Role of ISGF3 in modulating the anti-hepatitis B virus activity of interferon-alpha in vitro.
OBJECTIVE
Although interferon-alpha (IFN-alpha) is an effective treatment for hepatitis B virus (HBV) infection, its precise mechanism of action has not been identified. In this study, we investigated the role of signal transduction pathways in the activation of anti-HBV responses mediated by IFN-alpha.


METHODS
Using an oligo microarray, we found that four genes in the IFN-alpha signal pathway were markedly upregulated by IFN-alpha in human hepatoma cells regardless of whether they had been transfected with a plasmid containing the HBV genome: signal transducers and activators of transcription 1 (STAT1), interferon regulatory factor-9 (IRF-9, also called ISGF3gamma or P48), IFN-alpha-inducible protein 15 (IFI-15) and IFN-alpha-inducible protein 6-16 (IFI-6-16). We also investigated the role of IFN-stimulated gene factor3 (ISGF3) complex in IFN-alpha-mediated anti-HBV responses in human hepatoma cells by measuring the mRNA of the three genes within ISGF3 (STAT1, STAT2 and IRF-9) using semiquantitative reverse-transcription PCR (RT-PCR), and expression of the three proteins by western blot, and the mRNA and protein of dsRNA-dependent protein kinase (PKR).


RESULTS
STAT1, STAT2, IRF-9 and PKR mRNA as well as protein levels were upregulated by IFN-alpha treatment. When cells were pretreated with genistein, STAT1, STAT2 and IRF-9 mRNA levels remained unchanged after IFN-alpha stimulation, but PKR mRNA levels decreased, and the expression of the STAT1, P-STAT2, IRF-9 and PKR proteins decreased. Levels of HBV DNA decreased in the supernatants of cells treated with IFN-alpha, while ISGF3 levels increased. The quantity of HBV DNA remained unchanged by pretreating with genistein.


CONCLUSIONS
These observations suggested that the Janus tyrosine kinase-STAT (JAK-STAT) pathway may play a major role in mediating the effects of IFN-alpha against HBV, and that ISGF3 might be a key factor.
Zhang
Quan
Q
Hepatology Institute, Peking University People's Hospital, Beijing, China.
Wang
Yan
Y
Wei
Lai
L
Jiang
Dong
D
Wang
Jiang Hua
JH
Rao
Hui Ying
HY
Zhu
Ling
L
Chen
Hongsong
H
Fei
Ran
R
Cong
Xu
X
D000998:Antiviral Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D016207:Cytokines / Q000378:metabolism; D004279:DNA, Viral / Q000096:biosynthesis; D020869:Gene Expression Profiling / Q000379:methods; D019833:Genistein / Q000494:pharmacology; D006514:Hepatitis B Surface Antigens / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000378:metabolism; D006515:Hepatitis B virus / Q000187:drug effects* / Q000235:genetics / Q000276:immunology; D022781:Hepatocytes / Q000187:drug effects* / Q000276:immunology / Q000821:virology; D006801:Humans; D000077190:Interferon alpha-2; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit / Q000235:genetics / Q000378:metabolism*; D016898:Interferon-alpha / Q000494:pharmacology*; D053612:Janus Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins; D050794:STAT1 Transcription Factor / Q000378:metabolism; D050795:STAT2 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics; D013997:Time Factors; D014162:Transfection; D014452:Ubiquitins / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects; D019892:eIF-2 Kinase / Q000378:metabolism
D000998:Antiviral Agents; D016207:Cytokines; D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; C468762:IFI6 protein, human; C494227:IRF9 protein, human; D000077190:Interferon alpha-2; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha; D024101:Mitochondrial Proteins; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050795:STAT2 Transcription Factor; C494084:STAT2 protein, human; D014452:Ubiquitins; C074034:ISG15 protein, human; D019833:Genistein; D053612:Janus Kinases; D019892:eIF-2 Kinase