Mitochondrial protein-linked DNA breaks perturb mitochondrial gene transcription and trigger free radical-induced DNA damage

Breakage of one strand of DNA is the most common form of DNA damage. Most damaged DNA termini require end processing in preparation for ligation. The importance of this step is highlighted by the association of defects in the 3'-end processing enzyme tyrosyl DNA phosphodiesterase 1 (TDP1) and neurodegeneration, and by the cytotoxic induction of protein-linked DNA breaks (PDBs) and oxidised nucleic acid intermediates during chemo- and radiotherapy. Although much is known about the repair of PDBs in the nucleus, little is known about this process in the mitochondria. Here, we reveal that TDP1 resolves mitochondrial PDBs (mtPDBs), thereby promoting mitochondrial gene transcription. In the absence of TDP1, the imbalance in transcription of mitochondrial and nuclear encoded electron transport chain (ETC) subunits results in misassembly of the ETC complex III. Bioenergetics profiling further reveals that TDP1 promotes oxidative phosphorylation under both basal and high ATP demands. Mitochondrial dysfunction results in free radical leakage and nuclear DNA damage. Consequently, we report an increased accumulation of carbon-centred radicals in cells lacking TDP1, using electron spin resonance spectroscopy. Overexpression of the anti-oxidant enzyme superoxide dismutase 1 (SOD1) reduces carbon-centred adducts and protects TDP1 deficient cells from oxidative stress. Conversely, overexpression of the amyotrophic lateral sclerosis (ALS)-associated mutant SOD1G93A leads to marked sensitivity. Together, this data characterises a novel TDP1 driven mitochondrial PDB repair process and unravels its role in promoting mitochondrial gene transcription and oxygen consumption by oxidative phosphorylation, thus conferring cellular protection against ROS induced damage.