Mitogen-activated protein kinase activation during IgG-dependent phagocytosis in human neutrophils: inhibition by ceramide.
In FMLP-activated polymorphonuclear leukocytes (PMNs) challenged with IgG-opsonized erythrocytes (EIgG), the termination of phagocytosis correlates with an accumulation of ceramide, a product of sphingolipid metabolism. Furthermore, the exogenous addition of short chain ceramides inhibits EIgG-mediated phagocytosis. In the present study, we identified p42 and p44 mitogen-actived protein (MAP) kinases, referred to as extracellular signal-regulated kinases ERK2 and ERK1, respectively, as intracellular targets of ceramide action during Fc gammaR-mediated phagocytosis. The tyrosine phosphorylation of ERK1 and ERK2 increased within 30 s of addition of EIgG, with maximal phosphorylation by 1 to 5 min. By 30 min, ERK1 and ERK2 were almost completely dephosphorylated. The kinetics of ERK1 and ERK2 tyrosine phosphorylation indicated that MAP kinase activation preceded target ingestion. N-Acetylsphingosine (C2-ceramide) inhibited phagocytosis, reduced ERK1 and ERK2 phosphorylation to basal levels, and reduced ERK1 and ERK2 activity by 85 to 90% and 70 to 80%, respectively. In contrast, N-acetyldihydrosphingosine (dihydro-C2-ceramide) had no effect on either tyrosine phosphorylation or activity of ERK1 and ERK2. In the presence of the MAP kinase kinase (MEK) inhibitor, PD 098059, phagocytosis was reduced by approximately 50%, while ERK1 and ERK2 activity was reduced by 85 to 90%. Thus, engagement of Fc gammaRs led to ERK1 and ERK2 phosphorylation and activation, and the activation of these enzymes was critical for phagocytosis. Furthermore, the inhibition of phagocytosis by C2-ceramide correlated with the inhibition of tyrosine phosphorylation and activation of ERK1 and ERK2. These results suggest that ceramides generated during phagocytosis act on the MAP kinase signaling pathway, ultimately "turning off" the phagocytic response.
Suchard
S J
SJ
Department of Pediatrics, University of Michigan, Ann Arbor 48109, USA.
Mansfield
P J
PJ
Boxer
L A
LA
Shayman
J A
JA
D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002518:Ceramides / Q000494:pharmacology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000502:physiology; D048369:MAP Kinase Kinase 1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D010587:Phagocytosis; D019000:Phosphotyrosine / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011961:Receptors, Fc / Q000502:physiology
D002518:Ceramides; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007074:Immunoglobulin G; D011961:Receptors, Fc; D019000:Phosphotyrosine; D009240:N-Formylmethionine Leucyl-Phenylalanine; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one