Differentiation-dependent prolactin responsiveness and stat (signal transducers and activators of transcription) signaling in rat ovarian cells

PRL activates an important cytokine signaling cascade that is obligatory fo r maintaining luteal cell function in the rat ovary. To determine when spec ific components of this cascade are expressed and can be activated by PRL, we analyzed the expression of receptor subtypes (short, PRL-R-S, and long, PRL-R-L), the presence and kinetics of Stat (signal transducer and activato r of transcription) activation using the PRL-response element (PRL-RE) of t he alpha 2M (alpha 2-macroglobulin) gene, acid the content and hormonal reg ulation of three specific modulators of cytokine signaling; the tyrosine ph osphatases (SHP-1 and SHP-2), and the protein inhibitor of activated Stat3 (PIAS-3). These components were analyzed in differentiating granulosa/lutea l cells of hypophysectomized (H) rats and in corpora lutea of pregnant rats . Levels of PRL-R mRNAs increased as granulosa cells differentiated and rea ched maximal levels in luteal cells of pregnant rats where levels of PRL-R- S approached those of PRL-R,. The relative concentrations shifted from a 27 -fold excess of PRL-R, in preovulatory granulosa cells to a 3.7-fold differ ence in luteal cells during midgestation. Despite the increased PRL-R-L exp ression in differentiated granulosa cells, PRL did not stimulate detectable activation of Stats. Rather PRL activation of Stat5, principally Stat5b, o ccurred in association with luteinization. In contrast, granulosa cells of untreated immature and H rats contained a high level of DNA binding activit y, which was shown to be comprised entirely of activated, phosphorylated St at3. Treatment with estrogen and FSH reduced the amount of phosphorylated S tat3 and abolished its ability to bind DNA, an effect temporally related to increased PIAS-3. Expression of SHP-1 (but not SHP-2) was also hormonally regulated; SHP-1 mRNA and protein were high in granulosa cells of H rats, d ecreased by estrogen and FSH, and subsequently increased dramatically with luteinization. Of particular note, SHP-1 was localized in cytoplasm of gran ulosa cells in atretic follicles but was distinctly nuclear in luteal cells , indicative of different functional roles. Collectively, these results ind icate that Stat3 and Stat5 are activated by distinct cytokine-signaling pat hways modulated through differentiation-dependent transcriptional regulatio n of signaling pathway components and mediate distinct functional processes in the rat ovary: early follicle growth and atresia vs. luteinization.