Activation of protein kinase C down-regulates leukotriene C4 synthase activity and attenuates cysteinyl leukotriene production in an eosinophilic substrain of HL-60 cells.
An eosinophilic substrain of HL-60 cells (HL-60#7) predominantly synthesized cysteinyl leukotrienes after stimulation with the calcium ionophore A23187. Activation of protein kinase C (PKC) by phorbol 12-myristate 13-acetate (PMA) specifically attenuated cysteinyl leukotriene production without affecting the biosynthesis of non-cysteinyl leukotrienes. The inhibition of cysteinyl leukotriene biosynthesis was prevented only by specific PKC inhibitors (staurosporine and bisindolylmaleimide) but not by inhibitors of tyrosine kinases (genistein, tyrphostin 47, and herbimycin A), protein kinase A (KT5720), or the oxidative burst (apocynin). Similar results were obtained when LTC4 synthase enzymatic activity was measured directly in the presence of saturating concentrations of exogenously added substrates. Therefore, the inhibitory effects of PKC activation on cysteinyl leukotriene formation in intact cells was attributable to effects on the LTC4 synthase enzyme. The mechanism of inhibition of LTC4 synthase by PKC activation was determined by kinetic analysis to be noncompetitive in both eosinophil-like HL-60#7 cells and monocytic THP-1 cells. Contrary to the effect of PKC activation on cysteinyl leukotriene biosynthesis, the formation of prostaglandin E2 and thromboxane B2 was elevated twofold to threefold after PMA treatment, which was prevented by the PKC inhibitor, staurosporine. We propose a regulatory model in which PKC activation shifts the profile of eicosanoid mediators produced by eosinophils from cysteinyl leukotrienes to prostanoids.
Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.
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Ali
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Ford-Hutchinson
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Nicholson
D000470:Alkaloids / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D003545:Cysteine; D015536:Down-Regulation; D004804:Eosinophils / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism*; D015289:Leukotrienes / Q000096:biosynthesis*; D009570:Nitriles / Q000494:pharmacology; D010636:Phenols / Q000494:pharmacology; D011453:Prostaglandins / Q000096:biosynthesis; D011493:Protein Kinase C / Q000502:physiology*; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D020032:Tyrphostins
D000470:Alkaloids; D015289:Leukotrienes; D009570:Nitriles; D010636:Phenols; D011453:Prostaglandins; D020032:Tyrphostins; C060152:tyrphostin 47; D000001:Calcimycin; D005982:Glutathione Transferase; D011493:Protein Kinase C; C041116:leukotriene-C4 synthase; D019311:Staurosporine; D003545:Cysteine; D013755:Tetradecanoylphorbol Acetate