Section 7
Chapter 6,104

Phorbol myristate acetate (PMA) augments chemoattractant-induced diglyceride generation in human neutrophils but inhibits phosphoinositide hydrolysis. Implications for the mechanism of PMA priming of the respiratory burst

Tyagi, S.R.; Tamura, M.; Burnham, D.N.; Lambeth, J.D.

Journal of Biological Chemistry 263(26): 13191-13198


ISSN/ISBN: 0021-9258
PMID: 2843513
Accession: 006103779

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Pretreatment ("priming") of neutrophils with a non-activating concentration (2 nM) of phorbol myristate acetate (PMA) augments superoxide (O2-) production in response to the chemoattractant formylmethionylleucylphenylalanine (fMLP). We initially examined the effect of sphinganine, an inhibitor of protein kinase C (Ca2+/phospholipid-dependent enzyme), on activation of primed neutrophils. In both primed and unprimed cells activation by fMLP was blocked, and inhibition occurred at identical concentrations, supporting a common inhibited site. PMA also augmented (about 2-fold) fMLP-induced generation of sn-1,2-diglyceride (DG), the level of which correlated with O2- generation. In contrast to its effects on DG, PMA diminished by about 50% the magnitude of the fMLP-stimulated rise in cytosolic Ca2+. Thus, PMA priming dissociates the fMLP-stimulated Ca2+ increase from DG and O2- generation. The effect of PMA on Ca2+ levels appeared to be due in part to lowered levels of inositol trisphosphate. Lowering of inositol phosphate levels correlated with inhibition of fMLP-induced hydrolysis of inositol-containing phospholipids, particularly phosphatidylinositol 4,5-bisphosphate. PMA did not inhibit (and in fact augmented at early time points) formation of [32P] phosphatidic acid in response to fMLP, indicating that the increase in DG was not due to inhibition of cellular diglyceride kinase. Thus, the data suggest that PMA enhances fMLP-stimulated DG generation concomitant with switching the source of DG from phosphatidylinositol 4,5-bisphosphate to an alternative lipid(s). Increased DG and inhibition of activation by sphinganine are consistent with a role for protein kinase C in activation of the respiratory burst in PMA-primed neutrophils.

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