Activation of CYP11A and CYP11B gene promoters by the steroidogenic cell-specific transcription factor, Ad4BP.
We have examined the transcriptional activity of four cis-elements, Ad1(CRE), Ad2, Ad3, and Ad4, that are present in the promoter of the bovine CYP11B (11 beta-hydroxylase P-450) gene using beta-globin reporter gene constructs and transient transfection into steroidogenic and nonsteroidogenic cell types. Only Ad1(CRE), a CRE homolog, showed forskolin-dependent transcriptional activity in adrenal tumor Y-1 cells, whereas the other elements were not able to stimulate transcription by themselves. As Ad3 and Ad4 had previously been identified as the cis-elements required for full cAMP-dependent transcription of this gene, we examined the effect of combinations of different cis-elements on the transcription of the reporter gene. In Y-1 cells, Ad1(CRE) and four tandem copies of any one of the other cis-elements substantially activated transcription in response to forskolin treatment. The template carrying Ad1(CRE) and Ad4 was also active in testicular Leydig cells, I-10, whereas it was inactive in nonsteroidogenic PC-12 cells. Transcriptional activation by the 4xAd4/Ad1(CRE) combination presumably depended on the presence of Ad4-binding protein (Ad4BP), which is absent in PC-12 cells, as shown by immunoblot analysis. This was confirmed by cotransfecting an expression vector for Ad4BP into PC-12 cells, which caused forskolin-dependent transcription to increase in proportion to the amount of expression vector. In Y-1 cells, transcriptional activation by forskolin was mimicked by cotransfection of an expression vector for the catalytic subunit of protein kinase-A.(ABSTRACT TRUNCATED AT 250 WORDS)
Morohashi
K
K
Department of Molecular Biology Graduate School of Medical Science, Kyushu University, Fukuoka, Japan.
Zanger
U M
UM
Honda
S
S
Hara
M
M
Waterman
M R
MR
Omura
T
T
D000310:Adrenal Gland Neoplasms; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002460:Cell Line; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D051717:Fushi Tarazu Transcription Factors; D015971:Gene Expression Regulation, Enzymologic; D018398:Homeodomain Proteins; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis; D016716:PC12 Cells; D010957:Plasmids; D011401:Promoter Regions, Genetic; D018160:Receptors, Cytoplasmic and Nuclear; D015183:Restriction Mapping; D013252:Steroid 11-beta-Hydroxylase / Q000096:biosynthesis / Q000235:genetics*; D054339:Steroidogenic Factor 1; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D009838:Oligodeoxyribonucleotides; D018160:Receptors, Cytoplasmic and Nuclear; D054339:Steroidogenic Factor 1; D014157:Transcription Factors; D003577:Cytochrome P-450 Enzyme System; D013252:Steroid 11-beta-Hydroxylase