Protein Kinase C ζ Is Required for Epidermal Growth Factor–Induced Chemotaxis of Human Breast Cancer Cells

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Abstract
Chemotaxis plays an important role in cancer cell metastasis. In this study, we showed that epidermal growth factor (EGF) was a more potent chemoattractant than chemokine SDF-1α/CXCL12 for human breast cancer cell MDA-MB-231. Different inhibitors were used to evaluate the involvement of 12 protein kinase C (PKC) isotypes in the chemotactic signaling pathway. Chelerythrine chloride, an inhibitor of all PKC isotypes, blocked chemotaxis, whereas inhibitors of classic and novel PKC, such as Gö6976, Gö6850, or calphostin C, only impaired EGF-induced chemotaxis to a minor extent by ≯32% inhibition. These data suggested that atypical PKC were involved. The ligand-induced actin polymerization and cell adhesion were also similarly dependent on atypical PKC. Immunofluorescent staining showed an EGF-induced, LY294002-sensitive translocation of PKCζ from the cytosol to the plasma membrane, indicating that EGF was capable of activating PKCζ, probably via phosphoinositide 3 kinases. A myristoylated PKCζ pseudosubstrate blocked the chemotaxis with an IC50 of 20 μmol/L. To expand our investigation, we further showed that in MCF-7 and T47D, two additional human breast cancer cell lines, EGF-activated PKCζ and the PKCζ pseudosubstrate, inhibited chemotaxis. Taken together, our data suggest that PKCζ is an essential component of the EGF-stimulated chemotactic signaling pathway in human breast cancer cells.