The overall objective of this proposal is to examine the molecular mechanism(s) involved in the regulation of ovarian function by leuteinizing hormone (LH). The initial event is the interaction of LH with a specific receptor on the ovarian cell surface. Through this interaction with a specific cell surface receptor LH regulates ovarian follicular development, ovulation and corpus luteum function. By these means LH regulates fertility, implantation and early pregnancy. The present study focuses on the regulation of the expression of LH receptor on the cell surface. Specifically, the first specific aim examines the mechanism by which the steady state levels of LH receptor are regulated during follicle maturation and during the peri-ovulatory periods. The specific studies oroposedto examine this aim will elucidate the role of a newly identified protein, LH receptor mRNA binding protein, in the post- transcriptional regulation of LH receptor mRNA. These studies will determine the mechanism by which LH receptor mRNA is degraded by LH receptor mRNA binding protein. The specific studies proposed will also make use of an in vitro LH receptor mRNA decay assay to examine the LH receptor mRNA degradation rates, aid an electrophoretic mobility gel shift assay to examine the LH receptor mRNA binding activity of the binding protein. The second specific aim will determine the role of post-translational modification on the turnover of LH receptor on the cell surface, Specifically, this aim will address the role of palimitoylation/depalmitoylation of LH receptor on receptor turnover after interaction with LH.
This aim will also address the mechanism(s) involved in the arrestin-mediated internalization of the LH-bound receptor. The studies will utilize among other techniques, pulse labeling of epitope- tagged LH receptor and immunoprecipitation of the labeled receptor. The third specific aim will address the role of post-translational modification of the LH receptor on its folding and cell surface expression. The proposed experiments in specific aim 3 would involve mutation of the consensus glycosylation sites of the LH receptor and transfection of the receptor constructs to examine the role of glycosylation on folding and cell surface expression. These studies address novel questions central to reproductive endocrinology and are directly relevant to the diagnosis and treatment of infertility.
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