Activin is an ovarian-produced protein that regulates follicle function. The investigators hypothesize that the ability of follicles to respond to activin is controlled by the type of receptor subunits present on the cell membrane. Therefore, the investigators will investigate the presence of activin receptor subunits (RII-ligand binding and RI-signal transduction) in the granulosa cell at times following hormonal stimulation. Receptor activation stimulates cytoplasmic signaling molecules. Candidate signal proteins have been identified (Smad2 and Smad4) and the investigators have shown that these proteins are activated by activin in granulosa cells. They hypothesize that normal, activin-regulated granulosa cell function is dependent on the appropriate interaction of ligand with Smad2 or Smad4 and that alteration in the activin signal transduction pathway will lead to abnormal follicle function. These tenets will be tested in granulosa cell culture and in transgenic animals. The transcriptional response of activin-regulated promoters (p3TP and junB) will be measured following over-expression of normal or mutated signal proteins in a granulosa cell line, GRM02. The genetic sequence that is responsive to activin signals will be identified (activin-response element or ARE). Transgenic animals with ovarian-targeted (inhibin alpha subunit promoter) mutated signal protein (Smad2-dominant negative) or normal and mutated receptor (ActRII-dominant negative and ActRII-constitutively active) will be generated and the consequence of over expression of these proteins will be evaluated in terms of follicle growth, selection, and maturation. These studies will establish the causal relationship between the activin system and normal and abnormal follicle function.
