RNAi-mediated knockdown of protein kinase C-alpha inhibits cell migration in MM-RU human metastatic melanoma cell line

摘要

Protein kinase C (PKC) is a multigene family of serine/threonine protein kinases involved in cell signaling pathways of proliferation and motility. PKC interacts with Rho GTPases in the regulation of the actin cytoskeleton. The PKC-α isozyme binds the Rho GTPase cdc42, and both are coordinated with the Rac-phosphatidylinositol-3 kinase (PI3K) signaling pathway in melanoma cell invasion and migration on extracellular matrix proteins. To further define the role of PKC-α in melanoma cell migration, we tested the effect of PDBu and Ca2+ dependent activation of PKC-α as well as treatment with the PKC-α inhibitors calphostin C and Go6976. Furthermore, we transfected siRNA targeted against PKC-α into human melanoma cells and performed time-lapse analysis of cell migration followed by western immunoblotting. We found that significant enhancement of cell migration at 0.5 h after PDBu treatment directly correlated with Ca2+ dependent activation of PKC-α and was inhibited by the PKC-α inhibitor calphostin C. PKC-α siRNA transfection nearly abrogated PKC-α expression and significantly reduced melanoma cell migration compared with siRNA controls. These findings provide further evidence that PKC-α plays an important role in melanoma cell migration and may have implications in therapies designed to disrupt melanoma cell motility by alteration of PKC-α signaling.