LUTEINIZATION IN PRIMATES IS ACCOMPANIED BY LOSS OF A 43-KILODALTON ADENOSINE-3',5'-MONOPHOSPHATE RESPONSE ELEMENT-BINDING PROTEIN ISOFORM

Although granulosa cell differentiation and corpus luteum function are both regulated by cAMP, there are development-dependent differences, particularly at the level of gene expression and cell proliferation, b etween the responses of follicular granulosa cells and luteal cells to trophic hormone stimulation. In this study, we sought to determine wh ether these differences could be due to changes in the cellular expres sion of cAMP response element (CRE)-binding protein (CREB). Immunocyto chemical analysis of macaque ovaries revealed a development-related al teration in the subcellular distribution of CREB-immunoreactive materi al. Immunoreactive CREB was present in nuclei of follicular granulosa cells from maturing follicles, whereas after ovulation and luteinizati on, no CREB-immunoreactive proteins were visualized in luteal cell nuc lei. Anti-CREB immunoblotting of granulosa cell extracts from macaque preovulatory follicles as well as extracts of granulosa cells from lut einizing human follicles revealed a 43-kilodalton (kDa) protein, a siz e typical of native CREB. In contrast, whole cell extracts of monkey c orpora lutea collected during the early, mid-, and late luteal phases completely lacked a 43-kDa CREB signal. The absence of 43-kDa CREB iso forms in corpora lutea was confirmed using three different antisera di rected against different regions of CREB. Using a human collagenase ge ne CRE to probe Southwestern blots, a 43-kDa CREB was observed in foll icular cell extracts, whereas no CRE-binding activity was found in cor pora lutea extracts using this probe. We also sought to determine whet her the loss of expression of the 43-kDa CREB isoform may be functiona lly correlated with the cessation of cellular proliferation that accom panies luteinization. Expression of proliferating cell nuclear antigen (PCNA), an obligatory component of DNA polymerase delta, is essential for proliferation and has been shown by others to be CRE dependent. I mmunoblotting of follicle cell and luteal cell extracts with an anti-P CNA monoclonal antibody revealed PCNA expression in granulosa cells an d no detectable PCNA expression in corpora lutea. These findings indic ate that as follicular granulosa cells progress from the proliferative state to terminally differentiated luteal cells, there is a cessation of expression of a 43-kDa member of the CREB family of transcription factors, and there may be an association between the loss of CREB isof orms and cessation of PCNA expression.