The molecular basis for adhesion-mediated suppression of reactive oxygen species generation by human neutrophils

Abstract

Human neutrophil adherence to ECMs induces an initial inhibition of stimulated reactive oxygen species (ROS) formation, followed by an enhanced phase of oxidant production. The initial integrin-mediated suppression of ROS constitutes a mechanism to prevent inappropriate tissue damage as leukocytes migrate to inflammatory sites. The Rac2 guanosine 5′-triphosphatase (GTPase) is a critical regulatory component of the phagocyte NADPH oxidase. We show that activation of Rac2 is inhibited in adherent neutrophils, correlating with inhibition of ROS formation. Conversely, NADPH oxidase components p47 and p67 assemble normally, suggesting a specific action of adhesion on the Rac2 molecular switch. Reconstitution with activated Rac2 restored rapid NADPH oxidase activation kinetics to adherent neutrophils, establishing that inhibition was due to defective Rac2 activity. We provide evidence that integrins inhibit Rac2 activation via a membrane-associated guanine nucleotide exchange factor, likely to be Vav1. Activation of Vav1, but not its upstream activator, Syk, is suppressed by cell adhesion. Vav1 activity is inhibited due to dephosphorylation of the regulatory Tyr174 via enhanced tyrosine phosphatase activity in adherent cells. These studies identify an integrin-mediated pathway in which Vav1 is as a strong candidate for the critical regulatory point in suppression of Rac2 activation and ROS generation during inflammatory responses

Involvement of Rac1 in Activation of Multicomponent Nox1- and Nox3-Based NADPH Oxidases

“…Following cellular activation, these components assemble into the active, membrane-bound complex through distinct, but coordinated, signaling events. The flavocytochrome translocates to specific membrane domains; the cytosolic components are phosphorylated, thereby inducing conformational changes that favor interactions with the cytochrome b 558 ; and Rac dissociates from RhoGDI and translocates independently to the membrane following exchange of GDP for GTP (26,60). Extensive studies of cell-free oxidase reconstitution suggest that Rac can serve two functions in supporting the Nox2 complex: tethering p67…”

Blockade of Class IA Phosphoinositide 3-Kinase in Neutrophils Prevents NADPH Oxidase Activation- and Adhesion-dependent Inflammation

“…Inhibition of p47 phox phosphorylation markedly decreased the membrane translocation of p47 phox , association of p47 phox with Nox2 at the membrane, and activation of NADPH oxidase-induced oxidant generation (44). In addition, activation of the small GTPase Rac2, a prerequisite for NADPH oxidase assembly (35,45), was shown to be dependent on PI3K activity (46 -48).…”

Integrin α1β1 Controls Reactive Oxygen Species Synthesis by Negatively Regulating Epidermal Growth Factor Receptor-Mediated Rac Activation

“…In contrast to this finding, adherent leukocytes transiently suppress NADPH oxidase activity and ROS production (59). This effect is due to integrin-mediated inhibition of Rac2 as a result of enhanced tyrosine phosphatase activity and decreased Vav1 activation (59). All together, these data clearly suggest that integrin activation can either enhance or inhibit ROS production and this effect is cell and tissue specific.…”

“…In addition, activation of integrin ␣2␤1 induces NADPH oxidase-mediated ROS production in a p38-dependent manner (21). In contrast to this finding, adherent leukocytes transiently suppress NADPH oxidase activity and ROS production (59). This effect is due to integrin-mediated inhibition of Rac2 as a result of enhanced tyrosine phosphatase activity and decreased Vav1 activation (59).…”