New insights into the regulation of CYP2C9 gene expression: the role of the transcription factor GATA-4.
CYP2C9 is an important drug-metabolizing enzyme that metabolizes, e.g., warfarin, antidiabetics, and antiphlogistics. However, the endogenous regulation of this enzyme is largely unknown. In this study, we examined the role of GATA transcription factors in the gene expression of CYP2C9. We investigated four putative GATA binding sites within the first 200 base pairs of CYP2C9 promoter at the positions I: -173/-170, II: -167/-164, III: -118/-115, and IV: -106/-103. Luciferase activity driven by a wild-type CYP2C9 promoter construct was strongly up-regulated in Huh-7 cells upon cotransfection with expression plasmids for GATA-2 and GATA-4, whereas mutations introduced into GATA binding site III or I and II reduced this induction to a significant extent. Electrophoretic mobility shift assays revealed specific binding of GATA-4 and GATA-6 to the oligonucleotides containing GATA binding sites I and II. Furthermore, the association of GATA-4 with CYP2C9 promoter was confirmed by chromatin immunoprecipitation assays in HepG2 cells. Taken together, these data strongly suggest an involvement of liver-specific transcription factor GATA-4 in the transcriptional regulation of CYP2C9.
Mwinyi
Jessica
J
Karolinska Institutet, Department of Physiology and Pharmacology, Section of Pharmacogenetics, Stockholm, Sweden. Jessica.Mwinyi@ki.se
Nekvindová
Jana
J
Cavaco
Isa
I
Hofmann
Yvonne
Y
Pedersen
Rasmus Steen
RS
Landman
Ellie
E
Mkrtchian
Souren
S
Ingelman-Sundberg
Magnus
M
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D065729:Cytochrome P-450 CYP2C9; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D050989:GATA2 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050991:GATA4 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D050993:GATA6 Transcription Factor / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D004268:DNA-Binding Proteins; D050989:GATA2 Transcription Factor; C494711:GATA2 protein, human; D050991:GATA4 Transcription Factor; C484170:GATA4 protein, human; D050993:GATA6 Transcription Factor; C484171:Gata4 protein, mouse; D014157:Transcription Factors; C117427:ZFPM2 protein, human; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases