Characterization of the basal promoter element of the human type 5 17beta-hydroxysteroid dehydrogenase gene.
Testosterone biosynthesis from androstenedione is carried out by androgenic 17beta-hydroxysteroid dehydrogenase (17beta-HSD) activity. Of the androgenic 17beta-HSD isoenzymes, only type 5 (17beta-HSD5) is expressed ubiquitously, including the human adrenal gland and ovary. To characterize this gene promoter, luciferase constructs of the human 5'-flanking region were transiently transfected into the H295R human adrenal carcinoma cell line. A series of fragment deletion constructs and electrophoretic mobility shift assays suggested that a sequence of CCTCCTCCT at -65 to -58 bp is the core sequence and demonstrated Sp1/Sp3 binding to this CCT repeat. Forskolin stimulated the promoter activity of the HSD17B5 gene through this Sp1/Sp3 binding site. Mutation of this site resulted in a significant reduction of HSD17B5 promoter basal and forskolin-induced activity. Mithramycin A, which inhibits the binding of Sp1 and Sp3 to DNA, also remarkably decreased HSD17B5 mRNA expression in the H295R cell line. These results indicate that members of the Sp family of transcription factors play an important role in regulating constitutive and stimulated expression of the HSD17B5 gene in H295R cells.
Qin
Kenan
K
Departments of Pediatrics and Medicine, The University of Chicago, Chicago, IL 60637, USA.
Rosenfield
Robert L
RL
D015067:17-Hydroxysteroid Dehydrogenases / Q000235:genetics*; D015096:3-Hydroxysteroid Dehydrogenases; D024506:5' Flanking Region / Q000235:genetics; D000074425:Aldo-Keto Reductase Family 1 Member C3; D001483:Base Sequence; D015153:Blotting, Western; D045744:Cell Line, Tumor; D005576:Colforsin / Q000494:pharmacology; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D006911:Hydroxyprostaglandin Dehydrogenases; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D008926:Plicamycin / Q000031:analogs & derivatives* / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D018911:Trinucleotide Repeats / Q000235:genetics
D017931:DNA Primers; D004268:DNA-Binding Proteins; D009841:Oligonucleotides; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; D005576:Colforsin; C066851:mithramycin A; D015067:17-Hydroxysteroid Dehydrogenases; D015096:3-Hydroxysteroid Dehydrogenases; D006911:Hydroxyprostaglandin Dehydrogenases; C487341:AKR1C3 protein, human; D000074425:Aldo-Keto Reductase Family 1 Member C3; D008156:Luciferases; D008926:Plicamycin