Effects of Caveolin Disruption on Signaling from the hFSH Receptor

Follicle stimulating hormone is produced by the pituitary gland, regulating the function of both the ovaries and the testes. The inability of this hormone to bind to its receptor can cause infertility in both men and women. Human follicle stimulating hormone receptor (hFSHR) is a GPCR (g protein-coupled receptor) that plays a crucial role in the development of sperm and egg cells in the gonads. The successful interaction of FSH with its receptor results in activation of several downstream intracellular pathways including the production of cAMP. Currently, hFSHR signaling is thought to rely on the protein caveolin. Caveolin is a protein found in the caveolae of the cell membrane. Caveolae, a type of lipid raft, support a diverse group of receptors that are needed at the cell membrane. The ability of hFSHR to bind to the lipid raft is believed to be accomplished through an interaction with the transmembrane domain IV of the hFSHR with the caveolin protein. An interruption in the caveolin binding motif of the hFSHR could have a large effect on the development of sperm and egg cells and subsequently cause infertility. In this study, we introduced wild type and mutant peptides corresponding to the 4th transmembrane domain of the hFSHR into cells expressing the receptor. We expect the wild type peptide to interfere with the interaction between caveolin and its binding motif. Additionally, we expect the mutant peptide to not interfere with this interaction. Currently, we are using a quantitative fluorescent assay to measure cAMP production in order to determine if the peptide is interfering with the activation of the hFSHR. I hypothesize that if caveolin is prevented from binding to hFSHR, the receptor will become less active. I anticipate a decrease in levels of cAMP in the cells that are treated with the wild-type (interfering) peptide. There should be no difference in the level of cAMP in cells treated with the mutant peptide and the cells with no peptide. Preliminary trials have suggested that there is a difference in levels of cAMP produced between the two peptides. In the future, we plan on using scrambled peptides to validate our current observations.