alexa Inhibition of CD45 during neutrophil activation.
Immunology

Immunology

Journal of Clinical & Cellular Immunology

Author(s): Fialkow L, Chan CK, Downey GP

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Abstract Tyrosine phosphorylation represents a balance between the activity of tyrosine kinases and phosphatases. We have demonstrated recently that reactive oxygen intermediates (ROI) produced by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enhance tyrosine phosphorylation in neutrophils. As tyrosine phosphatase activity can be regulated by oxidants, we sought to determine whether endogenously generated ROI inhibited the activity of the leukocyte tyrosine phosphatase CD45. Addition of guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) to electropermeabilized neutrophils, conditions known to activate the oxidase, inhibited CD45, as determined by immunoprecipitation and an in vitro phosphatase assay. That this inhibition was a consequence of activation of the oxidase was supported by three observations: 1) GTPgammaS-induced inhibition of CD45 was NADPH dependent; 2) pretreatment of cells with diphenylene iodonium, an oxidase inhibitor, partially prevented the inhibition; and 3) inhibition of CD45 was diminished markedly in neutrophils from chronic granulomatous disease (CGD) patients. The inhibition could be partially prevented by treatment of the cells with the antioxidants N-acetylcysteine or DTT, but direct antioxidant treatment of CD45 immunoprecipitates could not restore activity. Exposure to PMA, a direct activator of protein kinase C that also induces an oxidative burst, inhibited CD45 in both normal and CGD neutrophils. However, the magnitude of inhibition was less and the kinetics delayed in CGD cells when compared with normal cells. We conclude that ROI produced by the NADPH oxidase can contribute to inhibition of tyrosine phosphatases such as CD45 by oxidant-mediated effects, but that alternate regulatory mechanisms also exist.
This article was published in J Immunol and referenced in Journal of Clinical & Cellular Immunology

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