Evidence for the presence of cholesterol side chain cleavage cytochrome P-450 and adrenodoxin in fresh granulosa cells. Effects of follicle-stimulating hormone and cyclic AMP on cholesterol side chain cleavage cytochrome P-450 synthesis and activity

B. Funkenstein, M. R. Waterman, B. S.S. Masters, E. R. Simpson

Research output: Contribution to journalArticlepeer-review

27 Scopus citations

Abstract

The synthesis of cholesterol side chain cleavage cytochrome P-450 (cytochrome P-450(scc) and adrenodoxin was studied both in freshly harvested bovine granulosa cells and in granulosa cells maintained in primary monolayer culture. In addition, the action of folicle-stimulating hormone (FSH) and cyclic AMP analogs to stimulate the synthesis of cytochrome P-450(scc) was investigated in cultured cells. Precursor forms of cytochrome P-450(scc) and adrenodoxin were immunoisolated from a cell-free translation system directed by RNA prepared from freshly obtained granulosa cells that were not luteinized. Furthermore, the presence of cytochrome P-450(scc) in lysates of granulosa cells freshly obtained from very small follicles (containing less than 0.1 ml of follicular fluid) and in mitochondria of freshly obtained granulosa cells was demonstrated by using an immunoblotting technique. Continuous treatment of cultured granulosa cells with FSH or with cyclic AMP analogs (dibutyryl cyclic AMP or 8-bromo cyclic AMP) for 72 h increased incorporation of [35S]methionine into immunoprecipitable cytochrome P-450(scc). Moreover, FSH, dibutyryl cyclic AMP, and 8-bromo cyclic AMP stimulated pregnenolone production by cultured granulosa cells (2.3-, 4.0-, and 7.5-fold increase over control, respectively), indicative of an increase in cholesterol side chain cleavage activity. The results of this study demonstrate for the first time the presence of two components of the cholesterol side chain cleavage system in freshly obtained granulosa cells, and provide direct evidence for the trophic effect of FSH and its presumed mediator, cyclic AMP, on the synthesis of cytochrome P-450(scc) in granulosa cells.

Original languageEnglish (US)
Pages (from-to)10187-10191
Number of pages5
JournalJournal of Biological Chemistry
Volume258
Issue number16
StatePublished - 1983

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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