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Nucleic Acids Res. 2016 Nov 2;44(19):9342-9357. Epub 2016 Sep 4.

Stable nuclear expression of ATP8 and ATP6 genes rescues a mtDNA Complex V null mutant.

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SENS Research Foundation Research Center, Mountain View, CA 94041, USA
SENS Research Foundation Research Center, Mountain View, CA 94041, USA.
Keck School of Medicine of USC, Los Angeles, CA 90033, USA.
Buck Institute for Research on Aging, Novato, CA 94945, USA.
Department of Biomedical Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
SENS Research Foundation Research Center, Mountain View, CA 94041, USA


We explore the possibility of re-engineering mitochondrial genes and expressing them from the nucleus as an approach to rescue defects arising from mitochondrial DNA mutations. We have used a patient cybrid cell line with a single point mutation in the overlap region of the ATP8 and ATP6 genes of the human mitochondrial genome. These cells are null for the ATP8 protein, have significantly lowered ATP6 protein levels and no Complex V function. Nuclear expression of only the ATP8 gene with the ATP5G1 mitochondrial targeting sequence appended restored viability on Krebs cycle substrates and ATP synthesis capabilities but, failed to restore ATP hydrolysis and was insensitive to various inhibitors of oxidative phosphorylation. Co-expressing both ATP8 and ATP6 genes under similar conditions resulted in stable protein expression leading to successful integration into Complex V of the oxidative phosphorylation machinery. Tests for ATP hydrolysis / synthesis, oxygen consumption, glycolytic metabolism and viability all indicate a significant functional rescue of the mutant phenotype (including re-assembly of Complex V) following stable co-expression of ATP8 and ATP6 Thus, we report the stable allotopic expression, import and function of two mitochondria encoded genes, ATP8 and ATP6, resulting in simultaneous rescue of the loss of both mitochondrial proteins.

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