title: Identification of CCAAT enhancer binding protein alpha binding sites on the human alpha-fetoprotein gene.
abstract: Development- and tissue-specific alpha-fetoprotein (AFP) gene expression is controlled by various transcription factors including hepatocyte nuclear factors (HNFs), and a number of cis-acting elements. We recently identified multiple CCAAT/enhancer binding protein (C/EBP) binding sites in the enhancer of the human AFP gene. In this study, we have identified and functionally characterized seven C/EBPalpha-binding sites in the promoter and enhancer regions. An electrophoretic mobility shift assay (EMSA) and DNase I footprinting analysis identified two and five C/EBPalpha-binding sites located in the promoter and enhancer regions, respectively. Chromatin immunoprecipitation analyses showed that C/EBPalpha binds both enhancer and promoter regions of the AFP gene in human AFP-producing hepatoma and stomach cancer cells, but not in non-AFP-producing cells. Reporter transfection assays showed that transcription was stimulated by C/EBPalpha binding to each of the elements. These results indicate that C/EBPalpha regulates AFP gene expression through direct binding to multiple sites in the human AFP gene in cultured human cells.
authors.affiliation: Department of Biochemistry, Hokkaido Graduate School of Medicine, Sapporo, Japan.
authors.forename: Hong Mei
authors.initials: HM
authors.lastname: Li
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ikeda
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Nakabayashi
authors.forename: Shinzo
authors.initials: S
authors.lastname: Nishi
authors.forename: Masaharu
authors.initials: M
authors.lastname: Sakai
mesh_terms: D000818:Animals; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D047369:Chromatin Immunoprecipitation; D018983:DNA Footprinting; D024202:Electrophoretic Mobility Shift Assay; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D006801:Humans; D011401:Promoter Regions, Genetic; D000509:alpha-Fetoproteins / Q000235:genetics*
chemical_list: D022762:CCAAT-Enhancer-Binding Proteins; C515848:CEBPA protein, human; D000509:alpha-Fetoproteins