Intragenotypic JFH1 based recombinant hepatitis C virus produces high levels of infectious particles but causes increased cell death

• Type of Material: Article
• Authors: Guaniri Mateu, Emory University; Ruben O. Donis, Centers for Disease Control and Prevention; Takaji Wakita, National Institute of Infectious Diseases, Japan; Jens Bukh, Copenhagen University Hospital; Arash Grakoui, Emory University
• Language: English
• Date: 2008-07-05
• Publisher: Elsevier
• Publication Version: Author Accepted Manuscript (After Peer Review)
• Copyright Statement: © 2008 Elsevier Inc. All rights reserved.
• License: Creative Commons Attribution-NonCommerical-NoDerivs 3.0 Unported
• Final Published Version (URL): http://dx.doi.org/10.1016/j.virol.2008.03.027
• Title of Journal or Parent Work: Virology
• ISSN: 0042-6822
• Volume: 376
• Issue: 2
• Start Page: 397
• End Page: 407
• Grant/Funding Information: JB was in part supported by grants from the Lundbeck Foundation, the Novo Nordisk Foundation, the Danish Medical Research Council and the A.P. Møller and the Chastine Mc-Kinney Møller Foundation.; The authors would like to acknowledge the support from the Cancer Research Institute Investigator Award, Woodruff Health Sciences Fund, EVC/CFAR Flow Cytometry Core P30 AI050409, Yerkes Research Center Base Grant RR-00165, and the public Health Service AI052448 and AI070101(AG).; TW was partially supported by a grant-in-aid for Scientific Research from the Japan Society for the Promotion of Science, from the Ministry of Health, Labor and Welfare of Japan and from the Ministry of Education, Culture, Sports, Science and Technology, and by the Research on Health Sciences Focusing on Drug Innovation from the Japan Health Sciences Foundation.
• Keywords: cell death; JFH; Hepatitis C virus; intragenotypic recombinant
• Research Categories: Biology, Virology

Abstract

The full-length hepatitis C virus (HCV) JFH1 genome (genotype 2a) produces moderate titers of infectious particles in cell culture but the optimal determinants required for virion production are unclear. It has been shown that intragenotypic recombinants encoding core to NS2 from J6CF in the context of JFH1 are more robust in the release of viral particles. To understand the contributions of structural and nonstructural genes to HCV replication potential and infectivity, we have characterized intragenotypic recombinant genotype 2a viruses with different portions of the J6 isolate engineered into the JFH1 infectious clone. All genomes produced high levels of intracellular HCV RNA and NS3 protein in Huh-7.5 transfected cells. However, JFH1 genomes containing J6 sequences from C to E2 (CE2) or C to p7 (Cp7) secreted up to 100-fold more infectious HCV particles than the parental JFH1 clone. Subsequent infection of naïve Huh-7.5 cells with each of the J6/JFH1 recombinants at a multiplicity of infection of 0.0003 resulted in high viral titers only for CE2 and Cp7 viruses. Comparison of virion production by the Cp7 J6/JFH1 recombinant to previously described J6/JFH1 recombinants showed flexibility of the chimeric junction. Moreover, NTRNS2 a chimeric virus equivalent to the previously reported FL-J6/JFH chimera, showed a 10- fold enhancement of virus titers compared to CNS2. NTRNS2 differs from CNS2 by three nucleotide differences residing in the 5’NTR and core coding sequence and all three nucleotide changes were necessary for increased virion production. Importantly, cells producing Cp7 virus showed increased apoptosis compared with JFH1, an effect correlating with virion production. These studies begin to unravel requirements for robust virus replication and the relationship between increased virion production and host cell viability.