Reversible glutathionylation of complex I increases mitochondrial superoxide formation.

Increased production of reactive oxygen species (ROS) by mitochondria is involved in oxidative damage to the organelle and in committing cells to apoptosis or senescence, but the mechanisms of this increase are unknown. Here we show that ROS production by mitochondrial complex I increases in response to oxidation of the mitochondrial glutathione pool. This correlates with thiols on the 51- and 75-kDa subunits of complex I forming mixed disulfides with glutathione. Glutathionylation of complex I increases superoxide production by the complex, and when the mixed disulfides are reduced, superoxide production returns to basal levels. Within intact mitochondria oxidation of the glutathione pool to glutathione disulfide also leads to glutathionylation of complex I, which correlates with increased superoxide formation. In this case, most of this superoxide is converted to hydrogen peroxide, which can then diffuse into the cytoplasm. This mechanism of reversible mitochondrial ROS production suggests how mitochondria might regulate redox signaling and shows how oxidation of the mitochondrial glutathione pool could contribute to the pathological changes that occur to mitochondria during oxidative stress.