Summary
Using a 5′ human P450scc cDNA probe and a human 57-mer oligonucleotide probe, 2 rat P450scc cDNA clones were identified and isolated from combined granulosa cell and luteal cell λgt11 cDNA expression libraries. One 350 bp clone contained the start codon. The other 750 bp clone overlapped on the 5′ end with the 350 bp clone and on the 3’ end with a previously isolated 1.2 kb cDNA clone (1), which contains a poly (A) tail, suggesting that the complete cDNA sequence for rat P450scc is encoded in these 3 cDNA clones. Using the same 5′ human P450scc probes, we have isolated a rat genomic P450scc clone from a rat liver genomic library. The genomic clone contained 3 distinct Bam HI fragments, 3.7, 4.0 and approximately 14 kb in size. The 3.7 and 4.0 kb Bam HI fragments hybridized to the 1.2 kb cDNA and appear to contain 3′ located exons. After subcloning, the 14 kb fragment yielded several transformants with inserts ranging from 8 to 0.5 kb in size. Hybridization with the rat cDNA clones revealed that the 8 kb fragment may represent 5′ upstream or intron sequences. A 6 kb fragment contained the first and probably the next 2 exons. Northern blot hybridization with the analysis, using the 750 bp cDNA probe, demonstrated that P450scc mRNA was low in granulosa cells of preovulatory (PO) follicles but could be induced by an ovulatory dose of LH/hCG in vivo or by forskolin added in vitro. Once induced by LH/hCG in vivo, P450scc mRNA and progesterone biosynthesis were constitutively maintained by luteinized cells in vivo or in vitro. Thus, the induction of P450scc mRNA in granulosa cells from developing follicles is hormone and cAMP dependent. In contrast, the expression of P450scc mRNA in luteinized granulosa cells has become cAMP independent.
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References
Goldring NB, Durica JM, Lifka J, et al. Cholesterol side-cleavage P450scc messenger ribonucleic acid: evidence for hormonal regulation in rat ovarian follicles and constitutive expression in corpora lutea. Endocrinology 1987; 120:1942–50.
Richards JS, Jahnsen T, Hedin L, et al. Ovarian follicular development: from physiology to molecular biology. Recent Prog Horm Res 1986; 43:231–76.
Richards JS, Hedin L, Caston L. Differentiation of rat ovarian thecal cells: evidence for functional luteinization. Endocrinology 1986; 118:1660–8.
Chung B, Matteson KJ, Voutilainen R, Mohandas TK, Miller WL. Human cholesterol side-chain cleavage enzyme, P450scc: cDNA cloning, assignment of the gene to chromosome 15, and expression in the placenta. Proc Natl Acad Sci USA 1986; 83:8962–6.
Morohashi K, Fujii-Kuriyama Y, Okada Y, et al. Molecular cloning and nucleotide sequence of cDNA for mRNA of mitochondrial cytochrome P-450 (SCC) of bovine adrenal cortex. Proc Natl Acad Sci USA 1984; 81:4647–51.
Mullis KB, Faloona FA. Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Methods Enzymol 1987; 155:335–50.
Saiki RK, Gelfand DH, Stoffel S, et al. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 1988; 239:487–91.
Pelham HRB. A regulatory upstream promoter element in the Drosophila Hsp 70 heat shock gene. Cell 1982; 40:517–28.
Morohashi K, Sogawa K, Omura T, Fujii-Kuriyama Y. Gene structure of human cytochrome P450 (SCC) cholesterol desmolase. J Biochem (Tokyo) 1987; 101:879–87.
Hedin L, Rodgers RJ, Simpson ER, Richards JS. Changes in content of cytochrome P450 17a, cytochrome P450scc, and 3-hydroxy-3-methylglutaryl CoA reductase in developing rat ovarian follicles and corpora lutea: correlation with theca cell steroidogenesis. BiolReprod 1987; 37:211–33.
Hickey GJ, Chen S, Besman MJ, et al. Hormonal regulation, tissue distribution, and content of aromatase cytochrome P450 messenger ribonucleic acid and enzyme in rat ovarian follicles and corpora lutea: relationship to estradiol biosynthesis. Endocrinology 1988; 122:1426–36.
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© 1989 Plenum Press, New York
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Oonk, R.B., Jansen, R., Hickey, G.J., Beattie, W.G., Richards, J.S. (1989). The Rat Cholesterol Side-Chain Cleavage Cytochrome P450 (P450scc) Gene: cAMP-Dependent and -Independent Regulation in Ovarian Cells. In: Hirshfield, A.N. (eds) Growth Factors and the Ovary. Springer, Boston, MA. https://doi.org/10.1007/978-1-4684-5688-2_41
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DOI: https://doi.org/10.1007/978-1-4684-5688-2_41
Publisher Name: Springer, Boston, MA
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