title: Protein kinase B (AKT) mediates phospholipase D activation via ERK1/2 and promotes respiratory burst parameters in formylpeptide-stimulated neutrophil-like HL-60 cells.
abstract: Phospholipase D (PLD), a major source of lipid second messengers (phosphatidic acid, diglycerides) in many cell types, is tightly regulated by protein kinases, but only a few of them have been identified. We show here that protein kinase B (AKT) is a novel major signaling effector of PLD activity induced by the formylpeptide f-Met-Leu-Phe (fMLP) in human neutrophil-like HL-60 cells (dHL-60 cells). AKT inhibition with the selective antagonist AKTib1/2 almost completely prevented fMLP-mediated activity of PLD, its upstream effector ERK1/2, but not p38 MAPK. Immunoprecipitation studies show that phosphorylated AKT, ERK, and PLD2 form a complex induced by fMLP, which can be prevented by AKTib1/2. In cell-free systems, AKT1 stimulated PLD activity via activation of ERK. AKT1 actually phosphorylated ERK2 as a substrate (K(m) 1 μm). Blocking AKT activation with AKTib1/2 also prevented fMLP- but not phorbol 12-myristate 13-acetate-mediated NADPH oxidase activation (respiratory burst, RB) of dHL-60 cells. Impaired RB was associated with defective membrane translocation of NADPH oxidase components p67(phox) and p47(phox), ERK, AKT1, AKT2, but not AKT3. Depletion of AKT1 or AKT2 with antisense oligonucleotides further indicates a partial contribution of both isoforms in fMLP-induced activation of ERK, PLD, and RB, with a predominant role of AKT1. Thus, formylpeptides induce sequential activation of AKT, ERK1/2, and PLD, which represents a novel signaling pathway. A major primarily role of this AKT signaling pathway also emerges in membrane recruitment of NOX2 components p47(phox), p67(phox), and ERK, which may contribute to assembly and activation of the RB motor system, NADPH oxidase.
authors.affiliation: Institut Cochin, Université Paris Descartes, CNRS UMR 8104, Paris, France.
authors.forename: Satyananda
authors.initials: S
authors.lastname: Patel
authors.forename: Bahia
authors.initials: B
authors.lastname: Djerdjouri
authors.forename: Yannick
authors.initials: Y
authors.lastname: Raoul-Des-Essarts
authors.forename: Pham My-Chan
authors.initials: PM
authors.lastname: Dang
authors.forename: Jamel
authors.initials: J
authors.lastname: El-Benna
authors.forename: Axel
authors.initials: A
authors.lastname: Périanin
mesh_terms: D004789:Enzyme Activation; D018922:HL-60 Cells / Q000187:drug effects / Q000502:physiology; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D019255:NADPH Oxidases / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016897:Respiratory Burst / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D007527:Isoenzymes; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D009240:N-Formylmethionine Leucyl-Phenylalanine; D019255:NADPH Oxidases; D051057:Proto-Oncogene Proteins c-akt; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020930:MAP Kinase Kinase Kinases; C069937:MAP3K8 protein, human; D010739:Phospholipase D