Interaction between p22 and Nox4 in the endoplasmic reticulum suggests a unique mechanism of NADPH oxidase complex formation.

The p22 protein is an essential component of the phagocytic- and inner ear NADPH oxidases but its relationship to other Nox proteins is less clear. We have studied the role of p22 in the TGF-β1-stimulated HO production of primary human and murine fibroblasts. TGF-β1 induced HO release of the examined cells, and the response was dependent on the expression of both Nox4 and p22. Interestingly, the p22 protein was present in the absence of any detectable Nox/Duox expression, and the p22 level was unaffected by TGF-β1. On the other hand, Nox4 expression was dependent on the presence of p22, establishing an asymmetrical relationship between the two proteins. Nox4 and p22 proteins localized to the endoplasmic reticulum and their distribution was unaffected by TGF-β1. We used a chemically induced protein dimerization method to study the orientation of p22 and Nox4 in the endoplasmic reticulum membrane. This technique is based on the rapamycin-mediated heterodimerization of the mammalian FRB domain with the FK506 binding protein. The results of these experiments suggest that the enzyme complex produces HO into the lumen of the endoplasmic reticulum, indicating that Nox4 contributes to the development of the oxidative milieu within this organelle.