Transcriptional regulation of the human cholesterol 7 alpha-hydroxylase gene (CYP7A) in HepG2 cells.
A stable HepG2 cell line harboring a human cholesterol 7 alpha-hydroxylase (CYP7A) minigene/luciferase reporter gene construct was selected for studying transcriptional regulation of CYP7A gene promoter. Insulin and phorbol 12-myristate-13-acetate (PMA) strongly repressed the promoter activity as measured with luciferase activity expressed in the cells. The promoter activity of the 5' progressive deletion/luciferase reporter gene constructs was studied in a transient transfection assay in HepG2 cells. PMA represses the promoter activity and the response elements were localized in the -184/-151 and -134/-81 regions. Insulin also represses the promoter activity and response element was mapped in the -298/-81 region. Surprisingly, glucocorticoid receptor (GR) strongly inhibited promoter activity in the presence of dexamethasone, and response elements were localized in the -298/-151 and the -150/+24 regions. Thyroid hormone receptor also repressed promoter activity and response elements were localized in the -150/+24 and upstream regions. Cotransfection of CYP7A chimeric constructs with an expression vector carrying liver-enriched transcription factor HNF3 alpha stimulated the reporter gene activity, but cotransfection with GR plasmid interfered with the HNF3 alpha-stimulated activity possibly through competition for binding to overlapping GR/HNF3 binding sites. Thus, human cholesterol 7 alpha-hydroxylase gene promoter is strongly repressed by insulin, PMA, and steroid/thyroid hormones and results in the low level of cholesterol 7 alpha-hydroxylase expression in the human liver.
Department of Biochemistry and Molecular Pathology, Northeastern Ohio Universities College of Medicine, Rootstown 44272, USA.
D P
DP
Wang
D
D
Stroup
M
M
Marrapodi
M
M
Crestani
G
G
Galli
J Y
JY
Chiang
D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002790:Cholesterol 7-alpha-Hydroxylase / Q000096:biosynthesis / Q000235:genetics*; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004794:Enzyme Repression; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017930:Genes, Reporter; D005938:Glucocorticoids / Q000494:pharmacology; D051541:Hepatocyte Nuclear Factor 3-alpha; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013963:Thyroid Hormones / Q000494:pharmacology; D014157:Transcription Factors; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; C496499:FOXA1 protein, human; D005938:Glucocorticoids; D051541:Hepatocyte Nuclear Factor 3-alpha; D007328:Insulin; D009687:Nuclear Proteins; D010703:Phorbol Esters; D011965:Receptors, Glucocorticoid; D011993:Recombinant Fusion Proteins; D013963:Thyroid Hormones; D014157:Transcription Factors; D002790:Cholesterol 7-alpha-Hydroxylase