ABSTRACT
This study was to explore estrogen receptor (ER) involvement in FSH and TGFbeta1-stimulated steroidogenesis in rat ovarian granulosa cells. We first determined the specific involvement of ERalpha and ERbeta in the process, and then investigated the molecular interaction of ERalpha and transcription coregulators in FSH and TGFbeta1 up-regulation of steroidogenic gene expression. Primary culture of ovarian granulosa cells from antral follicles of gonadotropin-primed immature rats was used. Interestingly, a selective ERalpha antagonist methyl-piperidino-pyrazole (MPP) [like ER antagonist ICI-182,780 (ICI)] decreased FSH +/- TGFbeta1-stimulated progesterone production, whereas an androgen receptor antagonist hydroxyflutamide and particularly a selective ERbeta antagonist 4-[2-Phenyl-5,7-bis(trifluoromethyl) pyrazolo [1,5-a] pyrimidin-3-yl] phenol had no significant effect. Consistent with this, a selective ERbeta agonist diarylpropionitrile (unlike 17beta-estradiol) also had no effect on FSH +/- TGFbeta1-stimulated progesterone production. Furthermore, a selective ERalpha agonist 4,4',4''-(4-Propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol (like 17beta-estradiol) enhanced FSH-stimulated progesterone production, and this was abolished by pretreatment with MPP. Immunoblotting and chromatin immunoprecipitation analyses indicate that MPP/ICI suppression of FSH +/- TGFbeta1 action is partly attributed to the reduced ERalpha-mediated expression of Hsd3b and Cyp11a1 genes, but not steroidogenic acute regulatory protein. Furthermore, FSH +/- TGFbeta1 increased ERalpha association with histone acetylases (CBP and SRC-1) and coactivator of peroxisome proliferator-activated receptor gamma (PGC-1alpha), and MPP/ICI dramatically reduced these interactions. In addition, FSH +/- TGFbeta1 increased CBP, SRC-1, and PGC-1alpha binding to Hsd3b and Cyp11a1 genes. Together, we demonstrate for the first time that ERalpha interaction with transcription coregulators, histone acetylases (CBP/SRC-1), and PGC-1alpha is crucial to FSH and TGFbeta1-up-regulated expression of Hsd3b and Cyp11a1, and, thus, progesterone production in rat ovarian granulosa cells.