17alpha-Hydroxylase gene expression in the bovine ovary: mechanisms regulating expression differ from those in adrenal cells.
17alpha-Hydroxylase cytochrome P450 (P450(17alpha)) is the enzyme which synthesizes C19 steroids in a two-step reaction in which 17alpha-OH pregnenolone is an intermediate. In the bovine and human adult female, 17alpha-hydroxylase is expressed in adrenocortical cells where 17alpha-OH pregnenolone and 17alpha-OH progesterone are precursors of cortisol, and in theca cells of the ovary where these intermediates are precursors of C19 steroids. In both adrenal cortex and theca, 17alpha-hydroxylase gene expression is stimulated by cyclic AMP (cAMP). The aim of this study was to determine the mechanism regulating 17alpha-hydroxylase gene expression in the bovine ovary. Our results indicate that the bovine 17alpha-hydroxylase gene is regulated in a tissue-specific fashion. Primer extension and S1 nuclease protection assays reveal that the start site of transcription in the theca is identical to that in the adrenal. Transfection studies employing beta-globin reporter gene constructs fused to successive deletions of the 5' regulatory region of the bovine 17alpha-hydroxylase gene indicate that sequences between -80 and -37 basepairs (bp) (CRS2) confer cAMP-regulated transcription in bovine theca cells in culture. These results are in contrast to similar studies conducted in bovine adrenocortical cells, which indicate that the major cAMP response element (referred to as CRS1) is located at -243 to -225 bp. The Ad4 element (AGGTCA, -42 to -37 bp) within CRS2, which has been shown to be involved in cAMP responsiveness in other steroidogenic P450 genes, cannot by itself confer cAMP-regulated reporter gene expression in bovine cells. These results indicate that in the cow, 17alpha-hydroxylase gene expression is regulated in a tissue-specific fashion, and that this regulation may be conferred, at least in part, by the use of tissue-specific cis-acting elements in the bovine 17alpha-hydroxylase gene.
Demeter Arlotto
M
M
Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas 75235-9051, USA.
Michael
M D
MD
Kilgore
M W
MW
Simpson
E R
ER
D000302:Adrenal Cortex / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D000242:Cyclic AMP / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000235:genetics; D004790:Enzyme Induction; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D015290:Second Messenger Systems; D013254:Steroid 17-alpha-Hydroxylase / Q000096:biosynthesis* / Q000235:genetics; D013799:Theca Cells / Q000378:metabolism*; D014162:Transfection
D011993:Recombinant Fusion Proteins; D003577:Cytochrome P-450 Enzyme System; D000242:Cyclic AMP; D013254:Steroid 17-alpha-Hydroxylase; C022433:pregnenolone 17-alpha-hydroxylase