Phosphoinositide 3-kinase regulates the phosphorylation of NADPH oxidase component p47(phox) by controlling cPKC/PKCdelta but not Akt

Biochem Biophys Res Commun. 2004 Apr 9;316(3):720-30. doi: 10.1016/j.bbrc.2004.02.108.

Abstract

Superoxide production by NADPH oxidase is essential for the bactericidal properties of phagocytes. Phosphorylation of p47(phox), one of the cytosolic components of NADPH oxidase, is a crucial step of the oxidase activation. Some evidences suggest that phosphoinositide 3-kinase (PI3K) is involved in p47(phox) phosphorylation, but it has not been fully understood how PI3K regulates it. The aim of this study was to examine the mechanism underlying the PI3K regulation of p47(phox) phosphorylation. Pharmacological inhibition of PI3K attenuated both fMLP-stimulated p47(phox) phosphorylation and NADPH oxidase activity in HL-60 cells differentiated to a neutrophil-like phenotype. Although fMLP elicited Akt activation in a PI3K-dependent manner, an Akt inhibitor had no effect on the oxidase activity triggered by fMLP. In vitro kinase assay revealed that Akt was unable to catalyze p47(phox) phosphorylation. Interestingly, the activation of cPKC and PKCdelta after fMLP stimulation was dependent on PI3K. Furthermore, PI3K inhibitors reduced the activation of phospholipase Cgamma2 without affecting tyrosine phosphorylation on it. These results suggest that PI3K regulates the phosphorylation of NADPH oxidase component p47(phox) by controlling diacylglycerol-dependent PKCs but not Akt.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Androstadienes / pharmacology
  • Animals
  • Blotting, Western
  • Cell Differentiation
  • Cell Membrane / metabolism
  • Chromones / pharmacology
  • Cytosol / enzymology
  • Cytosol / metabolism
  • DNA, Complementary / metabolism
  • Enzyme Activation
  • Enzyme Inhibitors / pharmacology
  • Glutathione Transferase / metabolism
  • HL-60 Cells
  • Humans
  • Immunoblotting
  • Macrophages / metabolism
  • Mice
  • Models, Biological
  • Monocytes / metabolism
  • Morpholines / pharmacology
  • N-Formylmethionine Leucyl-Phenylalanine / pharmacology
  • NADPH Oxidases / metabolism*
  • Neutrophils / cytology
  • Phenotype
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphatidylinositol 3-Kinases / physiology*
  • Phospholipase C gamma
  • Phosphoproteins / metabolism*
  • Phosphorylation*
  • Protein Kinase C / metabolism*
  • Protein Kinase C-delta
  • Protein-Serine-Threonine Kinases / metabolism*
  • Proto-Oncogene Proteins c-akt
  • Proto-Oncogene Proteins*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Subcellular Fractions / metabolism
  • Superoxides / metabolism
  • Time Factors
  • Type C Phospholipases / metabolism
  • Tyrosine / metabolism
  • Wortmannin

Substances

  • Androstadienes
  • Chromones
  • DNA, Complementary
  • Enzyme Inhibitors
  • Morpholines
  • Phosphoproteins
  • Proto-Oncogene Proteins
  • Recombinant Proteins
  • Superoxides
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Tyrosine
  • N-Formylmethionine Leucyl-Phenylalanine
  • NADPH Oxidases
  • neutrophil cytosolic factor 1
  • Glutathione Transferase
  • Phosphatidylinositol 3-Kinases
  • Prkcd protein, mouse
  • AKT1 protein, human
  • Protein-Serine-Threonine Kinases
  • Proto-Oncogene Proteins c-akt
  • PRKCD protein, human
  • Protein Kinase C
  • Protein Kinase C-delta
  • Type C Phospholipases
  • Phospholipase C gamma
  • Wortmannin