Platelet secretion induced by phorbol esters stimulation is mediated through phosphorylation of MARCKS: a MARCKS-derived peptide blocks MARCKS phosphorylation and serotonin release without affecting pleckstrin phosphorylation.
Previous experiments suggest that actin disassembly, perhaps at a specific site, is required for platelet secretion. Platelet stimulation by phorbol 12-myristate 13-acetate (PMA) induced pleckstrin phosphorylation, platelet aggregation, and secretion. Inhibition of protein kinase C (PKC) is accompanied by inhibition of pleckstrin phosphorylation and serotonin secretion. Here, we demonstrate the presence of myristoylated alanine-rich C kinase substrate (MARCKS), another PKC substrate, in platelets and its phosphorylation during PMA stimulation. MARCKS is known to bind actin and to cross-link actin filaments; the latter is inhibited by PKC-induced MARCKS phosphorylation. MARCKS phosphorylation and serotonin release from permeabilized platelets have the same time course and were blocked by a peptide (MPSD) with the amino acid sequence corresponding to the phosphorylation site domain of MARCKS. Pleckstrin and myosin light chain phosphorylation was not modified. A peptide (Ala-MPSD) in which the four serine residues of MPSD were substituted by alanines was ineffective. These results provide the first evidence that MARCKS may play a role in platelet secretion. Moreover, pleckstrin phosphorylation has a different time course than that of MARCKS or serotonin release and was not modified when MARCKS phosphorylation and serotonin release were inhibited, suggesting that pleckstrin is either not directly involved in secretion or that it might only be involved upstream in the cascade of events leading to exocytosis.
Elzagallaai
A
A
Secretory Process Research Program, Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.
Rosé
S D
SD
Trifaró
J M
JM
D000595:Amino Acid Sequence; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D001798:Blood Proteins / Q000378:metabolism; D004789:Enzyme Activation; D005089:Exocytosis / Q000187:drug effects; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D018994:Myosin Light Chains / Q000378:metabolism; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010446:Peptide Fragments / Q000494:pharmacology*; D010750:Phosphoproteins; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000502:physiology*; D012701:Serotonin / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D001798:Blood Proteins; D047908:Intracellular Signaling Peptides and Proteins; C000619710:MARCKS protein, human; D008565:Membrane Proteins; D018994:Myosin Light Chains; D010446:Peptide Fragments; D010750:Phosphoproteins; D011506:Proteins; C039077:platelet protein P47; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D012701:Serotonin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate