Establishment of a hepatitis C virus subgenomic replicon derived from human hepatocytes infected in vitro.
The hepatitis C virus (HCV) replicon system is a potent tool for understanding the mechanisms of HCV replication and proliferation, and for the development of treatments for patients with HCV. Recently, we established an HCV subgenomic replicon (50-1) using HCV genome RNA obtained from the cultured human T cell line MT-2C infected with HCV (isolate 1B-1) in vitro. In order to further obtain other HCV replicons without difficulty, we generated a replicon RNA library derived from human non-neoplastic hepatocytes infected with HCV (isolate 1B-2) in vitro. Upon transfection of the generated RNA library to "cured cells," from which the 50-1 subgenomic replicon was eliminated by prolonged treatment with interferon-alpha, we successfully established a new HCV subgenomic replicon, 1B-2R1. We characterized 1B-2R1 replicon in terms of efficiency of replication, HCV sequence, and sensitivity to interferons. The results revealed that the replication level of the 1B-2R1 replicon was comparable to that of the 50-1 replicon. We also found that the 1B-2R1 replicon possessed an HCV sequence distinct from those of other replicons established to date, and that the 1B-2R1 replicon was sensitive to interferon-alpha, interferon-beta, and interferon-gamma. Taken together, present results indicate that the replicon RNA library generated using an in vitro HCV infection system is useful for the establishment of an HCV subgenomic replicon.
Kato
Nobuyuki
N
Department of Molecular Biology, Okayama University Graduate School of Medicine and Dentistry, 2-5-1 Shikata-cho, 700-8558, Okayama, Japan. nkato@md.okayama-u.ac.jp
Sugiyama
Kazuo
K
Namba
Katsuyuki
K
Dansako
Hiromichi
H
Nakamura
Takashi
T
Takami
Marika
M
Naka
Kazuhito
K
Nozaki
Akito
A
Shimotohno
Kunitada
K
D002478:Cells, Cultured; D005814:Genes, Viral; D016174:Hepacivirus / Q000235:genetics* / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism / Q000821:virology*; D006801:Humans; D007372:Interferons / Q000378:metabolism; D008969:Molecular Sequence Data; D012367:RNA, Viral; D012093:Replicon / Q000235:genetics*; D014779:Virus Replication / Q000502:physiology*
D012367:RNA, Viral; D007372:Interferons