Lysosomal uptake of mtDNA mitigates heteroplasmy

Mitochondrial DNA is exposed to multiple insults produced by normal cellular function. Upon mtDNA replication stress the mitochondrial genome transfers to endosomes where it is degraded. Here, using proximity proteomics we found that mtDNA replication stress leads to the rewiring of the mitochondrial proximity proteome, increasing mitochondria association with lysosomal and vesicle-associated proteins, such as the GTPase RAB10 and the retromer. We found that upon mtDNA replication stress, RAB10 enhances mitochondrial fragmentation and relocates from the ER to lysosomes containing mtDNA. The retromer enhances and coordinates the expulsion of mitochondrial matrix components through mitochondrial-derived vesicles, and mtDNA with direct transfer to lysosomes. Using a Drosophila model carrying a long deletion on the mtDNA ({Delta}mtDNA), we evaluated in vivo the role of the retromer in mtDNA extraction and turnover in the larval epidermis. The presence of {Delta}mtDNA elicits the activation of a specific transcriptome profile related to counteract mitochondrial damage. Expression of the retromer component Vps35 is sufficient to restore mtDNA homoplasmy and mitochondrial defects associated with {Delta}mtDNA. Our data reveal novel regulators involved in the specific elimination of mtDNA. We demonstrate that modulation of the retromer in vivo is a successful mechanism to restore mitochondrial function associated with mtDNA damage.