Send to

Choose Destination
Nucleic Acids Res. 2016 Sep 19;44(16):7804-16. doi: 10.1093/nar/gkw676. Epub 2016 Jul 27.

Near-complete elimination of mutant mtDNA by iterative or dynamic dose-controlled treatment with mtZFNs.

Author information

MRC Mitochondrial Biology Unit, Cambridge, UK
MRC Cancer Unit, Cambridge, UK.
MRC Mitochondrial Biology Unit, Cambridge, UK.
MRC Mitochondrial Biology Unit, Cambridge, UK GABBA, University of Porto, Portugal.
JDRF/Wellcome Trust DIL, Cambridge Institute for Medical Research, University of Cambridge, UK.
INSERM U1154, CNRS UMR 7196, Muséum National d'Histoire Naturelle, Paris, France.
MRC Mitochondrial Biology Unit, Cambridge, UK


Mitochondrial diseases are frequently associated with mutations in mitochondrial DNA (mtDNA). In most cases, mutant and wild-type mtDNAs coexist, resulting in heteroplasmy. The selective elimination of mutant mtDNA, and consequent enrichment of wild-type mtDNA, can rescue pathological phenotypes in heteroplasmic cells. Use of the mitochondrially targeted zinc finger-nuclease (mtZFN) results in degradation of mutant mtDNA through site-specific DNA cleavage. Here, we describe a substantial enhancement of our previous mtZFN-based approaches to targeting mtDNA, allowing near-complete directional shifts of mtDNA heteroplasmy, either by iterative treatment or through finely controlled expression of mtZFN, which limits off-target catalysis and undesired mtDNA copy number depletion. To demonstrate the utility of this improved approach, we generated an isogenic distribution of heteroplasmic cells with variable mtDNA mutant level from the same parental source without clonal selection. Analysis of these populations demonstrated an altered metabolic signature in cells harbouring decreased levels of mutant m.8993T>G mtDNA, associated with neuropathy, ataxia, and retinitis pigmentosa (NARP). We conclude that mtZFN-based approaches offer means for mtDNA heteroplasmy manipulation in basic research, and may provide a strategy for therapeutic intervention in selected mitochondrial diseases.

[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Silverchair Information Systems Icon for PubMed Central
Loading ...
Support Center