Guanine nucleotide regulatory proteins (G proteins) are a family of homologous polypeptides that are required for receptor-mediated regulation of retinal cGMP phosphodiesterase, adenylyl cyclase, phospholipases C and A2, and subsets of K+ and Ca2+ ion channels. These signal-transducing molecules are involved in fundamental processes of cell modulation, as well as in highly specialized cell function, such as phototransduction and olfaction. Little is known about specific G proteins and the molecular biology of signal transduction in retinal pigment epithelial (RPE) cells. The RPE is a highly specialized monolayer of cells with a close functional relationship with photoreceptors. The multiple functions of the RPE, if disturbed, may lead to a variety of visual disorders, including serous retinal detachment and retinal degeneration. In proliferative vitreoretinopathy, RPE cells are believed to undergo abnormal chemotaxis, migration, and proliferation. A hypothesis of this proposal is that important behaviors of phagocytic RPE cells, such as chemotaxis, transformation, and proliferation, involve regulation by G proteins and phosphoinositide phospholipase C, since these signal-transducing molecules are the basis for the regulation of similar activities that have been studied intensively in macrophages, neutrophils, and other cell types. The long-term goal of this study is to determine the molecular basis for the regulation of phosphoinositide phospholipase C by RPE GTP-binding proteins and to develop specific reagents that interact with these signal-transducing molecules to modify cell response. To achieve the goals of this project, RPE cells from bovine and human tissue will be cultured by established methods. In these cultured cells, the regulation of phospholipase C by chemoattractants and growth factors will be studied, and the expression of RPE G proteins will be analyzed by molecular genetics. In addition, a human RPE cDNA library will be constructed from homogeneous cell cultures for the identification of novel G protein gene products in RPE. The effects of G protein antagonists, such as pertussis toxin, synthetic peptides, and inhibitory antibodies, on phospholipase C and RPE cell responses will be determined. This work may provide a basis for the rational development of reagents or drugs that interact with specific G proteins and other signal-transducing molecules in RPE.
Showing the most recent 10 out of 19 publications
