Cross-talk between cAMP and p53-generated signals in induction of differentiation and apoptosis in steroidogenic granulosa cells

In each menstrual cycle only very few follicles in the mammalian ovary undergo maturation and ovulation while most of the follicles degenerate in the process of atresia. Moreover, in the absence of pregnancy, the newly formed corpora lutea will degenerate and disappear in the process of luteolysis. Recent studies suggest that ovarian follicular atresia is associated with DNA fragmentation and degeneration of follicular cells, characteristic of programmed cell death (apoptosis). Apoptosis can be induced in vitro, in primary granulosa cell culture, by serum deprivation and by induction of a high intracellular level of cAMP. This induction of apoptosis can be blocked by fibroblast growth factor, suggesting that receptor-mediated activation of a tyrosine kinase can serve as a survival signal. Apoptosis can also be induced in immortalized steroidogenic granulosa cells, transformed by SV40 DNA and Ha- ras oncogene, by overexpression of the wild-type p53 tumor suppressor gene in cAMP-stimulated cells. Omitting the cAMP stimulus prevents the p53-induced apoptosis in these cells, suggesting cross-talk between p53 and cAMP-generated signals in the induction of apoptosis. Steroidogenic activity in these cells, as well as in nontransformed granulosa cells, does not decline during apoptosis but is rather significantly elevated before total cell collapse occurs. Cytochemical studies using confocal laser microscopy, electron microscopy, and three-dimensional reconstruction reveal a specific reorganization pattern of proteasomes, the most abundant nonlysosomal protease, and of the steroidogenic organelles, such as mitochondria and lipid droplets, in the apoptotic cell. Our results suggest that compartmentalization of intracellular organelles during apoptosis permits proteolysis without interfering with steroidogenesis, characteristic of the differentiated phenotype of the granulosa cell. Moreover, cytoskeletal rearrangement may serve as a barrier between these cellular activities.