Transcriptional regulation of human CYP2C genes: functional comparison of CYP2C9 and CYP2C18 promoter regions.
The cytochrome P4502C subfamily comprises a group of constitutive microsomal hemoproteins which are expressed primarily in liver. In humans, this subfamily is responsible for metabolism of a variety of therapeutic drugs such as warfarin, mephenytoin, omeprazole, and antiinflammatory drugs. In the present study, we analyzed the promoter activity of the 5'-flanking region of two human CYP2C genes, CYP2C9 and CYP2C18. The ability of the 2.2-kb 5'-flanking region of the CYP2C9 gene to direct expression of a luciferase reporter gene in HepG2 cells was 25 times greater than that of the 1.3-kb 5'-flanking region of CYP2C18. Deletional analysis of CYP2C9 indicated that the minimal promoter was located between the translation start site and nucleotide -155, and an HPF-1 domain consensus sequence was identified in this region. Gel shift analysis demonstrated that nuclear proteins from HepG2 cells had a high binding affinity for a 20-bp oligonucleotide containing the HPF-1 site of CYP2C9. Antiserum to rat HNF-4 supershifted this DNA--protein complex, and an oligonucleotide derived from an HNF-4 motif present in the human apolipoprotein CIII promoter competed for the supershifted complex. Cotransfection with an HNF-4 expression plasmid increased transcriptional activity of the CYP2C9 minimal promoter (approximately 2-fold) in HepG2 cells and elevated activity more substantially in nonhepatic NIH3T3 cells (26-fold) and Cos 1 cells (9-fold).(ABSTRACT TRUNCATED AT 250 WORDS)
Ibeanu
G C
GC
National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Goldstein
J A
JA
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D001665:Binding Sites; D006528:Carcinoma, Hepatocellular; D065729:Cytochrome P-450 CYP2C9; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins; D017353:Gene Deletion; D005786:Gene Expression Regulation; D000882:Haplorhini; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D008113:Liver Neoplasms; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins; D011401:Promoter Regions, Genetic; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism / Q000494:pharmacology; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; C403128:MLX protein, human; D010750:Phosphoproteins; C501970:Tcfl4 protein, mouse; D014157:Transcription Factors; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C082185:CYP2C18 protein, human; D039181:Steroid 16-alpha-Hydroxylase