This research has the long-range goal of understanding the relationships and interactions of circulating blood platelets in the process of blood coagulation and in the cellular responses of inflammation, repair, and atherosclerosis. During coagulation, platelets release the platelet derived growth factor (PDGF), the principal mitogenic protein in serum for cells of mesenchymal origin. Our research focuses on the biochemistry/mechanism of action of this protein. Because malignant cells respond differently to PDGF than do normal cells, our research also has the goal of understanding further the differences in the cellular responsiveness of normal and malignant cells in culture to PDGF. Our laboratory has purified PDGF to apparent homogeneity, studied in detail the binding of PDGF to surface receptors on Swiss mouse 3T3 cells, and described different responses of cultured cells to this potent mitogen. The present proposal builds upon previous work and is directed to the cell surface receptor for PDGF. We plan to further characterize the 3T3 cell membrane protein stimulated in tyrosine phosphorylation by PDGF (thought to be the PDGF receptor protein) by peptide mapping, to further identify the cell surface receptor by crosslinking the receptor protein to 125I-PDGF, to solubilize the PDGF receptor protein from 3T3 cell membranes, to purify the PDGF receptor protein to apparent homogeneity by affinity chromatography, to characterize the purified receptor protein by physical and chemical criteria, to characterize the anticipated associated receptor protein kinase activity by kinetic criteria and by identifying protein substrates for receptor kinase activity, to obtain monoclonal antibodies to the PDGF receptor protein, and to purify and characterize the PDGF receptor protein from other cell surfaces. These investigations will provide basic information on receptor function, on the regulation of normal cell growth, and on differences in cellular responsiveness of normal and malignant cells to mitogenic proteins. It is hoped that this information will provide further understanding of the processes of inflamation, repair, and atherosclerosis, and new insight into the understanding of how normal cells differ from the malignant cells.
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