Granulosa cells in growing ovarian follicles respond to follicle stimulating hormone (FSH) by the sequential expression of specific genes. Those that are induced rapidly by FHS appear related to granulosa cell proliferation whereas others that are induced in a delayed manner are associated with differentiation. Genes induced rapidly by FSH included cyclin D2 which regulates cyclin dependent kinases (cdks 4/6) and cell cycle progression, as well as sgk (serum and glucocorticoid induced kinase) and snk (serum induced kinase) which are members of a novel, inducible serine-threonine kinase family. Genes induced later by FSH include RIIbeta (the A-kinase type II beta regulatory subunit), aromatase cytochrome P450 (CYP 19), and the LH receptor. In addition, we have recently observed that granulosa cells cultured in the continuous presence of low levels of FSH or forskolin exhibit biphasic responses. Genes that are regulated by FSH in an oscillatory manner include cyclin D2 and sgk. Induction of early events coincides with the rapid production of cAMP, translocation of the catalytic (C) subunit of A-kinase to the nucleus and transient phosphorylation of the cAMP regulatory element binding protein (CREB). Differentiation of granulosa cells occurs between 24-48 h when cAMP levels are reduced but nuclear levels of phosphoCREB and A-kinase C subunit are again elevated. The preeminent role of cAMP and A-kinase in mediating the early, oscillatory and delayed actions of FSH in granulosa cells is countered by the apparent cAMP-independent function of these cells once they have undergone terminal differentiation to luteal cells. The focus of the current studies will be to determine what cellular mechanisms regulate the oscillatory pattern of FSH action via the A-kinase pathway to control gene activation during granulosa cell proliferation and differentiation. For this we will extend our current studies on FSH- regulated gene expression to determine: I. The cellular and molecular mechanisms regulating the oscillating patterns of A-kinase activity, localization and CREB phosphorylation in granulosa cells; II. The molecular mechanisms controlling expression and function of cell cycle related and inducible kinases; and III. The molecular mechanisms controlling the expression of aromatase. The results of these studies should provide novel, useful information on the essential role of FSH/cAMP/A-kinase, as well as the factors that they control, in regulating granulosa cell proliferation and differentiation; thereby promoting normal follicular growth while preventing abnormal proliferative activity.
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