Linear DNA-driven recombination in mammalian mitochondria

Abstract

Mitochondrial DNA (mtDNA) recombination in animals has remained enigmatic due to its uniparental inheritance and subsequent homoplasmic state, which excludes the biological need for genetic recombination, as well as limits tools to study it. However, molecular recombination is an important genome maintenance mechanism for all organisms, most notably being required for double-strand break repair. To demonstrate the existence of mtDNA recombination, we took advantage of a cell model with two different types of mitochondrial genomes and impaired its ability to degrade broken mtDNA. The resulting excess of linear DNA fragments caused increased formation of cruciform mtDNA, appearance of heterodimeric mtDNA complexes and recombinant mtDNA genomes, detectable by Southern blot and by long range PacBio® HiFi sequencing approach. Besides utilizing different electrophoretic methods, we also directly observed molecular complexes between different mtDNA haplotypes and recombination intermediates using transmission electron microscopy. We propose that the known copy-choice recombination by mitochondrial replisome could be sufficient for the needs of the small genome, thus removing the requirement for a specialized mitochondrial recombinase. The error-proneness of this system is likely to contribute to the formation of pathological mtDNA rearrangements.

Absence of both MGME1 and POLG EXO abolishes mtDNA whereas absence of either creates unique mtDNA duplications

“…A , frequency of duplication sizes found in Polg MM and Mgme1 KK heart mtDNA. B , mapping of mtDNA breakpoint start and end sequences superimposed onto a map of the mtDNA molecule by position ( 14 , 15 , 16 , 17 ). C , mtDNA duplications plots as predicted by mitoSALT analysis pipeline.…”

“…Currently, there is no consensus on the presence and mechanisms of mammalian mtDNA repair, particularly after DSB. However, recent reports suggest that the mtDNA replisome can mediate recombination of free DNA ends ( 15 ). Thus, the action of two replisome exonucleases, POLG EXO and MGME1, in eliminating broken mtDNA after DSB and their role in mtDNA maintenance has come into focus ( 7 , 8 , 10 , 11 ).…”