A human hepatoma cell line expressing hepatitis c virus nonstructural proteins tightly regulated by tetracycline.
Nonstructural (NS) proteins of hepatitis C virus (HCV) play major roles in viral replication and the pathogenesis of liver diseases. Current studies on antiviral strategies targeting these proteins have been hampered by the lack of efficient cell culture systems. Combining tetracycline-regulated gene expressing system and enhanced green fluorescent protein (EGFP), we generated a human hepatoma cell line inducibly expressing the HCV NS proteins. This cell line exhibited high induction of a full NS transcript ( approximately 7 kb). In the absence of tetracycline, NS proteins 3, 4A, and 5A of mature sizes were detected by immunoblot analysis and the induction of NS proteins 3 to 5B are confirmed by immunofluorescent staining. Using DNA microarray analysis, we characterized the changes in mRNA expression profile of 6416 genes and identified several genes, whose mRNAs are specifically upregulated by the induction of NS proteins. This cell line provides a unique in vitro hepatoma cell system for the investigation of structural and functional properties of HCV NS proteins.
Huang
Y
Y
Department of Molecular Biology and Medicine, RCAST, University of Tokyo, 4-6-1 Komaba, Tokyo, Meguro-ku, 153-8904, Japan.
Uchiyama
Y
Y
Fujimura
T
T
Kanamori
H
H
Doi
T
T
Takamizawa
A
A
Hamakubo
T
T
Kodama
T
T
D006528:Carcinoma, Hepatocellular / Q000473:pathology / Q000821:virology*; D005455:Fluorescent Antibody Technique; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D049452:Green Fluorescent Proteins; D016174:Hepacivirus / Q000378:metabolism*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology / Q000821:virology*; D008164:Luminescent Proteins / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000235:genetics; D013754:Tetracyclines / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D017361:Viral Nonstructural Proteins / Q000235:genetics*
D008164:Luminescent Proteins; D012333:RNA, Messenger; D013754:Tetracyclines; D017361:Viral Nonstructural Proteins; D049452:Green Fluorescent Proteins