EGF is known to affect adherens junctions and disrupt cell-cell adhesion in a variety of carcinomas but the underlying mechanisms are not completely understood. Using human tumor epithelial cells overexpressing EGFR we demonstrated that EGF-induced cell scattering was mediated by protein kinase C-delta (PKC-d). PKC-d knockdown by siRNA significantly inhibited EGF-induced internalization of E-cadherin into the cytoplasm and blocked cell scattering. EGF phosphorylated PKC-d at Y311 and ectopic expression of the mutant Y311F prevented PKC-dbinding to E-cadherin and EGF-induced cell-scattering. Moreover, blocking src or p38, but not p42/44 or JNK decreased EGF-induced PKC-d phosphorylation at Y311, consequently blocking its binding to E-cadherin and cell-scattering. Finally, EGF reduced expression of the tight junction protein, occludin, and this effect was also mediated by PKC-d. In summary, PKC-d mediated the effects of EGF on adherens and tight junctions thereby playing an important role in cell-cell adhesion with possible wider implications in tumor metastasis or epithelial-to-mesenchymal transition.