Journal
VIROLOGY
Volume 376, Issue 2, Pages 397-407Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.virol.2008.03.027
Keywords
hepatitis C virus; intragenotypic recombinant; JFH; cell death
Categories
Funding
- NCRR NIH HHS [RR-00165, P51 RR000165, P51 RR000165-48] Funding Source: Medline
- NIAID NIH HHS [AI052448, R21 AI052448, AI070101, R21 AI052448-01A1, P30 AI050409-07, P30 AI050409, R01 AI070101, R01 AI070101-01, R56 AI070101] Funding Source: Medline
- NIDDK NIH HHS [R01 DK083356] Funding Source: Medline
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The full-length hepatitis C virus (HCV)JFH1 genome (genotype 2a) produces moderate titers of infectious particles Received 6 February 2008 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 naive 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. (C) 2008 Elsevier Inc. All rights reserved.
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