title: Hepatitis B virus X protein regulates the mEZH2 promoter via the E2F1-binding site in AML12 cells.
abstract: Histone lysine methyltransferase EZH2 has been reported to be frequently overexpressed in hepatocellular carcinoma (HCC) tissues and associated with hepatocarcinogenesis. However, the exact mechanism of EZH2 up-regulation in HCC has not been determined. In this study, we used murine hepatocyte AML12 cells to investigate the role of hepatitis B virus X protein (HBx) in regulating the expression of mEZH2. Western blot analysis demonstrated that the expression level of mEZH2 protein in AML12 cells was up-regulated by HBx in a dose-dependent manner. To further investigate the mechanism of mEZH2 overexpression, the 2500 bp regulatory sequence upstream from the first exon of the mEZH2 gene was amplified from AML12 genomic DNA and constructed into a luciferase reporter plasmid. The luciferase activity of the mEZH2 promoter significantly increased in AML12 cells co-transfected with HBx plasmid, and deleting the -486/-214 promoter region decreased HBx-induced mEZH2 promoter activation by nearly 50%. The -486/-214 region was then analyzed in the TRANSFAC 6.0 database and a typical E2F1-binding site was found. Mutation of this E2F1-binding site or knockdown of E2F1 expression by RNAi led to a dramatic decrease in HBx-induced activation of the mEZH2 promoter and mEZH2 overexpression in AML12 cells. These results provide evidence that HBx up-regulates mEZH2 expression by transactivating the mEZH2 promoter through E2F1 transcription factor, thereby providing new epigenetic evidence for the carcinogenic effect of HBx.
authors: Beijing Institute of Biotechnology, Beijing 100071, P. R. China.
authors: Xiao-Yan
authors: XY
authors: Shi
authors: Ying-Ying
authors: YY
authors: Zhang
authors: Xiao-Wei
authors: XW
authors: Zhou
authors: Jian-Sheng
authors: JS
authors: Lu
authors: Ze-Kun
authors: ZK
authors: Guo
authors: Pei-Tang
authors: PT
authors: Huang
mesh_terms: D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D050687:E2F1 Transcription Factor / Q000235:genetics*; D000071221:Enhancer of Zeste Homolog 2 Protein; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D011495:Histone-Lysine N-Methyltransferase / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D010957:Plasmids; D063151:Polycomb Repressive Complex 2; D011401:Promoter Regions, Genetic / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D015854:Up-Regulation; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D050687:E2F1 Transcription Factor; D034741:RNA, Small Interfering; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D000071221:Enhancer of Zeste Homolog 2 Protein; C498410:Ezh2 protein, mouse; D011495:Histone-Lysine N-Methyltransferase; D063151:Polycomb Repressive Complex 2