Mechanism of inhibition of sequestration of protein kinase C alpha/betaII by ceramide. Roles of ceramide-activated protein phosphatases and phosphorylation/dephosphorylation of protein kinase C alpha/betaII on threonine 638/641.

摘要: Sustained activation of protein kinase C (PKC) isoenzymes alpha and betaII leads to their translocation a perinuclear region the formation pericentrion, PKC-dependent subset recycling endosomes. In MCF-7 human breast cancer cells, action PKC activator 4beta-phorbol-12-myristate-13-acetate (PMA) evokes ceramide formation, which in turn prevents PKCalpha/betaII pericentrion. this study we investigated mechanisms by negatively regulates PKCalpha/betaII. Upon PMA treatment, HEK-293 cells displayed dual phosphorylation at carboxyl-terminal sites (Thr-638/641 Ser-657/660), whereas were phosphorylated Ser-657/660 but not Thr-638/641. Inhibition synthesis fumonisin B1 overcame defect restored To determine involvement ceramide-activated phosphatases regulation, employed small interference RNA silence individual Ser/Thr phosphatases. Knockdown isoforms or beta catalytic subunits phosphatase 1 only increased Thr-638/641 also PKCbetaII Mutagenesis approaches revealed that mutation either Thr-641 Ser-660 Ala abolished sequestration PKC, implying indispensable roles those for Reciprocally, point Glu, mimics phosphorylation, inhibitory effects on PMA-stimulated cells. Therefore, results demonstrate novel role they disclose specific regulation autophosphorylation through 1.