GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR AND IL-5 ACTIVATE MITOGEN-ACTIVATED PROTEIN-KINASE THROUGH JAK2 KINASE AND PHOSPHATIDYLINOSITOL 3-KINASE IN HUMAN EOSINOPHILS

Mitogen-activated protein (MAP) kinases are activated by the sequentia l activation of Ras, Raf and MEK (MAP kinase kinase) and regulate a wi de variety of cell functions. To determine the kinase cascade for gran ulocyte-macrophage colony-stimulating factor (GM-CSF)- and IL-5-induce d MAP kinase activation in eosinophils, we studied the effect of inhib itors of Jak2 kinase, tyrosine kinases, phosphatidylinositol 3-kinase, and protein kinase C on GM-CSF-and IL-5-induced MAP kinase activation in human eosinophils. GMCSF and IL-5 activated 40, 42, and 44 kilodal ton MAP kinase isoforms in eosinophils. This was indicated by the elec trophoretic mobility shift of the three isoforms of MAP kinase in immu noblotting with anti-MAP kinase antibody and also by in-gel MAP kinase assay. MAP kinase activation was time-and dose-dependent, becoming ma ximal 3 to 15 minutes after stimulation. A Jak2 kinase inhibitor AG-49 0, a tyrosine kinase inhibitor genistein, and a phosphatidylinositol 3 -kinase inhibitor wortmannin inhibited GM-CSF- and IL-5-induced MAP ki nase activation in eosinophils, whereas a protein kinase C inhibitor s taurosporine had a weak inhibitory effect. Furthermore, AG-490 and gen istein prevented GM-CSF-induced tyrosine phosphorylation of Jak2 kinas e in eosinophils. Taken together, these results indicate that GMCSF an d IL-5 activate MAP kinases through the signaling pathway of Jak2 kina se-tyrosine phosphorylated beta chain-phosphatidylinositol 3-kinase-Ra s in eosinophils.