A small molecule inhibitor Mirin prevents TOP3A-dependent mtDNA breakage and segregation

Mirin, the chemical inhibitor of MRE11, has been recently reported to prevent immune response activation caused by mitochondrial DNA (mtDNA) breakage and release upon replication stalling. We show here that Mirin prevents mitochondrial replication fork breakage in mitochondrial 3’-exonuclease MGME1 deficient cells and the resulting innate immune response induction, but that this occurs independently of MRE11. Furthermore, Mirin also caused alteration of mtDNA supercoiling and accumulation of hemicatenated replication termination intermediates, hallmarks of topoisomerase dysfunction, as well as alleviated topological changes induced by the overexpression of mitochondrial TOP3A, including TOP3A-dependent strand breakage at the non-coding region of mtDNA, potentially explaining its protective effect in the MGME1-knockout cells. Although Mirin does not inhibit TOP3A in vitro , our results demonstrate its MRE11-independent effects in cells and give insight into the mechanisms of mtDNA segregation, as well as the maintenance of genomic integrity in mitochondria. Significance Statement ### Competing Interest Statement The authors have declared no competing interest. * 2D-AGE : Two-dimensional agarose gel electrophoresis BTR complex : BLM-TOP3A-RMI1-RMI2 complex DAMP : Damage-associated molecular pattern DSB : Double-strand break HR : Homologous recombination mtDNA : Mitochondrial DNA KO : Knockout NCR : Non-coding region (of mtDNA) OB-fold : Oligonucleotide-binding fold OriH : Origin of heavy-strand replication OriL : Origin of light-strand replication