The long term goals of this revised project are to understand the nature of cellular signalling events mediated by cellular receptors that recognize components of the extracellular matrix and/or other cell surface molecules. We will focus specifically on the role of tyrosine phosphorylation in regulating these pathways, seeking to define the molecular mechanisms by which tyrosine kinases contribute to the regulation and control of such pathways. The proposed experiments build and extend the progress made during the past four years. We have identified a novel protein tyrosine kinase, that is associated with cellular focal adhesions, designated Focal Adhesion Kinase, FAK. Evidence from our own laboratory as well as other has indicated that FAK plays a role in regulating signalling events initiated by interactions of surface integrins with extracellular matrix. In addition, our own studies have demonstrated a stable association of FAK with pp60src in src transformed cells. We outline three specific aims: First, we will define and characterize the mechanisms that lead to activation of FAK in response to engagement of integrins with defined extracellular ligands. These studies will include an analysis of the interaction of FAK with cytoplasmic domains of specific integrins and components of focal adhesions as well as possible functional interactions with proteins that regulate mitogen and hormone activated signal transduction pathways. Second, we will examine the role of FAK in the regulation of the formation and/or breakdown of cellular focal adhesions, in the regulation of other cellular activities (e.g., adhesion, cell spreading, cell migration) and the possible control of second messenger pathways. In these studies we will investigate the phenotypic and biochemical properties of cells expressing variant FAK proteins and attempt to correlate known defects in FAK activity with alterations in cellular metabolism. Finally, we will characterize the function interactions between FAK and pp60src in src transformed cells and extend this paradigm to the analysis of possible interactions of FAK and pp60src or other src family kinases in normal cells. These experiments seek to delineate the role of pp60src in the structural perturbation of focal adhesions in transformed cells and explore the possibility that FAK regulates or is regulated by src family kinases in normal cells.
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