Protein kinase C-alpha coordinately regulates cytosolic phospholipase A(2) activity and the expression of cyclooxygenase-2 through different mechanisms in mouse keratinocytes.
Transgenic mice (K5-PKC alpha) in which the keratin 5 promoter directs the expression of protein kinase C-alpha (PKC alpha) to epidermal keratinocytes display a 10-fold increase in PKC alpha protein in their epidermis and alterations in phorbol ester-induced cutaneous inflammation [J Cell Science 1999;112:3497-3506]. In the current study, we have used these K5-PKC alpha mice to examine the role of PKC alpha in keratinocyte phospholipid metabolism/eicosanoid production and cutaneous inflammation. Primary keratinocytes from wild-type and transgenic mice were prelabeled in culture with [(3)H]arachidonic acid (AA) and subsequently treated with TPA. Compared with wild-type keratinocytes, K5-PKC alpha keratinocytes displayed a 2-fold increase in AA release. TPA treatment resulted in the phosphorylation of cPLA(2). PKC inhibitors GF-109203X or H7, but not mitogen-activated protein/extracellular signal-regulated protein kinase (MEK) inhibitor PD 98059, could inhibit phosphorylation and AA release. Topical 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment of K5-PKC alpha mice resulted in a 5-fold increase in epidermal COX-2 induction and a 2- to 3-fold increase in prostaglandin (PG) E(2) levels above that observed in TPA-treated wild-type mice. PD 98059, GF-109203X, or H7 could block cyclooxygenase-2 (COX-2) induction by TPA. Because C/EBP beta, a basic leucine zipper transcription factor, can be activated via a PKC alpha/mitogen-activated protein kinase pathway and can influence COX-2 expression, we examined whether C/EBP beta is involved in TPA-induced epidermal COX-2 expression. TPA-induced COX-2 expression was similar in C/EBP beta nullizygous and wild-type mice. In summary, our results indicate that epidermal PKC alpha coordinately regulates cPLA(2) activity and COX-2 expression resulting in increased levels of AA and PGE(2). Furthermore, PKC alpha-induced AA release and cPLA(2) phosphorylation are independent of MEK, whereas PKC alpha-induced COX-2 expression and PGE(2) production are MEK-dependent and C/EBP beta-independent events.
Cell Signaling and Cancer Group, Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, North Carolina 27695-7633, USA.
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Wang
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Kim
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Tiano
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Langenbach
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Smart
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism; D002478:Cells, Cultured; D051546:Cyclooxygenase 2; D003600:Cytosol / Q000201:enzymology*; D015232:Dinoprostone / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007249:Inflammation / Q000378:metabolism; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism*; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007633:Keratins / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors; D010741:Phospholipases A / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D020033:Protein Isoforms / Q000235:genetics; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D012867:Skin / Q000187:drug effects / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D022782:CCAAT-Enhancer-Binding Protein-beta; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010743:Phospholipids; D020033:Protein Isoforms; D016718:Arachidonic Acid; D007633:Keratins; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D020929:Mitogen-Activated Protein Kinase Kinases; D010741:Phospholipases A; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate