Protein phosphatase 1α mediates ceramide-induced ERM protein dephosphorylation: a novel mechanism independent of phosphatidylinositol 4, 5-biphosphate (PIP2) and myosin/ERM phosphatase.
ERM (ezrin, radixin, and moesin) proteins are cytoskeletal interacting proteins that bind cortical actin, the plasma membrane, and membrane proteins, which are found in specialized plasma membrane structures such as microvilli and filopodia. ERM proteins are regulated by phosphatidylinositol 4, 5-biphosphate (PIP(2)) and by phosphorylation of a C-terminal threonine, and its inactivation involves PIP(2) hydrolysis and/or myosin phosphatase (MP). Recently, we demonstrated that ERM proteins are also subject to counter regulation by the bioactive sphingolipids ceramide and sphingosine 1-phosphate. Plasma membrane ceramide induces ERM dephosphorylation whereas sphingosine 1-phosphate induces their phosphorylation. In this work, we pursue the mechanisms by which ceramide regulates dephosphorylation. We found that this dephosphorylation was independent of hydrolysis and localization of PIP(2) and MP. However, the results show that ERM dephosphorylation was blocked by treatment with protein phosphatase 1 (PP1) pharmacological inhibitors and specifically by siRNA to PP1α, whereas okadaic acid, a PP2A inhibitor, failed. Moreover, a catalytic inactive mutant of PP1α acted as dominant negative of the endogenous PP1α. Additional results showed that the ceramide mechanism of PP1α activation is largely independent of PIP(2) hydrolysis and MP. Taken together, these results demonstrate a novel, acute mechanism of ERM regulation dependent on PP1α and plasma membrane ceramide.
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29425.
Daniel
D
Canals
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29425.
Patrick
P
Roddy
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29425. Electronic address: hannun@musc.edu.
Yusuf A
YA
Hannun
D002518:Ceramides / Q000235:genetics / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D008246:Lysophospholipids / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D019319:Okadaic Acid / Q000494:pharmacology; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects / Q000235:genetics; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives / Q000235:genetics / Q000378:metabolism
D002518:Ceramides; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D019269:Phosphatidylinositol 4,5-Diphosphate; D019319:Okadaic Acid; C060506:sphingosine 1-phosphate; C517189:PPP2CA protein, human; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2; D013110:Sphingosine