Growth factors, integrins, and the extracellular matrix (ECM) are known to play key roles in epidermal wound healing, although the interplay between these proteins is not fully understood. We show that growth factor macrophage stimulating protein (MSP)- and its receptor Ron-mediated PI3K activation in keratinocytes induces phosphorylation of both Ron and α6β4 integrin at specific 14-3-3 binding sites. Consequently, a Ron/α6β4 complex formed via 14-3-3 binding displaces α6β4 from its location at hemidesmosomes (structures supporting cell adhesion) and relocalizes it to lamellipodia. Concomitant activation of α3β1 and keratinocyte spreading/migration on laminin-5 occurs. Further, MSP-dependent β4 tyrosine phosphorylation evokes p38 and NF-κB signaling required for keratinocyte wound closure. Based on these results, we propose a mechanism based on MSP-Ron-dependent phosphorylation and 14-3-3 association, whereby the function of α6β4 switches from a mechanical adhesive device into a signaling component, and might be critically involved in human epidermal wound healing.