Characterization of the promoter region of the human peroxisomal multifunctional enzyme type 2 gene.
Peroxisomal multifunctional enzyme type 2 (perMFE-2) catalyzes conversion of (24E)-3alpha,7alpha, 12alpha-trihydroxy-5beta-cholest-24-enoyl-CoA to (24-keto)-3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoyl-CoA, which are physiological intermediates in cholic acid synthesis. In contrast to long chain fatty acid oxidizing enzymes clofibrate does not induce peroxisomal enzymes metabolizing bile acid intermediates. We proposed the existence of PPAR-independent regulation of cholesterol side chain oxidation in the process of bile acid synthesis. In the present study, we characterized the promoter region of the human perMFE-2 gene. The promoter contains the Sp1/AP2 binding site (-151/-142) within 197 base pairs upstream of the translation start site. Mutation of the Sp1/AP2 binding site decreases the promoter activity. Analysis by the luciferase assay revealed that the activity of the promoter region is strong in HepG2 and HeLa cell lines, although the activity in HepG2 cells was five- to sixfold higher than that in HeLa cells. Transient transfection assays have confirmed that AP2alpha and AP2gamma were able to transactivate the perMFE-2 promoter/luciferase chimeric gene. Cotransfections with Sp1 expression plasmid decreased the promoter activity. We suggest that perMFE-2 promoter activity is the result of both the abundance of AP2 and Sp1 family members and their relative ratios.
Novikov
D K
DK
Biocenter Oulu and Department of Biochemistry, University of Oulu, Oulu, FIN-90570, Finland. dnovikov@sun3.oulu.fi
Kamps
M E
ME
D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004746:Enoyl-CoA Hydratase / Q000235:genetics*; D020862:GC Rich Sequence / Q000235:genetics; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic / Q000502:physiology*; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D004268:DNA-Binding Proteins; D009097:Multienzyme Complexes; D016329:Sp1 Transcription Factor; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D015094:3-Hydroxyacyl CoA Dehydrogenases; D004746:Enoyl-CoA Hydratase