Repression of cAMP-induced expression of the mouse P450 17 alpha-hydroxylase/C17-20 lyase gene (Cyp17) by androgens.
In primary cultures of mouse Leydig cells, testosterone represses the cAMP-induced de novo synthesis of P450 17 alpha-hydroxylase/C17-20 lyase (P450c17) protein and the accumulation of P450c17 mRNA, via an androgen receptor (AR)-mediated mechanism. To examine the mechanism by which androgens repress the cAMP-induced expression of the mouse Cyp17 gene, constructs containing 5'-flanking sequences of the mouse Cyp17 linked to the chloramphenicol acetyltransferase (CAT) reporter gene were cotransfected into MA-10 tumor Leydig cells with a mouse AR expression plasmid. In the presence of dihydrotestosterone, the cAMP-induced expression of a reporter construct containing -1021 bp of Cyp17 promoter sequences was repressed. In contrast, no repression by dihydrotestosterone was observed when the -1021 bp Cyp17-CAT construct was cotransfected with a human AR expression plasmid missing the second zinc finger of the DNA-binding domain, indicating that DNA binding is involved in AR-mediated repression of Cyp17 expression. Analysis of deletions -346 bp of 5'-flanking region of the mouse Cyp17 promoter are sufficient to confer androgen repression of the cAMP-induced expression of Cyp17. Deoxyribonuclease I footprinting analysis indicated that the AR interacts with sequences between -330. and -278 bp of the Cyp17 promoter. This region overlaps with the previously identified cAMP-responsive region located between -346 and -245 bp of the Cyp17 promoter. These results suggest that AR-mediated repression involves binding of the AR to sequences in the cAMP-responsive region of the Cyp17 promoter, possibly interfering with the binding of the protein(s) that mediate cAMP induction of Cyp17.
Burgos-Trinidad
M
M
Department of Obstetrics and Gynecology, University of Michigan, Ann Arbor 48109, USA.
Youngblood
G L
GL
Maroto
M R
MR
Scheller
A
A
Robins
D M
DM
Payne
A H
AH
D000818:Animals; D000242:Cyclic AMP / Q000502:physiology; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000235:genetics; D018983:DNA Footprinting; D013196:Dihydrotestosterone / Q000494:pharmacology*; D004790:Enzyme Induction / Q000187:drug effects; D017930:Genes, Reporter; D006801:Humans; D007984:Leydig Cell Tumor / Q000473:pathology; D007985:Leydig Cells / Q000378:metabolism; D008297:Male; D051379:Mice; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D011944:Receptors, Androgen / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000502:physiology*; D015290:Second Messenger Systems; D013254:Steroid 17-alpha-Hydroxylase / Q000096:biosynthesis* / Q000235:genetics; D013736:Testicular Neoplasms / Q000473:pathology; D013739:Testosterone / Q000096:biosynthesis; D014162:Transfection; D014407:Tumor Cells, Cultured
D009363:Neoplasm Proteins; D011944:Receptors, Androgen; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D013196:Dihydrotestosterone; D013739:Testosterone; D003577:Cytochrome P-450 Enzyme System; D000242:Cyclic AMP; D013254:Steroid 17-alpha-Hydroxylase