Human Molecular Genetics Advance Access originally published online on August 18, 2004
Human Molecular Genetics 2004 13(20):2519-2534; doi:10.1093/hmg/ddh267
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Human Molecular Genetics, Vol. 13, No. 20 © Oxford University Press 2004; all rights reserved
Nuclear DNA-encoded tRNAs targeted into mitochondria can rescue a mitochondrial DNA mutation associated with the MERRF syndrome in cultured human cells
1CNRS-FRE 2375, Institute of Physiology and Biological Chemistry, Université Louis Pasteur, 21 René Descartes, 67084 Strasbourg, France, 2Department of Molecular Biology, Biology Faculty, Moscow State University, 119992 Moscow, Russia and 3Mitochondrial Research Group, Medical School, Neurology Department, University of Newcastle upon Tyne, Framlington Place, Newcastle upon Tyne NE2 4HH, UK
Received June 28, 2004; Revised July 29, 2004; Accepted August 4, 2004
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 tRNALys 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 tRNALys 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 tRNALys 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 tRNALys 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.
* To whom correspondence should be addressed: Tel: +33 390241481; Fax: +33 388417070; Email: i.tarassov{at}ibmc.u-strasbg.fr
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