NADPH oxidase controls phagosomal pH and antigen cross-presentation in human dendritic cells

Abstract

The phagocyte NADPH oxidase (NOX2) is critical for the bactericidal activity of phagocytic cells and plays a major role in innate immunity. We showed recently that NOX2 activity in mouse dendritic cells (DCs) prevents acidification of phagosomes, promoting antigen cross-presentation. In order to investigate the role of NOX2 in the regulation of the phagosomal pH in human DCs, we analyzed the production of reactive oxygen species (ROS) and the phagosomal/endosomal pH in monocytederived DCs and macrophages (MØs) from healthy donors or patients with chronic granulomatous disease (CGD). As expected, we found that human MØs acidify their phagosomes more efficiently than human DCs. Accordingly, the expression of the vacuolar proton ATPase (V-H ؉ -ATPase) was higher in MØs than in DCs. Phagosomal ROS production, however, was also higher in MØs than in DCs, due to higher levels of gp91phox expression and recruitment to phagosomes. In contrast, in the absence of active NOX2, the phagosomal and endosomal pH decreased. Both in the presence of a NOX2 inhibitor and in DCs derived from patients with CGD, the cross-presentation of 2 model tumor antigens was impaired. We conclude that NOX2 activity participates in the regulation of the phagosomal and endosomal pH in human DCs, and is required for efficient antigen crosspresentation. (Blood. 2008;112:4712-4722)

Lupus-associated causal mutation in neutrophil cytosolic factor 2 (NCF2) brings unique insights to the structure and function of NADPH oxidase

“…However, in other immune cells (e.g., antigen-presenting cells), ROS production is much more limited (26)(27)(28) and the role of ROS is not microbicidal. NADPH oxidase activity regulates phagosomal pH, and derivative ROS can function as signaling molecules within and between neighboring immune cells; reduced levels of ROS in these cells may thus influence antigen processing, immunoregulation, control of cell activation, and differentiation (28,29).…”

Receptor Signaling Lymphocyte-activation Molecule Family 1 (Slamf1) Regulates Membrane Fusion and NADPH Oxidase 2 (NOX2) Activity by Recruiting a Beclin-1/Vps34/Ultraviolet Radiation Resistance-associated Gene (UVRAG) Complex

“…Phagosome Isolation-Transfectant RAW264.7 cells were incubated with E. coli-containing magnetic particles for 90 min and then were disrupted in homogenization buffer (0.25 M sucrose, 10 mM HEPES, 3 mM MgCl 2 ⅐6H 2 O, EDTA-free protease inhibitor mixture) as described previously (11). Phagosomes were removed from the homogenate using a magnet and were washed three times in cold PBS.…”

NADPH Oxidase Modifies Patterns of MHC Class II–Restricted Epitopic Repertoires through Redox Control of Antigen Processing

“…It was recently discovered that the antimicrobial effector NADPH oxidase (NOX2) negatively regulates the levels of proteolysis within the maturing phagosome of macrophages (4,5) and DCs (3,6,7). NOX2, a multiprotein complex expressed in phagocytes, is rapidly assembled on the early phagosomal membrane where it oxidizes cytosolic NADPH to convert molecular oxygen to superoxide within the phagosomal lumen.…”