title: Aromatase cytochrome P450 in rat ovarian granulosa cells before and after luteinization: adenosine 3',5'-monophosphate-dependent and independent regulation. Cloning and sequencing of rat aromatase cDNA and 5' genomic DNA.
abstract: The complete nucleotide sequence for rat ovarian aromatase cytochrome P450 (P450arom) has been derived from four cDNA clones isolated from three rat granulosa/luteal cell lambda gt11 cDNA expression libraries. The composite P450arom cDNA extends 1597 basepairs, encodes a protein of 508 amino acids (calculated mol wt = 58,263), and hybridizes to three mRNA transcripts (3.3, 2.6, and 1.9 kilobases in size) in rat ovarian tissues. A 5' genomic fragment was isolated from a rat genomic library and shown to contain exon I and 538 basepairs of 5' flanking sequences, including putative promoter elements. Further, we document that P450arom mRNA and estradiol (E) biosynthesis are regulated by cAMP-dependent mechanisms in granulosa cells of preovulatory (PO) follicles, but are maintained by cAMP-independent mechanisms after LH/hCG-induced luteinization. The transition of the PO granulosa cell to the luteal cell (PO + hCG) phenotype requires 5 h of exposure to hCG in vivo. Once the luteal cell phenotype is programed, P450arom mRNA and E biosynthesis are maintained in the luteinized cells for up to 10 days in a constitutive manner in the absence of hormones or agents that increase intracellular cAMP. Furthermore, when PO + hCG (7 h) follicles were isolated and incubated for 1-3 h with reversible inhibitors of transcription (actinomycin-D) or translation (cycloheximide) before harvesting the granulosa cells, neither morphological nor functional luteinization of granulosa cells in culture was impaired. Thus, rapid cellular and molecular events occur in granulosa cells within 5-7 h after an ovulatory LH/hCG surge that alter the hormonal regulation of the aromatase gene.
authors.affiliation: Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: G J
authors.initials: GJ
authors.lastname: Hickey
authors.forename: J S
authors.initials: JS
authors.lastname: Krasnow
authors.forename: W G
authors.initials: WG
authors.lastname: Beattie
authors.forename: J S
authors.initials: JS
authors.lastname: Richards
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001141:Aromatase / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000494:pharmacology*; D004247:DNA / Q000235:genetics*; D005260:Female; D006065:Gonadotropins, Pituitary / Q000494:pharmacology*; D006107:Granulosa Cells / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D051381:Rats; D011919:Rats, Inbred Strains; D015183:Restriction Mapping
chemical_list: D006065:Gonadotropins, Pituitary; D004247:DNA; D000242:Cyclic AMP; D001141:Aromatase