Widespread Mitochondrial Depletion via Mitophagy Does Not Compromise Necroptosis

• 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

SummaryProgrammed 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.

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