TY - JOUR
T1 - Development of plaque assays for hepatitis C virus-JFH1 strain and isolation of mutants with enhanced cytopathogenicity and replication capacity
AU - Sekine-Osajima, Yuko
AU - Sakamoto, Naoya
AU - Mishima, Kako
AU - Nakagawa, Mina
AU - Itsui, Yasuhiro
AU - Tasaka, Megumi
AU - Nishimura-Sakurai, Yuki
AU - Chen, Cheng Hsin
AU - Kanai, Takanori
AU - Tsuchiya, Kiichiro
AU - Wakita, Takaji
AU - Enomoto, Nobuyuki
AU - Watanabe, Mamoru
N1 - Funding Information:
We are indebted to Dr. Francis V. Chisari for providing the Huh-7.5.1 cell line. This study was supported by grants from the Japan Society for the Promotion of Science, Miyakawa Memorial Research Foundation, and Viral Hepatitis Research Foundation of Japan.
PY - 2008/2/5
Y1 - 2008/2/5
N2 - HCV culture in vitro results in massive cell death, which suggests the presence of HCV-induced cytopathic effects. Therefore, we investigated its mechanisms and viral nucleotide sequences involved in this effect using HCV-JFH1 cell culture and a newly developed HCV plaque assay technique. The plaque assay developed cytopathic plaques, depending on the titer of the inoculum. In the virus-infected cells, the ER stress markers, GRP78 and phosphorylated eIF2-alpha, were overexpressed. Cells in the plaques were strongly positive for an apoptosis marker, annexin V. Isolated virus subclones from individual plaque showed greater replication efficiency and cytopathogenicity than the parental virus. The plaque-purified virus had 9 amino acid substitutions, of which 5 were clustered in the C terminal of the NS5B region. Taken together, the cytopathic effect of HCV infection involves ER-stress-induced apoptotic cell death. Certain HCV genomic structures may determine the viral replication capacity and cytopathogenicity.
AB - HCV culture in vitro results in massive cell death, which suggests the presence of HCV-induced cytopathic effects. Therefore, we investigated its mechanisms and viral nucleotide sequences involved in this effect using HCV-JFH1 cell culture and a newly developed HCV plaque assay technique. The plaque assay developed cytopathic plaques, depending on the titer of the inoculum. In the virus-infected cells, the ER stress markers, GRP78 and phosphorylated eIF2-alpha, were overexpressed. Cells in the plaques were strongly positive for an apoptosis marker, annexin V. Isolated virus subclones from individual plaque showed greater replication efficiency and cytopathogenicity than the parental virus. The plaque-purified virus had 9 amino acid substitutions, of which 5 were clustered in the C terminal of the NS5B region. Taken together, the cytopathic effect of HCV infection involves ER-stress-induced apoptotic cell death. Certain HCV genomic structures may determine the viral replication capacity and cytopathogenicity.
KW - Apoptosis
KW - ER stress
KW - HCV cell culture
KW - HCV-JFH1
KW - NS5B RNA-dependent RNA polymerase
KW - Plaque assay
KW - Unfolded protein responses
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U2 - 10.1016/j.virol.2007.09.019
DO - 10.1016/j.virol.2007.09.019
M3 - Article
C2 - 17949770
AN - SCOPUS:37849040237
SN - 0042-6822
VL - 371
SP - 71
EP - 85
JO - Virology
JF - Virology
IS - 1
ER -
