Neutrophil-mediated oxidative burst and host defense are controlled by a Vav-PLCγ2 signaling axis in mice
Daniel B. Graham, Charles M. Robertson, Jhoanne Bautista, Francesca Mascarenhas, M. Julia Diacovo, Vivianne Montgrain, Siu Kit Lam, Viviana Cremasco, W. Michael Dunne, Roberta Faccio, Craig M. Coopersmith, Wojciech Swat
Daniel B. Graham, Charles M. Robertson, Jhoanne Bautista, Francesca Mascarenhas, M. Julia Diacovo, Vivianne Montgrain, Siu Kit Lam, Viviana Cremasco, W. Michael Dunne, Roberta Faccio, Craig M. Coopersmith, Wojciech Swat
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Research Article Immunology

Neutrophil-mediated oxidative burst and host defense are controlled by a Vav-PLCγ2 signaling axis in mice

Abstract

Oxidative burst, a critical antimicrobial mechanism of neutrophils, involves the rapid generation and release of reactive oxygen intermediates (ROIs) by the NADPH oxidase complex. Genetic mutations in an NADPH oxidase subunit, gp91 (also referred to as NOX2), are associated with chronic granulomatous disease (CGD), which is characterized by recurrent and life-threatening microbial infections. To combat such infections, ROIs are produced by neutrophils after stimulation by integrin-dependent adhesion to the ECM in conjunction with stimulation from inflammatory mediators, or microbial components containing pathogen-associated molecular patterns. In this report, we provide genetic evidence that both the Vav family of Rho GTPase guanine nucleotide exchange factors (GEFs) and phospholipase C–γ2 (PLC-γ2) are critical mediators of adhesion-dependent ROI production by neutrophils in mice. We also demonstrated that Vav was critically required for neutrophil-dependent host defense against systemic infection by Staphylococcus aureus and Pseudomonas aeruginosa, 2 common pathogens associated with fatal cases of hospital-acquired pneumonia. We identified a molecular pathway in which Vav GEFs linked integrin-mediated signaling with PLC-γ2 activation, release of intracellular Ca2+ cations, and generation of diacylglycerol to control assembly of the NADPH oxidase complex and ROI production by neutrophils. Taken together, our data indicate that integrin-dependent signals generated during neutrophil adhesion contribute to the activation of NADPH oxidase by a variety of distinct effector pathways, all of which require Vav.

Authors

Daniel B. Graham, Charles M. Robertson, Jhoanne Bautista, Francesca Mascarenhas, M. Julia Diacovo, Vivianne Montgrain, Siu Kit Lam, Viviana Cremasco, W. Michael Dunne, Roberta Faccio, Craig M. Coopersmith, Wojciech Swat

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Figure 1

Adherent neutrophils require Vav to undergo oxidative burst in response to integrin ligands.

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Adherent neutrophils require Vav to undergo oxidative burst in response ...
ROI production in bone marrow–derived neutrophils was measured using lucigenin chemiluminescence. (A and B) WT and VavNULL neutrophils were stimulated in tubes coated with pRGD (A) or anti-CD18 (B). (C) Alternatively, neutrophils were stimulated in fibrinogen-coated tubes (100 μg/ml) with soluble TNF-α (50 ng/ml) (Fgn+TNF). (D) As a positive control, cells were stimulated with PMA (50 ng/ml) for 10–20 minutes or left untreated (Untx), and samples were performed in triplicate. Data shown are representative of at least 5 independent experiments. RLU, relative light units.