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. 1998 Jan 1;329(Pt 1):121–130. doi: 10.1042/bj3290121

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

P J Coffer 1, N Geijsen 1, L M'rabet 1, R C Schweizer 1, T Maikoe 1, J A Raaijmakers 1, J W Lammers 1, L Koenderman 1
PMCID: PMC1219022  PMID: 9405284

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.

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Selected References

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  1. Alessi D. R., Cuenda A., Cohen P., Dudley D. T., Saltiel A. R. PD 098059 is a specific inhibitor of the activation of mitogen-activated protein kinase kinase in vitro and in vivo. J Biol Chem. 1995 Nov 17;270(46):27489–27494. doi: 10.1074/jbc.270.46.27489. [DOI] [PubMed] [Google Scholar]
  2. Arcaro A., Wymann M. P. Wortmannin is a potent phosphatidylinositol 3-kinase inhibitor: the role of phosphatidylinositol 3,4,5-trisphosphate in neutrophil responses. Biochem J. 1993 Dec 1;296(Pt 2):297–301. doi: 10.1042/bj2960297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. BOYDEN S. The chemotactic effect of mixtures of antibody and antigen on polymorphonuclear leucocytes. J Exp Med. 1962 Mar 1;115:453–466. doi: 10.1084/jem.115.3.453. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berkow R. L., Wang D., Larrick J. W., Dodson R. W., Howard T. H. Enhancement of neutrophil superoxide production by preincubation with recombinant human tumor necrosis factor. J Immunol. 1987 Dec 1;139(11):3783–3791. [PubMed] [Google Scholar]
  5. Burgering B. M., Coffer P. J. Protein kinase B (c-Akt) in phosphatidylinositol-3-OH kinase signal transduction. Nature. 1995 Aug 17;376(6541):599–602. doi: 10.1038/376599a0. [DOI] [PubMed] [Google Scholar]
  6. Börsch-Haubold A. G., Kramer R. M., Watson S. P. Inhibition of mitogen-activated protein kinase kinase does not impair primary activation of human platelets. Biochem J. 1996 Aug 15;318(Pt 1):207–212. doi: 10.1042/bj3180207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chanock S. J., el Benna J., Smith R. M., Babior B. M. The respiratory burst oxidase. J Biol Chem. 1994 Oct 7;269(40):24519–24522. [PubMed] [Google Scholar]
  8. Chao W., Olson M. S. Platelet-activating factor: receptors and signal transduction. Biochem J. 1993 Jun 15;292(Pt 3):617–629. doi: 10.1042/bj2920617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clark-Lewis I., Sanghera J. S., Pelech S. L. Definition of a consensus sequence for peptide substrate recognition by p44mpk, the meiosis-activated myelin basic protein kinase. J Biol Chem. 1991 Aug 15;266(23):15180–15184. [PubMed] [Google Scholar]
  10. Coffer P. J., Koenderman L. Granulocyte signal transduction and priming: cause without effect? Immunol Lett. 1997 Jun 1;57(1-3):27–31. doi: 10.1016/s0165-2478(97)00067-9. [DOI] [PubMed] [Google Scholar]
  11. Dewald B., Thelen M., Baggiolini M. Two transduction sequences are necessary for neutrophil activation by receptor agonists. J Biol Chem. 1988 Nov 5;263(31):16179–16184. [PubMed] [Google Scholar]
  12. Didichenko S. A., Tilton B., Hemmings B. A., Ballmer-Hofer K., Thelen M. Constitutive activation of protein kinase B and phosphorylation of p47phox by a membrane-targeted phosphoinositide 3-kinase. Curr Biol. 1996 Oct 1;6(10):1271–1278. doi: 10.1016/s0960-9822(02)70713-6. [DOI] [PubMed] [Google Scholar]
  13. Ding J., Vlahos C. J., Liu R., Brown R. F., Badwey J. A. Antagonists of phosphatidylinositol 3-kinase block activation of several novel protein kinases in neutrophils. J Biol Chem. 1995 May 12;270(19):11684–11691. doi: 10.1074/jbc.270.19.11684. [DOI] [PubMed] [Google Scholar]
  14. Downey G. P. Mechanisms of leukocyte motility and chemotaxis. Curr Opin Immunol. 1994 Feb;6(1):113–124. doi: 10.1016/0952-7915(94)90042-6. [DOI] [PubMed] [Google Scholar]
  15. Dudley D. T., Pang L., Decker S. J., Bridges A. J., Saltiel A. R. A synthetic inhibitor of the mitogen-activated protein kinase cascade. Proc Natl Acad Sci U S A. 1995 Aug 15;92(17):7686–7689. doi: 10.1073/pnas.92.17.7686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dusi S., Della Bianca V., Donini M., Nadalini K. A., Rossi F. Mechanisms of stimulation of the respiratory burst by TNF in nonadherent neutrophils: its independence of lipidic transmembrane signaling and dependence on protein tyrosine phosphorylation and cytoskeleton. J Immunol. 1996 Nov 15;157(10):4615–4623. [PubMed] [Google Scholar]
  17. Dusi S., Donini M., Wientjes F., Rossi F. Tyrosine phosphorylation and subcellular redistribution of p125 ras guanosine triphosphatase-activating protein in human neutrophils stimulated with FMLP. FEBS Lett. 1996 Apr 1;383(3):181–184. doi: 10.1016/0014-5793(96)00248-7. [DOI] [PubMed] [Google Scholar]
  18. Eberle M., Traynor-Kaplan A. E., Sklar L. A., Norgauer J. Is there a relationship between phosphatidylinositol trisphosphate and F-actin polymerization in human neutrophils? J Biol Chem. 1990 Oct 5;265(28):16725–16728. [PubMed] [Google Scholar]
  19. El Benna J., Faust R. P., Johnson J. L., Babior B. M. Phosphorylation of the respiratory burst oxidase subunit p47phox as determined by two-dimensional phosphopeptide mapping. Phosphorylation by protein kinase C, protein kinase A, and a mitogen-activated protein kinase. J Biol Chem. 1996 Mar 15;271(11):6374–6378. doi: 10.1074/jbc.271.11.6374. [DOI] [PubMed] [Google Scholar]
  20. Faust L. R., el Benna J., Babior B. M., Chanock S. J. The phosphorylation targets of p47phox, a subunit of the respiratory burst oxidase. Functions of the individual target serines as evaluated by site-directed mutagenesis. J Clin Invest. 1995 Sep;96(3):1499–1505. doi: 10.1172/JCI118187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Fialkow L., Chan C. K., Rotin D., Grinstein S., Downey G. P. Activation of the mitogen-activated protein kinase signaling pathway in neutrophils. Role of oxidants. J Biol Chem. 1994 Dec 9;269(49):31234–31242. [PubMed] [Google Scholar]
  22. Franke T. F., Yang S. I., Chan T. O., Datta K., Kazlauskas A., Morrison D. K., Kaplan D. R., Tsichlis P. N. The protein kinase encoded by the Akt proto-oncogene is a target of the PDGF-activated phosphatidylinositol 3-kinase. Cell. 1995 Jun 2;81(5):727–736. doi: 10.1016/0092-8674(95)90534-0. [DOI] [PubMed] [Google Scholar]
  23. Gerard C., Gerard N. P. The pro-inflammatory seven-transmembrane segment receptors of the leukocyte. Curr Opin Immunol. 1994 Feb;6(1):140–145. doi: 10.1016/0952-7915(94)90045-0. [DOI] [PubMed] [Google Scholar]
  24. Gomez-Cambronero J., Huang C. K., Gomez-Cambronero T. M., Waterman W. H., Becker E. L., Sha'afi R. I. Granulocyte-macrophage colony-stimulating factor-induced protein tyrosine phosphorylation of microtubule-associated protein kinase in human neutrophils. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7551–7555. doi: 10.1073/pnas.89.16.7551. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Gonzalez F. A., Raden D. L., Davis R. J. Identification of substrate recognition determinants for human ERK1 and ERK2 protein kinases. J Biol Chem. 1991 Nov 25;266(33):22159–22163. [PubMed] [Google Scholar]
  26. Grinstein S., Furuya W. Chemoattractant-induced tyrosine phosphorylation and activation of microtubule-associated protein kinase in human neutrophils. J Biol Chem. 1992 Sep 5;267(25):18122–18125. [PubMed] [Google Scholar]
  27. Haslett C., Savill J. S., Meagher L. The neutrophil. Curr Opin Immunol. 1989 Oct;2(1):10–18. doi: 10.1016/0952-7915(89)90091-5. [DOI] [PubMed] [Google Scholar]
  28. Hawkins P. T., Eguinoa A., Qiu R. G., Stokoe D., Cooke F. T., Walters R., Wennström S., Claesson-Welsh L., Evans T., Symons M. PDGF stimulates an increase in GTP-Rac via activation of phosphoinositide 3-kinase. Curr Biol. 1995 Apr 1;5(4):393–403. doi: 10.1016/s0960-9822(95)00080-7. [DOI] [PubMed] [Google Scholar]
  29. Hayakawa T., Suzuki K., Suzuki S., Andrews P. C., Babior B. M. A possible role for protein phosphorylation in the activation of the respiratory burst in human neutrophils. Evidence from studies with cells from patients with chronic granulomatous disease. J Biol Chem. 1986 Jul 15;261(20):9109–9115. [PubMed] [Google Scholar]
  30. Honda Z., Takano T., Gotoh Y., Nishida E., Ito K., Shimizu T. Transfected platelet-activating factor receptor activates mitogen-activated protein (MAP) kinase and MAP kinase kinase in Chinese hamster ovary cells. J Biol Chem. 1994 Jan 21;269(3):2307–2315. [PubMed] [Google Scholar]
  31. Hulett M. D., Hogarth P. M. Molecular basis of Fc receptor function. Adv Immunol. 1994;57:1–127. doi: 10.1016/s0065-2776(08)60671-9. [DOI] [PubMed] [Google Scholar]
  32. Karin M., Hunter T. Transcriptional control by protein phosphorylation: signal transmission from the cell surface to the nucleus. Curr Biol. 1995 Jul 1;5(7):747–757. doi: 10.1016/s0960-9822(95)00151-5. [DOI] [PubMed] [Google Scholar]
  33. Klebanoff S. J., Vadas M. A., Harlan J. M., Sparks L. H., Gamble J. R., Agosti J. M., Waltersdorph A. M. Stimulation of neutrophils by tumor necrosis factor. J Immunol. 1986 Jun 1;136(11):4220–4225. [PubMed] [Google Scholar]
  34. Koenderman L., Kok P. T., Hamelink M. L., Verhoeven A. J., Bruijnzeel P. L. An improved method for the isolation of eosinophilic granulocytes from peripheral blood of normal individuals. J Leukoc Biol. 1988 Aug;44(2):79–86. doi: 10.1002/jlb.44.2.79. [DOI] [PubMed] [Google Scholar]
  35. Kotani K., Yonezawa K., Hara K., Ueda H., Kitamura Y., Sakaue H., Ando A., Chavanieu A., Calas B., Grigorescu F. Involvement of phosphoinositide 3-kinase in insulin- or IGF-1-induced membrane ruffling. EMBO J. 1994 May 15;13(10):2313–2321. doi: 10.1002/j.1460-2075.1994.tb06515.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Kramer R. M., Roberts E. F., Strifler B. A., Johnstone E. M. Thrombin induces activation of p38 MAP kinase in human platelets. J Biol Chem. 1995 Nov 17;270(46):27395–27398. doi: 10.1074/jbc.270.46.27395. [DOI] [PubMed] [Google Scholar]
  37. Leyravaud S., Bossant M. J., Joly F., Bessou G., Benveniste J., Ninio E. Biosynthesis of paf-acether. X. Phorbol myristate acetate-induced paf-acether biosynthesis and acetyltransferase activation in human neutrophils. J Immunol. 1989 Jul 1;143(1):245–249. [PubMed] [Google Scholar]
  38. Lin L. L., Wartmann M., Lin A. Y., Knopf J. L., Seth A., Davis R. J. cPLA2 is phosphorylated and activated by MAP kinase. Cell. 1993 Jan 29;72(2):269–278. doi: 10.1016/0092-8674(93)90666-e. [DOI] [PubMed] [Google Scholar]
  39. Nahas N., Molski T. F., Fernandez G. A., Sha'afi R. I. Tyrosine phosphorylation and activation of a new mitogen-activated protein (MAP)-kinase cascade in human neutrophils stimulated with various agonists. Biochem J. 1996 Aug 15;318(Pt 1):247–253. doi: 10.1042/bj3180247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Nobes C. D., Hawkins P., Stephens L., Hall A. Activation of the small GTP-binding proteins rho and rac by growth factor receptors. J Cell Sci. 1995 Jan;108(Pt 1):225–233. doi: 10.1242/jcs.108.1.225. [DOI] [PubMed] [Google Scholar]
  41. Okada T., Sakuma L., Fukui Y., Hazeki O., Ui M. Blockage of chemotactic peptide-induced stimulation of neutrophils by wortmannin as a result of selective inhibition of phosphatidylinositol 3-kinase. J Biol Chem. 1994 Feb 4;269(5):3563–3567. [PubMed] [Google Scholar]
  42. Panayotou G., Waterfield M. D. Phosphatidyl-inositol 3-kinase: a key enzyme in diverse signalling processes. Trends Cell Biol. 1992 Dec;2(12):358–360. doi: 10.1016/0962-8924(92)90042-l. [DOI] [PubMed] [Google Scholar]
  43. Pick E., Mizel D. Rapid microassays for the measurement of superoxide and hydrogen peroxide production by macrophages in culture using an automatic enzyme immunoassay reader. J Immunol Methods. 1981;46(2):211–226. doi: 10.1016/0022-1759(81)90138-1. [DOI] [PubMed] [Google Scholar]
  44. Pillinger M. H., Feoktistov A. S., Capodici C., Solitar B., Levy J., Oei T. T., Philips M. R. Mitogen-activated protein kinase in neutrophils and enucleate neutrophil cytoplasts: evidence for regulation of cell-cell adhesion. J Biol Chem. 1996 May 17;271(20):12049–12056. doi: 10.1074/jbc.271.20.12049. [DOI] [PubMed] [Google Scholar]
  45. Ridley A. J., Paterson H. F., Johnston C. L., Diekmann D., Hall A. The small GTP-binding protein rac regulates growth factor-induced membrane ruffling. Cell. 1992 Aug 7;70(3):401–410. doi: 10.1016/0092-8674(92)90164-8. [DOI] [PubMed] [Google Scholar]
  46. Sandborg R. R., Smolen J. E. Early biochemical events in leukocyte activation. Lab Invest. 1988 Sep;59(3):300–320. [PubMed] [Google Scholar]
  47. Schweizer R. C., van Kessel-Welmers B. A., Warringa R. A., Maikoe T., Raaijmakers J. A., Lammers J. W., Koenderman L. Mechanisms involved in eosinophil migration. Platelet-activating factor-induced chemotaxis and interleukin-5-induced chemokinesis are mediated by different signals. J Leukoc Biol. 1996 Mar;59(3):347–356. doi: 10.1002/jlb.59.3.347. [DOI] [PubMed] [Google Scholar]
  48. Segal A. W., Abo A. The biochemical basis of the NADPH oxidase of phagocytes. Trends Biochem Sci. 1993 Feb;18(2):43–47. doi: 10.1016/0968-0004(93)90051-n. [DOI] [PubMed] [Google Scholar]
  49. Segal A. W. The NADPH oxidase and chronic granulomatous disease. Mol Med Today. 1996 Mar;2(3):129–135. doi: 10.1016/1357-4310(96)88723-5. [DOI] [PubMed] [Google Scholar]
  50. Sha'afi R. I., Molski T. F. Activation of the neutrophil. Prog Allergy. 1988;42:1–64. doi: 10.1159/000318681. [DOI] [PubMed] [Google Scholar]
  51. Sisson J. H., Prescott S. M., McIntyre T. M., Zimmerman G. A. Production of platelet-activating factor by stimulated human polymorphonuclear leukocytes. Correlation of synthesis with release, functional events, and leukotriene B4 metabolism. J Immunol. 1987 Jun 1;138(11):3918–3926. [PubMed] [Google Scholar]
  52. Stephens L., Smrcka A., Cooke F. T., Jackson T. R., Sternweis P. C., Hawkins P. T. A novel phosphoinositide 3 kinase activity in myeloid-derived cells is activated by G protein beta gamma subunits. Cell. 1994 Apr 8;77(1):83–93. doi: 10.1016/0092-8674(94)90237-2. [DOI] [PubMed] [Google Scholar]
  53. Tartaglia L. A., Goeddel D. V. Two TNF receptors. Immunol Today. 1992 May;13(5):151–153. doi: 10.1016/0167-5699(92)90116-O. [DOI] [PubMed] [Google Scholar]
  54. Thelen M., Wymann M. P., Langen H. Wortmannin binds specifically to 1-phosphatidylinositol 3-kinase while inhibiting guanine nucleotide-binding protein-coupled receptor signaling in neutrophil leukocytes. Proc Natl Acad Sci U S A. 1994 May 24;91(11):4960–4964. doi: 10.1073/pnas.91.11.4960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Thompson H. L., Marshall C. J., Saklatvala J. Characterization of two different forms of mitogen-activated protein kinase kinase induced in polymorphonuclear leukocytes following stimulation by N-formylmethionyl-leucyl-phenylalanine or granulocyte-macrophage colony-stimulating factor. J Biol Chem. 1994 Apr 1;269(13):9486–9492. [PubMed] [Google Scholar]
  56. Torres M., Hall F. L., O'Neill K. Stimulation of human neutrophils with formyl-methionyl-leucyl-phenylalanine induces tyrosine phosphorylation and activation of two distinct mitogen-activated protein-kinases. J Immunol. 1993 Feb 15;150(4):1563–1577. [PubMed] [Google Scholar]
  57. Treisman R. Regulation of transcription by MAP kinase cascades. Curr Opin Cell Biol. 1996 Apr;8(2):205–215. doi: 10.1016/s0955-0674(96)80067-6. [DOI] [PubMed] [Google Scholar]
  58. Vlahos C. J., Matter W. F., Brown R. F., Traynor-Kaplan A. E., Heyworth P. G., Prossnitz E. R., Ye R. D., Marder P., Schelm J. A., Rothfuss K. J. Investigation of neutrophil signal transduction using a specific inhibitor of phosphatidylinositol 3-kinase. J Immunol. 1995 Mar 1;154(5):2413–2422. [PubMed] [Google Scholar]
  59. Vlahos C. J., Matter W. F. Signal transduction in neutrophil activation. Phosphatidylinositol 3-kinase is stimulated without tyrosine phosphorylation. FEBS Lett. 1992 Sep 14;309(3):242–248. doi: 10.1016/0014-5793(92)80781-b. [DOI] [PubMed] [Google Scholar]
  60. Waterman W. H., Sha'afi R. I. Effects of granulocyte-macrophage colony-stimulating factor and tumour necrosis factor-alpha on tyrosine phosphorylation and activation of mitogen-activated protein kinases in human neutrophils. Biochem J. 1995 Apr 1;307(Pt 1):39–45. doi: 10.1042/bj3070039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Wennström S., Hawkins P., Cooke F., Hara K., Yonezawa K., Kasuga M., Jackson T., Claesson-Welsh L., Stephens L. Activation of phosphoinositide 3-kinase is required for PDGF-stimulated membrane ruffling. Curr Biol. 1994 May 1;4(5):385–393. doi: 10.1016/s0960-9822(00)00087-7. [DOI] [PubMed] [Google Scholar]
  62. Worthen G. S., Avdi N., Buhl A. M., Suzuki N., Johnson G. L. FMLP activates Ras and Raf in human neutrophils. Potential role in activation of MAP kinase. J Clin Invest. 1994 Aug;94(2):815–823. doi: 10.1172/JCI117401. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. de Rooij J., Bos J. L. Minimal Ras-binding domain of Raf1 can be used as an activation-specific probe for Ras. Oncogene. 1997 Feb 6;14(5):623–625. doi: 10.1038/sj.onc.1201005. [DOI] [PubMed] [Google Scholar]
  64. de Vries-Smits A. M., Burgering B. M., Leevers S. J., Marshall C. J., Bos J. L. Involvement of p21ras in activation of extracellular signal-regulated kinase 2. Nature. 1992 Jun 18;357(6379):602–604. doi: 10.1038/357602a0. [DOI] [PubMed] [Google Scholar]
  65. el Benna J., Faust L. P., Babior B. M. The phosphorylation of the respiratory burst oxidase component p47phox during neutrophil activation. Phosphorylation of sites recognized by protein kinase C and by proline-directed kinases. J Biol Chem. 1994 Sep 23;269(38):23431–23436. [PubMed] [Google Scholar]

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