Nuclear DNA-encoded tRNAs targeted into mitochondria can rescue a mitochondrial DNA mutation associated with the MERRF syndrome in cultured human cells

Mitochondrial DNA (mtDNA) mutations are an important cause of human disease for which there is no efficient treatment. Our aim was to determine whether the A8344G mitochondrial tRNA(Lys) mutation, which can cause the MERRF (myoclonic epilepsy with ragged-red fibers) syndrome, could be complemented by targeting tRNAs into mitochondria from the cytosol. Import of small RNAs into mitochondria has been demonstrated in many organisms, including protozoans, plants, fungi and animals. Although human mitochondria do not import tRNAs in vivo, we previously demonstrated that some yeast tRNA derivatives can be imported into isolated human mitochondria. We show here that yeast tRNA(Lys) derivatives expressed in immortalized human cells and in primary human fibroblasts are partially imported into mitochondria. Imported tRNAs are correctly aminoacylated and are able to participate in mitochondrial translation. In transmitochondrial cybrid cells and in patient-derived fibroblasts bearing the MERRF mutation, import of tRNA(Lys) is accompanied by a partial rescue of mitochondrial functions affected by the mutation such as mitochondrial translation, activity of respiratory complexes, electrochemical potential across the mitochondrial membrane and respiration rate. Import of a tRNA(Lys) with a mutation in the anticodon preventing recognition of the lysine codons does not lead to any rescue, whereas downregulation of the transgenic tRNAs by small interfering RNA (siRNA) transiently abolishes the functional rescue, showing that this rescue is due to the import. These findings prove for the first time the functionality of imported tRNAs in human mitochondria in vivo and highlight the potential for exploiting the RNA import pathway to treat patients with mtDNA diseases.