The human hepatitis B virus transactivator X gene product regulates Sp1 mediated transcription of an insulin-like growth factor II promoter 4.
Human hepatitis B virus (HBV) is one of the causative agents of hepatocellular carcinoma (HCC). The virus encodes a 17 kDa protein, X, which is known to be a causative agent in the formation of HCC. An insulin-like growth factor-II (IGF-II) is expressed during the formation of HCC. Among the four promoters of the IGF-II gene, promoters 2, 3 and 4 become activated during the formation of HCC. The high frequency of detection of hepatitis B virus X (HBV-X) antigen in liver cells from patients with chronic hepatitis, cirrhosis, and liver cancer suggested that the expressions of HBV-X and IGF-II are associated. Studies were carried out to test the relationship between the HBV-X gene product and the activation of IGF-II promoter 4. We demonstrated that the HBV-X protein increases the endogenous IGF-II expression from promoter 3 and 4 of IGF-II gene. Analysis of the fourth promoter of IGF-II gene showed that the HBV-X gene product positively regulates transcription. Two copies of a motif are responsible for conferring HBV-X regulation on the fourth promoter of IGF-II. These motifs have been identified as Sp1 binding sites. Sp1 binding to IGF-II P4 promoter was identified by gel mobility shift assay using purified Sp1. By using a GAL4-Sp1 fusion protein it was demonstrated that HBV-X positively regulates the Spl mediated transcriptional activity of IGF-II in vivo. A protein-affinity chromatography experiment showed that HBV-X protein does not bind directly to Sp1, but HBV-X does augment the DNA binding activity of the phosphorylated form of Sp1 in HepG2 cells. Sp1 was phosphorylated by HBV-X and its DNA-binding activity was up-regulated upon HBV-X transfections. Various HBV-X mutant expression vectors were used for the demonstration of specific interactions between Sp1 and HBV-X. These results indicate that HBV-X functions as a positive regulator of transcription, and that Sp1 is a direct target for the transcriptional regulation of IGF-II. Increasing the DNA binding ability of the phosphorylated form of Sp1 by HBV-X might be an important mechanism for regulating the IGF-II gene expression and possibly promoting cell division during hepatic carcinogenesis. Our experimental results suggest that expression of HBV-X might induce the expression of IGF-II and the IGF-II might play a role in hepatitis B virus pathogenesis during the formation of HCC.
Lee
Y I
YI
Molecular Cell Biology Research Division, Korea Research Institute of Bioscience and Biotechnology, Korea Institute of Science and Technology, Taejon.
Lee
S
S
Lee
Y
Y
Bong
Y S
YS
Hyun
S W
SW
Yoo
Y D
YD
Kim
S J
SJ
Kim
Y W
YW
Poo
H R
HR
D001665:Binding Sites; D006528:Carcinoma, Hepatocellular / Q000209:etiology / Q000235:genetics; D016384:Consensus Sequence; D004252:DNA Mutational Analysis; D005786:Gene Expression Regulation; D006515:Hepatitis B virus; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000096:biosynthesis* / Q000235:genetics; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D054334:Viral Regulatory and Accessory Proteins
D016329:Sp1 Transcription Factor; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D007335:Insulin-Like Growth Factor II