The phosphoinositide-binding protein p40 phox activates the NADPH oxidase during FcγIIA receptor-induced phagocytosis

Chang Il Suh, Natalie D. Stull, Jun Li Xing, Wei Tian, Marianne O. Price, Sergio Grinstein, Michael B. Yaffe, Simon Atkinson, Mary C. Dinauer

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112 Scopus citations

Abstract

Superoxide produced by the phagocyte reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is essential for host defense. Enzyme activation requires translocation of p67 phox, p47 phox, and Rac-GTP to flavocytochrome b 558 in phagocyte membranes. To examine the regulation of phagocytosis-induced superoxide production, flavocytochrome b 558, p47 phox, p67 phox, and the FcγIIA receptor were expressed from stable transgenes in COS7 cells. The resulting COS phoxFcγR cells produce high levels of superoxide when stimulated with phorbol ester and efficiently ingest immunoglobulin (Ig)G-coated erythrocytes, but phagocytosis did not activate the NADPH oxidase. COS7 cells lack p40 phox, whose role in the NADPH oxidase is poorly understood. p40 phox contains SH3 and phagocyte oxidase and Bem1p (PB1) domains that can mediate binding to p47 phox and p67 phox, respectively, along with a PX domain that binds to phosphatidylinositol-3- phosphate (PI(3)P), which is generated in phagosomal membranes. Expression of p40 phox was sufficient to activate superoxide production in COS phoxFcγR phagosomes. FcγIIA-stimulated NADPH oxidase activity was abrogated by point mutations in p40 phox that disrupt PI(3)P binding, or by simultaneous mutations in the SH3 and PB1 domains. Consistent with an essential role for PI(3)P in regulating the oxidase complex, phagosome NADPH oxidase activation in primary macrophages ingesting IgG-coated beads was inhibited by phosphatidylinositol 3 kinase inhibitors to a much greater extent than phagocytosis itself. Hence, this study identifies a role for p40 phox and PI(3)P in coupling FcγR-mediated phagocytosis to activation of the NADPH oxidase. JEM

Original languageEnglish
Pages (from-to)1915-1925
Number of pages11
JournalJournal of Experimental Medicine
Volume203
Issue number8
DOIs
StatePublished - Aug 7 2006
Externally publishedYes

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    Suh, C. I., Stull, N. D., Xing, J. L., Tian, W., Price, M. O., Grinstein, S., Yaffe, M. B., Atkinson, S., & Dinauer, M. C. (2006). The phosphoinositide-binding protein p40 phox activates the NADPH oxidase during FcγIIA receptor-induced phagocytosis. Journal of Experimental Medicine, 203(8), 1915-1925. https://doi.org/10.1084/jem.20052085