Abstract
Protein kinase C (PKC) plays a prominent role in immune signaling. To elucidate the signal transduction in a respiratory burst and isoform-specific function of PKC during FcgammaR-mediated phagocytosis, we used live, digital fluorescence imaging of mouse microglial cells expressing GFP-tagged molecules. betaI PKC, epsilonPKC, and diacylglycerol kinase (DGK) beta dynamically and transiently accumulated around IgG-opsonized beads (BIgG). Moreover, the accumulation of p47(phox), an essential cytosolic component of NADPH oxidase and a substrate for betaI PKC, at the phagosomal cup/phagosome was apparent during BIgG ingestion. Superoxide (O(2)(-)) production was profoundly inhibited by Gö6976, a cPKC inhibitor, and dramatically increased by the DGK inhibitor, R59949. Ultrastructural analysis revealed that BIgG induced O(2)(-) production at the phagosome but not at the intracellular granules. We conclude that activation/accumulation of betaI PKC is involved in O(2)(-) production, and that O(2)(-) production is primarily initiated at the phagosomal cup/phagosome. This study also suggests that DGKbeta plays a prominent role in regulation of O(2)(-) production during FcgammaR-mediated phagocytosis.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Cell Line, Transformed
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Diacylglycerol Kinase / genetics
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Diacylglycerol Kinase / metabolism
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Diacylglycerol Kinase / ultrastructure
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Green Fluorescent Proteins
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Humans
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Isoenzymes / genetics
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Isoenzymes / metabolism
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Isoenzymes / physiology
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Isoenzymes / ultrastructure
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Luminescent Proteins / genetics
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Luminescent Proteins / metabolism
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Luminescent Proteins / ultrastructure
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Mice
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Microglia / enzymology*
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Microglia / immunology*
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Microglia / metabolism
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Microglia / ultrastructure
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Microspheres
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Oxidants / biosynthesis
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Phagocytes / enzymology
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Phagocytes / immunology
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Phagocytes / metabolism
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Phagocytes / ultrastructure
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Phagocytosis / genetics
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Phagocytosis / immunology*
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Phagosomes / enzymology
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Phagosomes / genetics
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Phagosomes / metabolism*
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Phagosomes / ultrastructure
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Protein Kinase C / genetics
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Protein Kinase C / metabolism
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Protein Kinase C / physiology*
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Protein Kinase C / ultrastructure
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Protein Kinase C beta
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Protein Kinase C-epsilon
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Receptors, IgG / physiology*
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Superoxides / metabolism*
Substances
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Isoenzymes
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Luminescent Proteins
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Oxidants
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Receptors, IgG
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Superoxides
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Green Fluorescent Proteins
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Prkce protein, mouse
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Diacylglycerol Kinase
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PRKCE protein, human
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Protein Kinase C
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Protein Kinase C beta
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Protein Kinase C-epsilon