To evaluate the mechanisms by which epidermal growth factor (EGF) regulates actin-based cellular processes such as cell migration, we first examined the effects of EGF on cell adhesion, which is essential for cell migration. In mouse B82L fibroblasts transfected with the full-length EGF receptor, EGF promotes cell rounding and attenuates cell spreading on fibronectin, laminin, and vitronectin, and thus appears to reduce the strength of cell adhesion. Moreover, EGF synergizes with multiple extracellular matrix (ECM) components in the promotion of integrin-mediated cell migration of several different cell types, including fibroblasts and various carcinoma and osteosarcoma cell lines. Interestingly, co-presentation (co-positioning) of EGF with laminin or fibronectin is essential for EGF-stimulated migration. When EGF is mixed with the cells instead of the ECM components, it has little effect on cell migration. These results suggest that co-presentation of EGF with ECM components can enhance the polarization events required for directional cell movement. To identify the EGF receptor elements critical for the EGF stimulation of cell migration, B82L fibroblasts were transfected with either mutated or wild-type EGF receptors. Surprisingly, we found that B82L-Parental cells that lack the EGF receptor are not able to migrate to fibronectin, even though they can adhere to fibronectin. However, the introduction of wild-type EGF receptors into these fibroblasts enables them to migrate toward fibronectin even in the absence of EGF. The requirement of the EGF receptor for cell migration does not appear to result from the secretion of EGF or TGF-␣ by the cells transfected with the EGF receptor. Furthermore, cells expressing EGF receptors that are kinase-inactive, or Cterminally truncated, exhibit little migration toward fibronectin, indicating that an intact EGF receptor kinase is required for fibronectin-induced cell migration. In addition, neutralizing anti-EGF receptor antibodies attenuate cell migration in the presence of EGF, and inhibit migration to fibronectin or laminin alone. These results further suggest that the EGF receptor is downstream of integrin activation in the signal transduction pathways leading to fibroblast migration. EGF1 triggers many biological responses, including cell proliferation and differentiation (1). In addition, EGF has been shown to induce the reorganization of the actin cytoskeleton, and the EGF receptor has been found to be associated with actin filaments (2-7). In this regard, EGF has been reported to stimulate rapid cell rounding, extensive membrane ruffling, extension of filopodia, retraction of cells from the substratum (8, 9), extensive cortical actin polymerization, and depolymerization of actin stress fibers (10, 11). Moreover, numerous studies have shown that activation of the EGF receptor leads to increased cell motility (12-19) and production of ECM degrading proteases (20 -23), thereby supporting a role for the EGF receptor in normal development and pathophysiological events such...