Signaling and Signal Transduction in Endocrine Tissues
(M757) SIGNALING MECHANISM OF FSH AND TGF1 IN STIMULATING STEROIDOGENESIS IN RAT OVARIAN GRANULOSA CELLS.
Chen, Yun Ju 1, Ke, Ferng-Chun 2, Lee, Ming-Ting3, Lee, Ping-Ping 3, Hwang, Jiuan-Jiuan 1, 1 Institute of Physiology, National Yang-Ming University, Taipei, Taiwan2 Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan3 Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
ABSTRACT- Previous studies demonstrate that transforming growth factor 1 (TGF1) augmented FSH-induced progesterone production in rat ovarian granulosa cells. The post-receptor signaling relaying FSH action rests mainly on cAMP followed by protein kinase A (PKA)-dependent and/or PKA-independent pathway possibly involving phosphatidylinositol-3-kinase (PI3K). This study was conducted to investigate the role of cAMP/PKA and PI3K pathways in FSH and TGF1-regulated steroidogenesis. Granulosa cells of eCG-primed immature rats were pretreated with PKA inhibitor (PKAI), PI3K inhibitor (wortmannin), and/or inhibitor of mammalian target of rapamycin/mTOR (rapamycin) for 1 h, and then treated with FSH and/or TGF1 (1) for 48 h to determine their effects on progesterone levels and protein levels of steroidogenic acute regulatory protein (StAR) and cholesterol side-chain-cleavage enzyme (P450scc), and (2) for 1 h to determine their effects on the activation of the PI3K downstream signaling molecules including Akt/PKB, serum and glucocorticoid-induced kinase (Sgk), mTOR, p70 ribosomal protein S6 kinase (S6K) and 4E-binding protein 1 (4E-BP1). PKAI, wortmannin and rapamycin suppressed FSH + TGF1-induced progesterone production. And only PKAI and wortmannin reduced FSH or 8-Br-cAMP stimulatory effect. Furthermore, the inhibitory effect of PKAI and wortmannin in combination was similar to those of either treatment alone. In addition, TGF1 did not augment 8-Br-cAMP effect as it did on FSH effect. And unlike 8-Br-cAMP, activation of cAMP-GEF with 8-CPT-2,-O-Me-cAMP had no influence on progesterone production. The suppressive effect of PKAI and wortmannin on FSH + TGF1-induced progesterone production may attribute partly to their effects on P450scc and StAR protein levels. We further demonstrate that FSH ± TGF1 increased the phosphorylation of Akt, mTOR and p70 S6K (but not Sgk and 4E-BP1), and this was suppressed by wortmannin. Also, rapamycin reduced the phosphorylation of p70 S6K. In conclusion, FSH may act through cAMP/PKA- and PI3K-mediated signaling in stimulating steroidogenesis in rat granulosa cells, and TGF1 appear to act at upstream of cAMP and/or cAMP-independent manner to augment FSH effect.
KEY WORDS: FSH & TGF1, steroidogenesis, PKA & PI3K, ovarian granulosa cells