Abstract
Cytosolic phospholipase A(2) (cPLA(2)) is an enzyme involved in the formation of proinflammatory mediators by catalyzing the release of arachidonic acid, thereby mediating eicosanoid biosynthesis. Using HaCaT keratinocytes as a model system, we present experimental evidence that in these cells, cPLA(2) is constitutively phosphorylated and that the degree of phosphorylation dramatically increases in cells under hyperosmotic stress induced by sorbitol. In parallel, a rapid release of arachidonic acid followed by prostaglandin E(2) formation was detected. Elucidating the mechanism of cPLA(2) upregulation, we observed that it is mediated via epidermal growth factor receptor (EGFR) activation, since tyrphostin AG1478, a selective inhibitor of EGFR tyrosine kinase, completely inhibited cPLA(2) phosphorylation. Furthermore, addition of PD98059, which is an inhibitor of MEK1 activation, but not of SB203580, which is an inhibitor of p38 stress kinase, inhibited cPLA(2) phosphorylation, indicating that the ras-raf-MEK cascade is the major signalling pathway involved in cPLA(2) phosphorylation. In addition, depletion of the cells from intracellular calcium does not prevent sorbitol-elicited cPLA(2) phosphorylation, suggesting that this process is independent of the presence of calcium. Together, our results demonstrate that hyperosmotic stress phosphorylates cPLA(2) in human keratinocytes by an EGFR-mediated process.
Copyright 2002 Elsevier Science Inc.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Arachidonic Acid / metabolism
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Calcium / deficiency
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Cell Compartmentation / drug effects
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Cell Compartmentation / physiology
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Cell Line, Transformed
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Cell Nucleus / drug effects
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Cell Nucleus / enzymology
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Dinoprostone / metabolism
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Drug Eruptions / enzymology*
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Drug Eruptions / physiopathology
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Enzyme Inhibitors / pharmacology
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Epidermal Growth Factor / metabolism
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ErbB Receptors / drug effects
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ErbB Receptors / metabolism*
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Humans
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Keratinocytes / drug effects*
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Keratinocytes / enzymology*
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MAP Kinase Kinase 1
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MAP Kinase Signaling System / drug effects
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MAP Kinase Signaling System / physiology
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Mitogen-Activated Protein Kinase Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinase Kinases / metabolism
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Mitogen-Activated Protein Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinases / drug effects
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Mitogen-Activated Protein Kinases / metabolism
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Osmotic Pressure / drug effects*
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Phospholipases A / drug effects
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Phospholipases A / metabolism*
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Phospholipids / metabolism*
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Phosphorylation / drug effects
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Phosphotransferases / antagonists & inhibitors
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Phosphotransferases / metabolism
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Protein-Serine-Threonine Kinases / antagonists & inhibitors
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Protein-Serine-Threonine Kinases / metabolism
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Sorbitol / toxicity
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Stress, Physiological / chemically induced
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Stress, Physiological / enzymology*
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Stress, Physiological / physiopathology
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Tyrosine / metabolism
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p38 Mitogen-Activated Protein Kinases
Substances
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Enzyme Inhibitors
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Phospholipids
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Arachidonic Acid
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Tyrosine
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Sorbitol
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Epidermal Growth Factor
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Phosphotransferases
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ErbB Receptors
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Protein-Serine-Threonine Kinases
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Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 1
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MAP2K1 protein, human
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Mitogen-Activated Protein Kinase Kinases
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Phospholipases A
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Dinoprostone
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Calcium