The transcription factor GATA-4 regulates cytochrome P4502C19 gene expression.
OBJECTIVE
Cytochrome P4502C19 (CYP2C19) is an important enzyme involved in the metabolism of antiulcer drugs and antidepressants. However, despite the well documented drug-dependent variability of CYP2C19 expression, the mechanisms underlying the regulation of the enzyme remain unknown. In this study we investigated whether the transcription factor family GATA is involved in the regulation of CYP2C19 gene expression.


METHODS
We identified a novel putative GATA binding site at position -165/-156 within the CYP2C19 gene promoter. 5'-Deletion fragments of the CYP2C19 promoter containing wild type or mutant variants of this GATA binding site were co-transfected with expression vectors encoding the transcription factors GATA-4 or GATA-2 and analyzed using dual luciferase gene reporter assays in HepG2 and Huh-7 hepatoma cells. Electrophoretic Mobility Shift Assay (EMSA) and Chromatin Immunoprecipitations (ChIP) were performed to proof a sequence-specific interaction of GATA proteins with the putative GATA binding site.


RESULTS
The wild type fragments of CYP2C19 promoter were highly upregulated by GATA-4 and GATA-2 in luciferase gene reporter assay, whereas mutations introduced into the GATA binding sites caused a significant activity loss. Similar attenuation was observed upon co-transfection of GATA-4 with a known co-regulator of GATA activity, FOG-2. EMSA analysis revealed a sequence-specific binding of GATA-4 and GATA-6 to the wild type GATA binding site. In addition, the association of GATA-4 with the CYP2C19 promoter was confirmed by ChIP analysis.


CONCLUSIONS
These data indicate that GATA-4 plays an important role in the transcriptional regulation of CYP2C19 expression.
Mwinyi
Jessica
J
Karolinska Institutet, Department of Physiology and Pharmacology, Section of Pharmacogenetics, 17177 Stockholm, Sweden. Jessica.Mwinyi@ki.se
Hofmann
Yvonne
Y
Pedersen
Rasmus Steen
RS
Nekvindová
Jana
J
Cavaco
Isa
I
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; D065731:Cytochrome P-450 CYP2C19; D024202:Electrophoretic Mobility Shift Assay; D050989:GATA2 Transcription Factor / Q000378:metabolism*; D050991:GATA4 Transcription Factor / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic; D056945:Hep G2 Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics; D051379:Mice; D009154:Mutation; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic
D050989:GATA2 Transcription Factor; D050991:GATA4 Transcription Factor; D008156:Luciferases; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19