Genetic screens have been used extensively to probe interactions between nuclear genes and their impact on phenotypes. Probing interactions between mitochondrial genes and their phenotypic outcome, however, has not been possible due to a lack of tools to map the responsible polymorphisms. Here, using a toolkit we previously established in Drosophila, we isolate over 300 recombinant mitochondrial genomes and map a naturally occurring polymorphism at the cytochrome c oxidase III residue 109 (CoIII109) that fully rescues the lethality and other defects associated with a point mutation in cytochrome c oxidase I (CoIT300I). Through lipidomics profiling, biochemical assays and phenotypic analyses, we show that the CoIII109 polymorphism modulates cardiolipin binding to prevent complex IV instability caused by the CoIT300I mutation. This study demonstrates the feasibility of genetic interaction screens in animal mitochondrial DNA. It unwraps the complex intra-genomic interplays underlying disorders linked to mitochondrial DNA and how they influence disease expression.

Cambridge Institute for Medical Research University of Cambridge Cambridge CB2 0XY UK

Department of Genetics University of Cambridge Downing Street Cambridge CB2 3EH UK

Guangdong Provincial Key Laboratory of Protein Function and Regulation in Agricultural Organisms South China Agricultural University Tianhe District 510642 Guangzhou Guangdong P R China

Laboratory of Mitochondrial Physiology Institute of Physiology of the Czech Academy of Sciences Videnska 1083 142 20 Prague Czech Republic

Laverock Therapeutics Stevenage Bioscience Catalyst Gunnels Wood Road Stevenage SG1 2FX UK

School of Biosciences University of Birmingham Birmingham B15 2TT UK

University College London Queen Square Institute of Neurology Royal Free Hospital Rowland Hill Street London NW3 2PF UK

Wellcome Cancer Research UK Gurdon Institute Tennis Court Road Cambridge CB2 1QN UK

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