Protein kinases involved in the activation of the NADPH oxidase by Fcγ receptors in neutrophils were studied. Of three different protein kinase C (PKC) inhibitors, Gö 6976 inhibited the NADPH oxidase completely, whereas bisindolylmaleimide I and Ro 31-8220 caused a 70–80% inhibition. Thus a Gö 6976-sensitive, bisindolylmaleimide I/Ro 31-8220-insensitive component contributes to NADPH oxidase activation induced by Fcγ receptors. Down-regulation of PKC isotypes resulted in inhibition of Fcγ-receptor-activated NADPH oxidase, but a down-regulation-insensitive component was still present. This component was sensitive to Gö 6976, but insensitive to Ro 31-8220. It has been shown previously that protein kinase D/PKC-μ (PKD) shows this same pharmacology in vitro. We show that PKD is present in neutrophils and that, in contrast with PKC isotypes, PKD is not down-regulated. Therefore PKD may participate in NADPH oxidase activation. To obtain direct evidence for this we adopted an antisense approach. Antisense PKD inhibited NADPH oxidase induced by Fcγ-receptor stimulation by 50% and the Ro 31-8220-insensitive component in the activation was inhibited by antisense PKD. In vitro kinase assays showed that PKD is activated by presenting IgG-opsonized particles to neutrophils. Furthermore, PKD localizes to the area of particle intake in the cell and phosphorylates two of the three cytosolic components of the NADPH oxidase, p40phox and p47phox. Taken together, these data indicate that Fcγ receptors engage PKD in the regulation of the NADPH oxidase.
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April 2002
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Research Article|
March 22 2002
Involvement of protein kinase D in Fcγ-receptor activation of the NADPH oxidase in neutrophils
Jan K. DAVIDSON-MONCADA;
Jan K. DAVIDSON-MONCADA
Centre for Molecular Medicine, Department of Medicine, University College London, The Rayne Institute, 5 University Street, London WC1E 6JJ, U.K.
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Guillermo LOPEZ-LLUCH;
Guillermo LOPEZ-LLUCH
1
Centre for Molecular Medicine, Department of Medicine, University College London, The Rayne Institute, 5 University Street, London WC1E 6JJ, U.K.
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Anthony W. SEGAL;
Anthony W. SEGAL
Centre for Molecular Medicine, Department of Medicine, University College London, The Rayne Institute, 5 University Street, London WC1E 6JJ, U.K.
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Lodewijk V. DEKKER
Lodewijk V. DEKKER
2
Centre for Molecular Medicine, Department of Medicine, University College London, The Rayne Institute, 5 University Street, London WC1E 6JJ, U.K.
2To whom correspondence should be addressed, at Ionix Pharmaceuticals, 185 Cambridge Science Park, Milton Road, Cambridge CB4 0GA, U.K. (e-mail ldekker@ionixpharma.com).
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Publisher: Portland Press Ltd
Received:
September 13 2001
Revision Received:
December 20 2001
Accepted:
January 29 2002
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2002
2002
Biochem J (2002) 363 (1): 95–103.
Article history
Received:
September 13 2001
Revision Received:
December 20 2001
Accepted:
January 29 2002
Citation
Jan K. DAVIDSON-MONCADA, Guillermo LOPEZ-LLUCH, Anthony W. SEGAL, Lodewijk V. DEKKER; Involvement of protein kinase D in Fcγ-receptor activation of the NADPH oxidase in neutrophils. Biochem J 1 April 2002; 363 (1): 95–103. doi: https://doi.org/10.1042/bj3630095
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