Phosphorylation of MARCKS (80-kDa) protein, a major substrate for protein kinase C in oligodendroglial progenitors.
We have recently reported a potent mitogenic stimulation of oligodendroglial (OL) progenitors by the protein kinase C (PKC) activating phorbol ester, i.e., phorbol 12-myristate 13-acetate (PMA) (Bhat NR, J Neurosci Res 22:20-27, 1989). The present study deals with PMA-induced protein phosphorylation reactions in cultured OL progenitors. The phorbol ester induced the phosphorylation of several cytosol and membrane-associated proteins, including a major protein with an apparent molecular weight of 80 kDa. In both control and PMA-treated cultures, phosphorylation level of the 80-kDa protein in cytosol was higher than that in the particulate fraction. Okadaic acid, an inhibitor of protein phosphatases, also increased the phosphorylation of several proteins and substantially enhanced protein phosphorylation induced by PMA. In vitro incubation of the cell membranes with phosphatidylserine and diacylglycerol (a physiological activator of PKC) in the presence of [gamma 32p]-ATP resulted in an increased phosphorylation of the 80-kDa protein. The induction of phosphorylation of the 80-kDa protein under both in situ and in vitro conditions was subject to inhibition by 1-[5[isoquinolinyl sulfonyl)-3-methylpiperazine (H-7), a potent inhibitor of PKC. The 80-kDa phosphoprotein was identified as the prominent PKC substrate, i.e., myristoylated alanine-rich C-kinase substrate (MARCKS) protein by immunoprecipitation with anti-MARCKS antibodies.
Bhat
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NR
Department of Biochemistry, University of Kentucky College of Medicine, Lexington.
D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D015151:Immunoblotting; D007120:Immunochemistry; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D009836:Oligodendroglia / Q000201:enzymology*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D013234:Stem Cells / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D047908:Intracellular Signaling Peptides and Proteins; C000619712:Marcks protein, rat; D008565:Membrane Proteins; D010761:Phosphorus Radioisotopes; D011506:Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D004247:DNA; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate