REGULATION OF FOCAL ADHESION STRUCTURE

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The long-term goal of this work is to understand how cell adhesion to the extracellular matrix is regulated. We will study the protein interactions in the focal adhesions, which are the sites where cells adhere to the extracellular matrix and where the cytoskeleton is anchored to the membrane. Cell adhesion plays an important role during embryonic development, when cells are stimulated either to migrate or adhere based on signals conveyed from the extracellular matrix. Cell adhesion is also a major factor in human disease: improper regulation of adhesion in blood platelets can generate arterial blood clots that cause strokes and heart attacks, and the loss of adhesion that typically occurs in cancer cells contributes to tumor metastasis. Cell adhesions involves three groups of proteins: proteins outside the cell that form the extracellular matrix, transmembrane proteins that bind to the extracellular matrix (the integrins), and cytoskeleton proteins that anchor the cytoskeleton to the integrins. This project is focused on the integrins and their linkage to the cytoskeleton. The specific aims of this proposal are to characterize in detail the interactions between the cytoplasmic tails of the alpha and beta subunits of intergrin with the cytoskeleton. Synthetic peptides will be used to represent the integrin cytoplasmic tails. These peptides will be used in co-precipitation assays, solid-phase binding assays, and affinity chromatography experiments to identify cytoskeletal proteins that bind to the integrin cytoplasmic domains. The data obtained from these experiments will be used to generate models representing potential linkages between integrins and the cytoskeleton. These models will be tested in living cell using the techniques of immunoprecipitation and microinjection.