LH Receptor C-terminus in Receptor Desensitization

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DESCRIPTION (provided by applicant): The desensitization of luteinizing hormone (LH) receptors, namely the appearance of a hormone-unresponsive state after hormone-induced signaling, is essential for completion of oocyte meiosis following the LH surge and for receptor internalization at other times in the ovarian cycle. Brief exposure to either LH or human chorionic gonadotropin (hCG) causes LH receptor desensitization and clustering of LH receptors in large, microscopically-visible structures. Receptors remain desensitized to hormone-triggered signaling until such time as these visible structures dissipate. The applicant hypothesizes that the C-terminus of the rat LH receptor contains information critical for desensitization of the receptor and for targeting of LH receptors to specific plasma membrane microdomains. To evaluate the role of the LH receptor C- terminus, the applicant will use chimeric receptors derived from "desensitization-resistant" gonadotropin releasing hormone (GnRH) receptors, which lack an intracellular C-terminus and which do not exhibit desensitization. The applicant will link these receptors to the LH receptor C-terminus (GnRH/cLH receptors) and determine whether the addition of this domain confers desensitization capacity on the GnRH receptor. In Specific Aim 1, the applicant will determine whether these chimeric receptors exhibit hormone-induced desensitization and, if so, how long this state, and related receptor self-association, persists. In Specific Aim 2, the applicant will assess whether hormone-treated GnRH/cLH receptors form large, visible clusters complexes like desensitized LH receptors or, if not, whether they form isolated dimers or small oligomers. In Specific Aim 3, the applicant will use single-particle tracking to determine whether desensitized GnRH/cLH and wild type LH receptors both occupy similar-sized membrane domains and whether these domain sizes change upon resensitization. These studies will draw on the experience of the Principal Investigator as well as the expertise of Dr. Colin Clay, a member of the Animal Reproduction and Biotechnology Laboratory, and of Dr. George Barisas, a biophysicist at Colorado State University. Although she will develop and utilize several state-of-the- art biophysical approaches to study G-protein receptor function, this project is, nonetheless, of somewhat limited scope and likely to be completed within two years.