title: Chemoattractant-induced signaling via the Ras-ERK and PI3K-Akt networks, along with leukotriene C4 release, is dependent on the tyrosine kinase Lyn in IL-5- and IL-3-primed human blood eosinophils.
abstract: Human blood eosinophils exhibit a hyperactive phenotype in response to chemotactic factors after cell "priming" with IL-5 family cytokines. Earlier work has identified ERK1/2 as molecular markers for IL-5 priming, and in this article, we show that IL-3, a member of the IL-5 family, also augments fMLP-stimulated ERK1/2 phosphorylation in primary eosinophils. Besides ERK1/2, we also observed an enhancement of chemotactic factor-induced Akt phosphorylation after IL-5 priming of human blood eosinophils. Administration of a peptide antagonist that targets the Src family member Lyn before cytokine (IL-5/IL-3) priming of blood eosinophils inhibited the synergistic increase of fMLP-induced activation of Ras, ERK1/2 and Akt, as well as the release of the proinflammatory factor leukotriene C(4). In this study, we also examined a human eosinophil-like cell line HL-60 clone-15 and observed that these cells exhibited significant surface expression of IL-3Rs and GM-CSFRs, as well as ERK1/2 phosphorylation in response to the addition of IL-5 family cytokines or the chemotactic factors fMLP, CCL5, and CCL11. Consistent with the surface profile of IL-5 family receptors, HL-60 clone-15 recapitulated the enhanced fMLP-induced ERK1/2 phosphorylation observed in primary blood eosinophils after priming with IL-3/GM-CSF, and small interfering RNA-mediated knockdown of Lyn expression completely abolished the synergistic effects of IL-3 priming on fMLP-induced ERK1/2 phosphorylation. Altogether, our data demonstrate a central role for Lyn in the mechanisms of IL-5 family priming and suggest that Lyn contributes to the upregulation of the Ras-ERK1/2 and PI3K-Akt cascades, as well as the increased leukotriene C(4) release observed in response to fMLP in "primed" eosinophils.
authors.affiliation: Molecular and Cellular Pharmacology Program, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Yiming
authors.initials: Y
authors.lastname: Zhu
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Bertics
mesh_terms: D000293:Adolescent; D000328:Adult; D000595:Amino Acid Sequence; D001249:Asthma / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D004804:Eosinophils / Q000276:immunology* / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D007377:Interleukin-3 / Q000502:physiology*; D015848:Interleukin-5 / Q000502:physiology*; D017997:Leukotriene C4 / Q000378:metabolism*; D008875:Middle Aged; D019950:Mitogen-Activated Protein Kinase 1 / Q000502:physiology*; D048052:Mitogen-Activated Protein Kinase 3 / Q000502:physiology*; D008969:Molecular Sequence Data; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology*; D015398:Signal Transduction / Q000276:immunology; D018631:ras Proteins / Q000502:physiology*; D019061:src-Family Kinases / Q000502:physiology*
chemical_list: C508596:IL3 protein, human; C508599:IL5 protein, human; D007377:Interleukin-3; D015848:Interleukin-5; D017997:Leukotriene C4; D009240:N-Formylmethionine Leucyl-Phenylalanine; D019869:Phosphatidylinositol 3-Kinases; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D018631:ras Proteins