Antigen activation of mitogen-activated protein kinase in mast cells through protein kinase C-dependent and independent pathways.
We demonstrate discrete pathways for activation of mitogen-activated protein (MAP) kinase in cultured RBL-2H3 mast cells through protein kinase C (PKC), cytosolic calcium, and a third pathway that provides sustained signals for activation in Ag-stimulated cells. Thus, p42 MAP kinase was activated by increasing intracellular free Ca2+ with thapsigargin or by stimulating PKC with PMA. The latter stimulation was selectively blocked by the protein kinase C inhibitor, Ro31-7549. Stimulation of p42 MAP kinase by Ag resulted in relatively sustained activation of MAP kinase which was only partially suppressed by Ro31-7549. Kinetic studies revealed two components of the MAP kinase response to Ag: a rapid but transient component that was Ro31-7549 sensitive and presumably PKC dependent; and a more sustained component that was Ro31-7549 resistant and presumably PKC independent. Similarly, Ro31-7549 inhibited the early but not late release of arachidonic acid, a finding that was consistent with the known regulation of phospholipase A2 by MAP kinase. Early tyrosine phosphorylation events which were thought to be essential for Ag-induced activation of p42 MAP kinase and release of arachidonic acid were unaffected by Ro31-7549. The findings suggested that release of arachidonic acid was regulated primarily through MAP kinase but that PKC may transiently influence this release, either directly or indirectly through MAP kinase.
Zhang
C
C
Laboratory of Molecular Immunology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Hirasawa
N
N
Beaven
M A
MA
D000818:Animals; D000941:Antigens / Q000276:immunology; D016718:Arachidonic Acid / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000502:physiology; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008301:Maleimides / Q000494:pharmacology; D008407:Mast Cells / Q000201:enzymology*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D019000:Phosphotyrosine / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051567:Proto-Oncogene Proteins c-vav; D051381:Rats; D000072377:Syk Kinase; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology
D000941:Antigens; D018797:Cell Cycle Proteins; D004792:Enzyme Precursors; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D008301:Maleimides; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; C496620:Vav1 protein, rat; D019000:Phosphotyrosine; D016718:Arachidonic Acid; D019284:Thapsigargin; D002217:Carbachol; D011505:Protein-Tyrosine Kinases; D000072377:Syk Kinase; C000606898:Syk protein, rat; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D019204:GTP-Binding Proteins; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; C064755:Ro 31-7549