This proposal is a request for funding of years 13 through 17 of an ongoing project designed to study membrane proteins involved in platelet aggregation. Platelet aggregation is a fundamental reaction in hemostasis and thrombosis and appears to be mediated by the glycoprotein (GP) IIb-IIIa complex in platelet membranes. Previous work from this laboratory has i) purified this glycoprotein complex and studied its physical, chemical and morphological properties; ii) identified and characterized some of the biological properties of GP IIb-IIIa such as binding of ligands and interaction with actin filaments; and iii) demonstrated that GP IIb-IIIa-like glycoproteins exist in endothelial cells and smooth muscle cells. The present study is designed to extend this later finding by characterizing the structural and functional properties of the GP IIb-IIIa-like glycoproteins of endothelial cell and smooth muscle cell membranes and by testing the hypothesis that these properties are related to those of GP IIb-IIIa in platelets. Monoclonal antibodies to the GP IIb-IIIa-like complex in endothelial cells and smooth muscle cells will be raised and used to purify the glycoproteins from these two cells and to test for potential functions of these glycoproteins. Biochemical and receptor-binding properties of the isolated glycoproteins will be determined and compared to platelet GP IIb-IIIa. Studies are outlined to deduce the primary amino acid sequences of GP IIb- and Gp IIIa-like glycoproteins using recombinant cDNA libraries prepared from endothelial cells. The primary sequences of the glycoproteins in nucleated cells will be compared to partial sequences determined for platelet glycoproteins to determine whether platelet and endothelial cell GP IIb-IIIa are products of the same gene or are members of a family of closely related genes. It is anticipated that this study will establish new information on the structure and function of the membrane glycoproteins in the cells of the blood vessel wall and will initiate a molecular biology approach to the study of regulation and organization of the genome(s) of the GP IIb-IIIa complex.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Method to Extend Research in Time (MERIT) Award (R37)
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Cor Therapeutics, Inc.
South San Francisco
United States
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Parise, L V; Steiner, B; Nannizzi, L et al. (1993) Evidence for novel binding sites on the platelet glycoprotein IIb and IIIa subunits and immobilized fibrinogen. Biochem J 289 ( Pt 2):445-51
Phillips, D R; Fitzgerald, L; Parise, L et al. (1992) Platelet membrane glycoprotein IIb-IIIa complex: purification, characterization, and reconstitution into phospholipid vesicles. Methods Enzymol 215:244-63
Phillips, D R (1992) Surface labeling of platelet membrane glycoproteins. Methods Enzymol 215:412-9
Lanza, F; Wolf, D; Fox, C F et al. (1991) cDNA cloning and expression of platelet p24/CD9. Evidence for a new family of multiple membrane-spanning proteins. J Biol Chem 266:10638-45
Steiner, B; Parise, L V; Leung, B et al. (1991) Ca(2+)-dependent structural transitions of the platelet glycoprotein IIb-IIIa complex. Preparation of stable glycoprotein IIb and IIIa monomers. J Biol Chem 266:14986-91
Lanza, F; Kieffer, N; Phillips, D R et al. (1990) Characterization of the human platelet glycoprotein IIIa gene. Comparison with the fibronectin receptor beta-subunit gene. J Biol Chem 265:18098-103
Sosnoski, D M; Emanuel, B S; Hawkins, A L et al. (1988) Chromosomal localization of the genes for the vitronectin and fibronectin receptors alpha subunits and for platelet glycoproteins IIb and IIIa. J Clin Invest 81:1993-8
Fitzgerald, L A; Phillips, D R (1988) The molecular biology of platelet and endothelial cell adhesion receptors. Prog Clin Biol Res 283:387-418
Smart, J L; Deth, R C (1988) Influence of alpha 1-adrenergic receptor stimulation and phorbol esters on hepatic Na+/K+-ATPase activity. Pharmacology 37:94-104
Phillips, D R; Charo, I F; Parise, L V et al. (1988) The platelet membrane glycoprotein IIb-IIIa complex. Blood 71:831-43

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