Interleukin-8-induced priming of neutrophil oxidative burst requires sequential recruitment of NADPH oxidase components into lipid rafts.
The superoxide-producing phagocyte NADPH oxidase consists of a membrane-bound flavocytochrome b(558), the cytosol factors p47(phox), p67(phox), p40(phox), and the small GTPase Rac2, which translocate to the membrane to assemble the active complex following neutrophil activation. Interleukin-8 (IL-8) does not activate NADPH oxidase, but potentiates the oxidative burst induced by stimuli such as formyl-methionyl-leucyl-phenylalanine (fMLP) via a priming mechanism. The effect of IL-8 on the components of NADPH oxidase during the priming process has never been investigated in human neutrophils. Here we showed that within 3 min, IL-8 treatment enhanced the Btk- and ERK1/2-dependent phosphorylation of p47(phox), as well as the recruitment of flavocytochrome b(558), p47(phox), and Rac2 into cholesterol-enriched detergent-resistant microdomains (or lipid rafts). Conversely, IL-8 treatment lasting 15 min failed to recruit flavocytochrome b(558), p47(phox), or Rac2, but did enhance the Btk- and p38 MAPK-dependent phosphorylation and the translocation of p67(phox) into detergent-resistant microdomains. Moreover, methyl-beta-cyclodextrin, which disrupts lipid rafts, inhibited IL-8-induced priming in response to fMLP. Our findings indicate that IL-8-induced priming of the oxidative burst in response to fMLP involves a sequential assembly of the NADPH oxidase components in the lipid rafts of neutrophils.
Guichard
Cécile
C
Unité INSERM 683, Facultédemédecine Xavier BICHAT, BP 416, 75870 Paris Cedex 18, France. guichard@bichat.inserm.fr
Pedruzzi
Eric
E
Dewas
Cédric
C
Fay
Michèle
M
Pouzet
Cécile
C
Bens
Marcelle
M
Vandewalle
Alain
A
Ogier-Denis
Eric
E
Gougerot-Pocidalo
Marie-Anne
MA
Elbim
Carole
C
D000077329:Agammaglobulinaemia Tyrosine Kinase; D003573:Cytochrome b Group / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000494:pharmacology*; D008055:Lipids; 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*; D009504:Neutrophils / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D016897:Respiratory Burst; D013481:Superoxides / Q000378:metabolism*; D047392:beta-Cyclodextrins / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism; D000097766:RAC2 GTP-Binding Protein
D000077329:Agammaglobulinaemia Tyrosine Kinase; D003573:Cytochrome b Group; D016209:Interleukin-8; D008055:Lipids; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D009240:N-Formylmethionine Leucyl-Phenylalanine; D019255:NADPH Oxidases; D010750:Phosphoproteins; D011505:Protein-Tyrosine Kinases; D013481:Superoxides; D047392:beta-Cyclodextrins; D048051:p38 Mitogen-Activated Protein Kinases; D020744:rac GTP-Binding Proteins; D000097766:RAC2 GTP-Binding Protein; C108732:methyl-beta-cyclodextrin; C085072:neutrophil cytosol factor 40K; C079789:neutrophil cytosol factor 67K; C036888:cytochrome b558; C060505:neutrophil cytosolic factor 1; C000625949:BTK protein, human