Mutations in the
mitochondrial DNA (
mtDNA) can cause a variety of human diseases. In most cases, such mutations are heteroplasmic (i.e. mutated and wild-type
mtDNA coexist) and a small percentage of wild-type sequences can have a strong protective effect against a metabolic defect. Because a genetic approach to correct
mtDNA mutations is not currently available, the ability to modulate heteroplasmy would have a major impact in the phenotype of many patients with
mitochondrial disorders. We show here that a
restriction endonuclease targeted to mitochondria has this ability. A mitochondrially targeted PstI degraded
mtDNA harboring PstI sites, in some cases leading to a complete loss of mitochondrial genomes. Recombination between
DNA ends released by PstI was not observed. When expressed in a heteroplasmic rodent cell line, containing one
mtDNA haplotype with two sites for PstI and another haplotype having none, the mitochondrial PstI caused a significant shift in heteroplasmy, with an accumulation of the
mtDNA haplotype lacking PstI sites. These experiments provide proof of the principle that
restriction endonucleases are feasible tools for genetic therapy of a sub-group of
mitochondrial disorders. Although this approach is limited by the presence of mutation-specific restriction sites, patients with
neuropathy, ataxia and retinitis pigmentosa (NARP) could benefit from it, as the T8399G mutation creates a unique restriction site that is not present in wild-type human
mitochondrial DNA.