Mitochondrial ATP synthase is a target for TNBS-induced protein carbonylation in XS-106 dendritic cells.

ROS are produced in dendritic cells (DCs) during antigen presentation in contact hypersensitivity (CHS). As a result, ROS cause a number of nonenzymatic protein modifications, including carbonylation, which is the most widely used marker of oxidative stress. 2,4,6-Trinitrobenzene sulfonic acid (TNBS) is a well-characterized contact allergen that results in the formation of ROS. However, proteins that are carbonylated in DCs in response to TNBS have not been identified. To study ROS-dependent protein carbonylation in response to TNBS, we used the well-established mouse DC line, XS-106. We focused on the effects of TNBS on oxidation by examining selected oxidative markers. We identified TNBS-induced ROS and myeloperoxidase (MPO) proteins and demonstrated that the increase in ROS resulted in IL-12 production. The increase in oxidation was further confirmed by an oxidation-dependent increase in protein modifications, such as carbonylation. In fact, TNBS strongly induced carbonylation of mitochondrial adenosine triphosphate (ATP) synthase in XS-106 DCs, as determined by MALDI-TOF analysis and 2-D Western blotting. ROS production and protein carbonylation were confirmed in human monocyte-derived DCs (Mo-DCs). Furthermore, glutathione (GSH) decreased ROS and protein carbonylation in Mo-DCs. Carbonylation of ATP synthase in DCs may contribute to the pathophysiology of CHS.