Widespread Mitochondrial Depletion via Mitophagy Does Not Compromise Necroptosis

Programmed necrosis (or necroptosis) is a form of cell death triggered by the activation of receptor interacting protein kinase-3 (RIPK3). Several reports have implicated mitochondria and mitochondrial reactive oxygen species (ROS) generation as effectors of RIPK3-dependent cell death. Here, we directly test this idea by employing a method for the specific removal of mitochondria via mitophagy. Mitochondria-deficient cells were resistant to the mitochondrial pathway of apoptosis, but efficiently died via tumor necrosis factor (TNF)-induced, RIPK3-dependent programmed necrosis or as a result of direct oligomerization of RIPK3. Although the ROS scavenger butylated hydroxyanisole (BHA) delayed TNF-induced necroptosis, it had no effect on necroptosis induced by RIPK3 oligomerization. Furthermore, although TNF-induced ROS production was dependent on mitochondria, the inhibition of TNF-induced necroptosis by BHA was observed in mitochondria-depleted cells. Our data indicate that mitochondrial ROS production accompanies, but does not cause, RIPK3-dependent necroptotic cell death. [Display omitted] •Parkin-induced mitochondrial depletion can be used to define mitochondrial functions•Mitochondrial depletion prevents TNF-induced ROS, but not necroptosis•Activated RIPK3-induced necroptosis proceeds normally in mitochondria-depleted cells•ROS scavenger BHA delays TNF-induced necroptosis in mitochondria-depleted cells Mitochondria have been implicated as key effectors in the execution of RIPK3-dependent necroptosis. Tait, Green, and colleagues directly test the importance of mitochondria in necroptosis by utilizing mitophagy to generate mitochondria-deficient cells. Mitochondrial depletion inhibits ROS production, but it does not affect necroptosis induced either by TNF or through direct activation of RIPK3. These results indicate that mitochondrial ROS accompanies rather than causes necroptosis and argue against a major role for mitochondria in executing necroptotic cell death.