The overall goal of this project is to characterize the molecular mechanisms of vascular cell signaling by the Class B scavenger receptor CD36. CD36 is expressed on macrophages and capillary endothelial cells, where it has been implicated in important vascular processes. As a macrophage receptor for oxidatively modified low density lipoprotein particles it is involved in directly internalization of bioactive lipids and foam cell formation during atherogenesis. As a receptor on phagocytes for apoptotic cells, it participates in their clearance and in mediating the accompanying cellular response. As a receptor on endothelium for the matrix glycoprotein thrombospondin-1 it mediates a potent anti- angiogenic activity. The molecular mechanisms by which CD36 mediates diverse actions in different cell types remain largely unknown and is the focus of this application. Dissecting these signaling pathways will have important implications in understanding regulation of macrophage and endothelial cell activation, and in defining potential new therapeutic targets for atherosclerosis inflammation, and cancer. These studies will take advantage of several unique reagents, including cells isolated from CD36 null mice, peptides derived from the cytoplasmic domain of CD36, and CD36 homolog is identified in the nematode Caenorhabditis elegans. Specific plans are to identify candidate signaling molecules and pathways involved in CD36- mediated cellular events by characterizing proteins that interact physically with CD36 in vascular cells. Strategies will include affinity chromatography on CD36 cytoplasmic peptides, co- immunoprecipitation, and yeast 2-hybrid screens. The phenotype induced by loss of expression of CD36 homologs in C. elegans will be characterized to identify the signaling pathways utilized by the homologs in engulfment of apoptotic cells and cell migration. Once candidate signaling partners are identified, their role in CD36-mediated signal transduction will be studied in vascular cells. Practical attention will be paid to changes in protein activity and localization induced by ligation of CD36 by its ligands and to their effects on angiogenesis and macrophage function. This project involves many collaborations and synergisms with investigators in other projects within the Program. Strategies to identify candidate signaling proteins by peptide sequencing will involve collaboration with Drs. Gross, K. Kajjar, and R. Upmacis (projects 5, 1 and 6), who are experts in MS/MS and protein chemistry. Studies of cytoskeletal regulation and signaling by Crk, DOCK180 and Rac will involve collaboration with Dr. Hempstead (project 4), and studies of G- protein signaling will involve collaboration with Dr. D. Hajjar. Studies of endothelial cell responses will involve collaboration with Drs. Kraemer and Hempstead (project 4).
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