Adult epidermal keratinocytes migrate by crawling, a process that requires protrusion of the plasma membrane at the front of the cell and contraction of the cell body at the rear. We have found that epidermal growth factor (EGF) and insulin-like growth factor 1 (IGF-1) influence keratinocyte shape differently. Whereas IGF-1 stimulates membrane protrusion and facilitates cell spreading, EGF induces contraction of keratinocytes. The effects of each growth factor on keratinocyte shape are mediated by distinct signal transduction pathways: EGF stimulates the activity of the classical mitogen-activated protein kinase pathway and IGF-1 stimulates phosphatidylinositol-3-kinase. Activation of these kinases is both necessary and sufficient to induce cell shape changes upon growth factor treatment. In addition, IGF-1-stimulated keratinocyte spreading depends on the activation of Rho family proteins. In vitro assays of wound re-epithelialization show that both growth factors stimulate migration of keratinocytes, and the activity of the respective signalling pathways is required for this re-epithelialization process. When added simultaneously, IGF-1 and EGF have additive effects on wound epithelialization. Our results show that IGF-1 and EGF can influence different components of the keratinocyte migration machinery that determines the speed of wound epithelialization.