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Department of Cell Biology, Baylor College of Medicine Houston, Texas 77030
Department of Obstetrics and Gynecology, Baylor College of Medicine Houston, Texas 77030
Address requests for reprints to: Dr. JoAnne S. Richards, Department of Cell Biology, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030.
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 
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.
FOOTNOTES
This work was supported by NIH Grants HD-16229 and HD-16272 (to J.S.R.) and the American Cancer Society (BC627; to J.S.R.).
Received for publication September 18, 1989. Revision received October 18, 1989. Accepted for publication October 23, 1989.
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