Regulation of TNF-induced oxygen radical production in human neutrophils: role of delta-PKC.
In human neutrophils, TNF-elicited O(2)(-) production requires adherence and integrin activation. How this cooperative signaling between TNFRs and integrins regulates O(2)(-) generation has yet to be fully elucidated. Previously, we identified delta-PKC as a critical early regulator of TNF signaling in adherent neutrophils. In this study, we demonstrate that inhibition of delta-PKC with a dominant-negative delta-PKC TAT peptide resulted in a significant delay in the onset time of TNF-elicited O(2)(-) generation but had no effect on Vmax, indicating an involvement of delta-PKC in the initiation of O(2)(-) production. In contrast, fMLP-elicited O(2)(-) production in adherent and nonadherent neutrophils was delta-PKC-independent, suggesting differential regulation of O(2)(-) production. An important step in activation of the NADPH oxidase is phosphorylation of the cytosolic p47phox component. In adherent neutrophils, TNF triggered a time-dependent association of delta-PKC with p47phox, which was associated with p47phox phosphorylation, indicating a role for delta-PKC in regulating O(2)(-) production at the level of p47phox. Activation of ERK and p38 MAPK is also required for TNF-elicited O(2)(-) generation. TNF-mediated ERK but not p38 MAPK recruitment to p47phox was delta-PKC-dependent. delta-PKC activity is controlled through serine/threonine phosphorylation, and phosphorylation of delta-PKC (Ser643) and delta-PKC (Thr505) was increased significantly by TNF in adherent cells via a PI3K-dependent process. Thus, signaling for TNF-elicited O(2)(-) generation is regulated by delta-PKC. Adherence-dependent cooperative signaling activates PI3K signaling, delta-PKC phosphorylation, and delta-PKC recruitment to p47phox. delta-PKC activates p47phox by serine phosphorylation or indirectly through control of ERK recruitment to p47phox.
Department of Physiology and Lung Center, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. laurie.kilpatrick@temple.edu
Laurie E
LE
Kilpatrick
Shuang
S
Sun
Haiying
H
Li
Thomas C
TC
Vary
Helen M
HM
Korchak
D000328:Adult; D002448:Cell Adhesion; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D019255:NADPH Oxidases / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000502:physiology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism*; D016897:Respiratory Burst / Q000187:drug effects / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000502:physiology; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000494:pharmacology
D010446:Peptide Fragments; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D054322:tat Gene Products, Human Immunodeficiency Virus; C497137:tat peptide (47-57), Human immunodeficiency virus 1; D009240:N-Formylmethionine Leucyl-Phenylalanine; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D051745:Protein Kinase C-delta; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases