Hyperosmolality induces activation of cPKC and nPKC, a requirement for ERK1/2 activation in NIH/3T3 cells.
Protein kinase C (PKC) has been reported to be associated with the activation of extracellular signal-regulated kinase (ERK) by hyperosmolality. However, it is unclear whether hyperosmolality induces PKC activation and which PKC isoforms are involved in ERK activation. In this study, we demonstrate that NaCl increases total PKC activity and induces PKCalpha, PKCdelta, and PKCepsilon translocation from the cytosol to the membrane in NIH/3T3 cells, suggesting that hyperosmotic stress activates conventional PKC (cPKC) and novel PKC (nPKC). Further studies show that NaCl-inducible ERK1 and ERK2 (ERK1/2) activation is a consequence of cPKC and nPKC activation, because either downregulation with 12-O-tetradecanoylphorbol 13-acetate or selective inhibition of cPKC and nPKC by GF-109203X and rottlerin largely inhibited the stimulation of ERK1/2 phosphorylation by NaCl. In addition, we show that NaCl increases diacylglycerol (DAG) levels and that a phospholipase C (PLC) inhibitor, U-73122, inhibits NaCl-induced ERK1/2 phosphorylation. These results, together, suggest that a hyperosmotic NaCl-induced signaling pathway that leads to activation of ERK1/2 may sequentially involve PLC activation, DAG release, and cPKC and nPKC activation.
Zhuang
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Department of Molecular Biology, Yokohama City University School of Medicine, Kanazawa-Ku, Yokohama 236, Japan.
Hirai
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Ohno
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D016475:3T3 Cells / Q000201:enzymology; D000818:Animals; D002867:Chromones / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D005110:Extracellular Space / Q000378:metabolism; D006982:Hypertonic Solutions / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009025:Morpholines / Q000494:pharmacology; D009994:Osmolar Concentration; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D011760:Pyrrolidinones / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*; D014882:Water-Electrolyte Balance / Q000502:physiology*
D002867:Chromones; D004075:Diglycerides; D004791:Enzyme Inhibitors; D004963:Estrenes; D006982:Hypertonic Solutions; D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D009025:Morpholines; D010726:Phosphodiesterase Inhibitors; D011760:Pyrrolidinones; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D012965:Sodium Chloride; D019869:Phosphatidylinositol 3-Kinases; C497259:Prkcd protein, mouse; C497256:Prkce protein, mouse; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D010738:Type C Phospholipases; C070515:bisindolylmaleimide I