The Role of Mitochondrial DNA Replication Fidelity in Toxicity Initiated by Drugs that Impair Mitochondrial Function

Mitochondrial DNA (mtDNA) is crucial for proper mitochondrial function in part because it encodes for 13 subunits of the electron transport chain (ETC). mtDNA is replicated exclusively by DNA polymerase gamma (Polg), which has an exonuclease proof-reading domain critical for maintaining mtDNA replication fidelity. Drug-induced mitochondrial toxicity is a common problem that can occur by many mechanisms, including ETC inhibition and mtDNA damage. Gene-environment interactions can modulate mitochondrial toxicity, and individuals with POLG mutations are at risk for toxicity from valproic acid, a drug that impairs mitochondrial function. We hypothesize that mtDNA replication fidelity protects against toxicity initiated by drugs that impair mitochondrial function. I observed that mouse embryonic fibroblasts (MEFs) with impaired polymerase gamma exonuclease activity (Polgm/m) and increased mtDNA mutation load have increased susceptibility to toxicity caused by drugs that impair mitochondrial function. The ETC complex I inhibitor rotenone caused increased apoptosis in Polgm/m MEFs relative to wild type. This sensitivity was also observed for other ETC complex inhibitors and clinical drugs with off-target ETC complex I inhibition. A mitochondria-penetrating peptide (MPP) was used to target the alkylating agent chlorambucil (Cbl) specifically to mitochondria. Mitochondria-targeted Cbl (mt-Cbl) caused bioenergetic differences between Polgm/m and wild type MEFs, but was not differentially toxic at the level of cell viability. Increasing mitochondrial autophagy with rapamycin attenuated rotenone toxicity in Polgm/m MEFs, while changes to mitochondrial fusion/fission dynamics did not alter toxicity. Overall, these data suggest that mtDNA replication fidelity maintained by polymerase gamma exonuclease activity protects against bioenergetic stress caused by ETC inhibition.Ph