NADPH oxidase subunit p22phox: A marker of oxidase-dependent oxidative stress and target for stress suppression in nonphagocytic cells.

Reactive oxygen species (ROS)-producing NADPH oxidase (Nox) family proteins are involved in host defense. The overproduction of ROS leads to oxidative stress, which is associated with a myriad of diseases. The Nox subunit p22phox is essential for Nox1-4 activity, and p22phox and Nox2 proteins are mutually stabilized in phagocytic cells. This study investigated the suitability of p22phox protein as a marker of Nox activity. To avoid contamination by phagocytic p22phox, we developed global Cybb (encoding Nox2)-knockout mice and analyzed p22phox stability and the expression profiles of Nox proteins in lysates of various tissues. We found that consistent with Nox2 in phagocytic cells, p22phox protein was detected when Nox1-4 were coexpressed in nonphagocytic cells. Furthermore, p22phox protein degradation was suppressed by Nox1-4, suggesting that p22phox is a suitable marker of Nox family-dependent oxidative stress. Thus, we examined p22phox protein levels in tissue lysates prepared from Cybb-knockout mice to avoid the contamination of phagocytic p22phox. Cybb-knockout mice show moderately reduced p22phox protein levels in lung tissue, suggesting that Nox2 and other Nox family members stabilized p22phox protein. Paradoxically, this result implied that p22phox knockdown concurrently suppressed various Nox family-dependent oxidative stress mechanisms, and this was confirmed by the suppression of Nox family-dependent directed migration in p22phox-knockdown A549 human lung epithelial cells. Therefore, p22phox not only served as a marker of Nox-dependent oxidative stress but also as a target to suppress this stress in various tissues and cells.