Comparison of the roles of mitogen-activated protein kinase kinase and phosphatidylinositol 3-kinase signal transduction in neutrophil effector function

Abstract

Although it is known that many stimuli can activate mitogen-activated protein kinases (MAPKs) and phosphatidylinositol 3-kinases (PI3K) in human neutrophils, little is known concerning either the mechanisms or function of this activation. We have utilized a selective inhibitor of MAPK kinase (MEK), PD098059, and two inhibitors of PI3K, wortmannin and LY294002, to investigate the roles of these kinases in the regulation of neutrophil effector functions. Granulocyte/macrophage colony-stimulating factor, platelet-activating factor (PAF) and N-formylmethionyl-leucyl-phenylalanine are capable of activating both p44ERK1 and p42ERK2 MAPKs and phosphotyrosine-associated PI3K in human neutrophils. The activation of extracellular signal-related protein kinases (ERKs) is correlated with the activation of p21ras by both tyrosine kinase and G-protein-coupled receptors as measured by a novel assay for GTP loading. Wortmannin and LY294002 inhibit, to various degrees, superoxide generation, neutrophil migration and PAF release. Incubation with PD098059, however, inhibits only the PAF release stimulated by serum-treated zymosan. This demonstrates that, while neither MEK nor ERK kinases are involved in the activation of respiratory burst or neutrophil migration, inhibition of PAF release suggests a potential role in the activation of cytosolic phospholipase A2. PI3K isoforms, however, seem to have a much wider role in regulating neutrophil functioning.

Examination of the signal transduction pathways leading to activation of extracellular signal‐regulated kinase by formyl‐methionyl‐leucyl‐phenylalanine in rat neutrophils

“…However, the signal transduction events upstream to ERK occurring in neutrophils in response to fMLP have been incompletely delineated. Based on immunoblot analysis with anti-phosphop44/42 MAPK antibodies, we found that exposure of neutrophils to fMLP rapidly induced the activation of ERK, reconcile the earlier reports [14,15], and these responses were attenuated by 0.3 WM U0126 ( Fig. 1) as well as 3 WM PD98059 (data not shown), both selective MEK inhibitors [16,17].…”

A new phosphospecific cell-based ELISA for p42/p44 mitogen-activated protein kinase (MAPK), p38 MAPK, protein kinase B and cAMP-response-element-binding protein

“…Concentration-and temporal-dependent, PD98059-sensitive, fMLP-induced enzymic product formation was readily detected (Figure 2). The p42\p44 MAPK activation by fMLP closely matched that observed in primary isolated human granulocytes using classical kinase assays or Western-blot analysis [11]. Thus PACE is a specific and sensitive method for detection of p42\p44 MAPK phosphoryl- ation in both adherent and suspension cells.…”

“…Blood was obtained from healthy volunteers. Mixed granulocytes were isolated as described earlier [11].…”

Regulation of Proliferation, Differentiation and Survival by the IL-3/IL-5/GM-CSF Receptor Family

“…However, IL-3 been shown to be activated by IL-3/IL-5/GM-CSF in multifails to activate SEK-1 in FDC-P2 cells [108]. Downstream ple primary cellular lineages and cell lines [22,[86][87][88][89][90].…”

“…Indeed, IL-3/IL-5/GMand DSE-dependent transcription induced by IL-5 [106]. CSF cytokines rapidly induce activation of Ras [22, [89][90][91][92][93] Blocking JNK/SAPK activation with a serine/threonine/tyand c-Raf [22, [94][95][96]. Activation of this pathway will evenrosine phosphatase suggests that this pathway contributes to tually result in enhanced transcription of c-fos and c-jun IL-3-dependent growth but not survival [111].…”