The mammary adenocarcinoma cell line MTLn3 is chemotactic towards epidermal growth factor (EGF), and this induced motility is thought to promote breast cancer invasion and metastasis. Stimulation of MTLn3 cells with EGF results in the extension of a flat, thin structure filled with filamentous actin and termed a lamellipod. Lamellipod extension is dependent on actin polymerization and is localized to the border of adherent cells. The structure of EGF-stimulated lamellipods in MTLn3 cells is well suited to analysis of chemoattractant-stimulated protrusion. Actin polymerization occurs within 200 nm of the extending edge of the lamellipod. Although extension of the lamellipod is not dependent upon interaction with the substratum, stabilization of the extended lamellipod is dependent on an adhesive substratum. Dorsal ruffling is suppressed during lamellipod extension. Tyrosine phosphorylation is reduced in preexisting focal contacts compared to new contacts induced by EGF stimulation. The coordination of turnover of focal contacts with lamellipod extension is proposed to result in polarized cell motility in response to gradients of chemoattractants.