Importance of MEK in neutrophil microbicidal responsiveness.
Exposure of neutrophils to inflammatory stimuli such as the chemoattractant FMLP leads to activation of responses including cell motility, the oxidative burst, and secretion of proteolytic enzymes. A signaling cascade involving sequential activation of Raf-1, mitogen-activated protein kinase (MEK), and extracellular signal regulated kinase (ERK) is also rapidly activated after agonist exposure. The temporal relationship between these events suggests that the kinases may be involved in triggering the effector functions, but direct evidence of a causal relationship is lacking. To assess the role of the MEK/ERK pathway in the activation of neutrophil responses, we studied the effects of PD098059, a potent and selective inhibitor of MEK. Preincubation of human neutrophils with 50 microM PD098059 almost completely (>90%) inhibited the FMLP-induced activation of MEK-1 and MEK-2, the isoforms expressed by neutrophils. This dose of PD098059 virtually abrogated chemoattractant-induced tyrosine phosphorylation and activation of ERK-1 and ERK-2, implying that MEKs are the predominant upstream activators of these mitogen-activated protein kinases. Pretreatment of neutrophils with the MEK antagonist inhibited the oxidative burst substantially and phagocytosis only moderately. In addition, PD098059 antagonized the delay of apoptosis induced by exposure to granulocyte-macrophage CSF. However, the effects of PD098059 were selective, as it failed to inhibit other responses, including chemoattractant-induced exocytosis of primary and secondary granules, polymerization of F-actin, chemotaxis, or activation of phospholipase A2. We conclude that MEK and ERK contribute to the activation of the oxidative burst and phagocytosis, and participate in cytokine regulation of apoptosis.
Downey
G P
GP
Toronto Hospital, and Department of Medicine, University of Toronto, Ontario, Canada. gregory.downey@utoronto.ca
Butler
J R
JR
Tapper
H
H
Fialkow
L
L
Saltiel
A R
AR
Rubin
B B
BB
Grinstein
S
S
D000199:Actins / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001770:Blood Bactericidal Activity; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015550:Cell Degranulation / Q000187:drug effects; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000037:antagonists & inhibitors; D006801:Humans; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D009504:Neutrophils / Q000201:enzymology*; D010587:Phagocytosis / Q000187:drug effects; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D019000:Phosphotyrosine / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism / Q000502:physiology*; D016897:Respiratory Burst / Q000187:drug effects
D000199:Actins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D019000:Phosphotyrosine; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; C482128:MAP2K2 protein, human; 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; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; D010741:Phospholipases A; D054467:Phospholipases A2; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one