Ezrin is a downstream effector of trafficking PKC-integrin complexes involved in the control of cell motility

Protein kinase C (PKC) a has been implicated in beta1 integrin-mediated cell migration. Stable expression of PKC alpha is shown here to enhance wound closure. This PKC-driven migratory response directly correlates with increased C-terminal threonine phosphorylation of ezrin/moesin/radixin (ERM) at the wound edge. Both the wound migratory response and ERM phosphorylation are dependent upon the catalytic function of PKC and are susceptible to inhibition by phosphatidylinositol 3-kinase blockade. Upon phorbol 12,13-dibutyrate stimulation, green fluorescent protein-PKC alpha and beta1 integrins co-sediment with ERM proteins in low-density sucrose gradient fractions that are enriched in transferrin receptors, Using fluorescence lifetime imaging microscopy, PKC alpha is shown to form a molecular complex with ezrin, and the PKC-co-precipitated endogenous ERR I is hyperphosphorylated at the C-terminal threonine residue, i.e, activated. Electron microscopy showed an enrichment of both proteins in plasma membrane protrusions. Finally, overexpression of the C-terminal threonine phosphorylation site mutant of ezrin has a dominant inhibitory effect on PKC alpha -induced cell migration. We provide the first evidence that PKC alpha or a PKC alpha -associated serine/threonine kinase can phosphorylate the ERM C-terminal threonine residue within a kinase-ezrin molecular complex in vivo.