Rapid Induction of Prostaglandin Endoperoxide Synthase in Rat Preovulatory Follicles by Luteinizing Hormone and cAMP is Blocked by Inhibitors of Transcription and Translation

Abstract

Prostaglandin endoperoxide synthase (PGS) catalyzes the rate-limiting step in the synthesis of prostaglandins E and F2 alpha that are obligatory for ovulation. To understand the molecular mechanisms by which LH regulates the induction of PGS in rat preovulatory (PO) follicles, we established an in vitro system which mimics in vivo induction of the enzyme. We show that the rapid increase in PGS enzyme: 1) is stimulated by LH, FSH, and forskolin (cAMP) in a time- and dose-dependent manner that is distinct from changes in steroidogenic enzymes analyzed in the same follicles; 2) is unaltered by end products (PGE and PGF2 alpha) of the reaction or inhibitors (indomethacin) of enzyme activity; 3) is blocked by inhibitors of transcription (alpha-amanitin) and translation (cycloheximide) at a step distal to production of cAMP and activation of A-kinase. Analyses of PGS mRNA by Northern blots using a mouse PGS cDNA probe revealed a PGS transcript of 2.8 kilobases that was present but in low abundance in PO follicles and decreased rapidly (1-4 h) as a consequence of LH/human CG stimulation. This hormone-induced decrease in PGS mRNA appeared to be transient because PGS transcripts were present in corpora lutea at a level similar to that in PO follicles. These results raise the possibility that the marked increase in PGS enzyme in granulosa cells of PO follicles that occurs as a consequence of the LH/human CG surge may not involve a cAMP regulated increase in transcription of the PGS gene itself. Or, if increased transcription of PGS mRNA does occur, it is rapid and coupled with cotranslational degradation of the mRNA. Alternatively, transcriptional (alpha-amanitin-sensitive) regulation of a separate gene may be obligatory for increased translation of PGS mRNA or posttranslational modification (stabilization?) of the enzyme. In summary, the LH-stimulated appearance of PGS in granulosa cells of PO follicles before ovulation is mediated by cAMP in a complex manner involving transcriptional regulation (PGS gene?) and translational control of PGS mRNA. The transient appearance of the PGS enzyme represents a unique pattern of response by granulosa cells of PO follicles to LH/cAMP and thereby may involve novel intracellular factors and regulatory processes.