The human hepatitis B virus transactivator X gene product regulates Sp1 mediated transcription of an insulin-like growth factor II promoter 4

Abstract

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 identi®ed as Sp1 binding sites. Sp1 binding to IGF-II P4 promoter was identi®ed by gel mobility shift assay using puri®ed Sp1. By using a GAL4-Sp1 fusion protein it was demonstrated that HBV-X positively regulates the Sp1 mediated transcriptional activity of IGF-II in vivo. A protein-anity 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 speci®c 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.

Transcriptional Regulation of the TFIIH Transcription Repair Components XPB and XPD by the Hepatitis B Virus x Protein in Liver Cells and Transgenic Liver Tissue

“…9A). HBx has been shown to interact with Sp1 and to affect its DNA binding activity (9). Studies with Sp1 mutated promoter sequences in human liver cells were not possible as we observed that these sites were indispensable for XPB basal expression; this is in agreement with an earlier study (42).…”

“…Several transcription factor responsive elements are present in both XPB and XPD promoters including Sp1, Oct, TATA box-binding protein, and ETS-1. The Sp1 transcription factor has been shown to be a specific target for HBx resulting in impairment of its DNA binding properties (9). To address a possible role for Sp1, we used the Drosophila cell line SL2 which has an Sp1-deficient background.…”

Hepatitis B virus-X protein upregulates the expression of p21wafl/cipl and prolongs G1→S transition via a p53-independent pathway in human hepatoma cells

“…One possibility is the direct interactions of ets transcription factor with HBV-X resulting in the activation of p21 as in the case of Maguire et al (1991). Another possibility is the activations of ets binding to its binding site through the post-transcriptional protein modi®cations through signal transduction pathways (Lee et al, 1998b). It has been known that Ets1 and Ets2 are speci®c nuclear targets of Ras signaling events and that phosphorylation of a conserved threonine residue is a necessary molecular component of Ras-mediated activations of these transcription factors (Yang et al, 1996).…”

The H19 endodermal enhancer is required for Igf2 activation and tumor formation in experimental liver carcinogenesis

“…The arrows indicate the DNase I cleavage products of the 7.2 Kb HindIII and 11.4 BamHI full-length fragments. A diagram showing the relative positions of the H19 gene (box), the two H19 endodermal enhancer elements described by Yoo-Warren et al (1988) (circles), relevant restriction sites, probes, and identi®ed hypersensitive sites (HS1 ± 4, indicated by vertical arrows) is reported in (a) cell proliferation (Zhang et al, 1996(Zhang et al, , 1998Lee et al, 1998;Caricasole and Ward, 1993;Drummond et al, 1992;Kim et al, 1992;Dugimont et al, 1998).…”

“…Di erential activation of these promoters has been observed in human and experimental cancers (Ueno et al, 1988;Zarrilli et al, 1994). Although several transcription factors involved in neoplastic transformation have been reported to bind and regulate in vitro-transfected IGF2 promoter fragments, no information has been obtained so far on the mechanisms responsible for the activation of this gene during in vivo carcinogenesis (Zhang et al, 1996(Zhang et al, , 1998Lee et al, 1998;Caricasole and Ward, 1993;Drummond et al, 1992;Kim et al, 1992).…”