A library of base editors for the precise ablation of all protein-coding genes in the mouse mitochondrial genome.
The development of curative treatments for mitochondrial diseases, which are often caused by mutations in mitochondrial DNA (mtDNA) that impair energy metabolism and other aspects of cellular homoeostasis, is hindered by an incomplete understanding of the underlying biology and a scarcity of cellular and animal models. Here we report the design and application of a library of double-stranded-DNA deaminase-derived cytosine base editors optimized for the precise ablation of every mtDNA protein-coding gene in the mouse mitochondrial genome. We used the library, which we named MitoKO, to produce near-homoplasmic knockout cells in vitro and to generate a mouse knockout with high heteroplasmy levels and no off-target edits. MitoKO should facilitate systematic and comprehensive investigations of mtDNA-related pathways and their impact on organismal homoeostasis, and aid the generation of clinically meaningful in vivo models of mtDNA dysfunction.
Acknowledgements: We acknowledge the members of the Mitochondrial Genetics Group (MRC-MBU, University of Cambridge) for useful discussion during the course of this research. We thank J. Carroll for help in obtaining the data for Fig. 6f. All authors disclose funding support for the research described in this study from Medical Research Council UK (MC_UU_00015/4 and MC_UU_00028/3). P.S-P. discloses support for the research described in this study from The Champ Foundation (G112428). P.A.N. discloses support for the research described in this study from The Lily Foundation (G101554).