title: The CYP2B1 proximal promoter contains a functional C/EBP regulatory element.
abstract: Cytochromes P450 2B1 and 2B2 (CYP2B1 and CYP2B2) are well-known phenobarbital-inducible genes in rat liver. Potential transcriptional regulatory elements in the proximal promoter regions of rat CYP2B genes were analyzed by transfection in HepG2 hepatoma cells and by binding of nuclear proteins. Deletion of sequences from -1,400 to -110 had modest effects on promoter activity, but further deletion to -57 decreased the transcriptional activity by more than 90%, suggesting the presence of strong cis-acting elements in this region. Sequences similar to a basal transcription element (BTE) in CYP1A1 and a proposed phenobarbital responsive element (Barbie box) are present from -89 to -67. However, no protection was detected in these regions by DNase I footprinting assay. Instead, a region (FP1) from -64 to -45 was protected by liver nuclear extracts. Mutation of either the BTE or FP1 sequences of CYP2B1, or both, reduced transcriptional activity by 70-80% in HepG2 cells. FP1 was identified as a functional C/EBP site by co-transfection of C/EBP expression vectors and supershift assays with C/EBP antisera. Binding of liver nuclear proteins to sequences within the -110 to +1 region was not detectably altered by pretreatment of rats with phenobarbital.
authors.affiliation: Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 61801, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Park
authors.forename: B
authors.initials: B
authors.lastname: Kemper
mesh_terms: D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D022762:CCAAT-Enhancer-Binding Proteins; D006528:Carcinoma, Hepatocellular; D002457:Cell Extracts; D002467:Cell Nucleus; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D008099:Liver; D008297:Male; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D017208:Rats, Wistar; D017384:Sequence Deletion; D013250:Steroid Hydroxylases / Q000235:genetics*; D014158:Transcription, Genetic / Q000235:genetics; D015533:Transcriptional Activation / Q000235:genetics; D014407:Tumor Cells, Cultured
chemical_list: D022762:CCAAT-Enhancer-Binding Proteins; D002457:Cell Extracts; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C025619:steroid 16-beta-hydroxylase