ABSTRACT
This chapter describes experimental protocols for studies on tyrosine phosphorylation of Cas in response to integrin-mediated cell adhesion and for detection of tyrosine phosphorylation-dependent interactions mediated by Cas. It focuses on defining the functional significance of the Cas-Crk complex formation in intracellular signaling. Cas has been found to become tyrosine-phosphorylated following cell adhesion to ECM substrates, but not to polylysine, which is consistent with the phosphorylation being mediated by integrins. The time course of Cas tyrosine phosphorylation in response to integrin ligand binding is relatively slow and persistent; maximal phosphorylation in fibroblasts occurs at around 15 min after plating, which coincides with cell spreading and actin filament reorganization, and the phosphorylation levels are maintained with a slow decline for the duration of cell adhesion.
