products are effectors of the type I interferon antiviral response. Nature 472:481-5
Dorner, Marcus; Horwitz, Joshua A; Robbins, Justin B et al. (2011) A genetically humanized mouse model for hepatitis C virus infection. Nature 474:208-11
Jones, Christopher T; Catanese, Maria Teresa; Law, Lok Man J et al. (2010) Real-time imaging of hepatitis C virus infection using a fluorescent cell-based reporter system. Nat Biotechnol 28:167-71
Sheahan, Timothy; Jones, Christopher T; Ploss, Alexander (2010) Advances and challenges in studying hepatitis C virus in its native environment. Expert Rev Gastroenterol Hepatol 4:541-50
Law, Lok Man J; Albin, Owen R; Carroll, John-William N et al. (2010) Identification of a dominant negative inhibitor of human zinc finger antiviral protein reveals a functional endogenous pool and critical homotypic interactions. J Virol 84:4504-12
Kopp, Martina; Murray, Catherine L; Jones, Christopher T et al. (2010) Genetic analysis of the carboxy-terminal region of the hepatitis C virus core protein. J Virol 84:1666-73
Ploss, Alexander; Khetani, Salman R; Jones, Christopher T et al. (2010) Persistent hepatitis C virus infection in microscale primary human hepatocyte cultures. Proc Natl Acad Sci U S A 107:3141-5
Murray, Catherine L; Jones, Christopher T; Tassello, Jodie et al. (2007) Alanine scanning of the hepatitis C virus core protein reveals numerous residues essential for production of infectious virus. J Virol 81:10220-31