Mitochondria have a major role in energy production via oxidative phosphorylation, which is dependent on the expression of critical genes encoded by mitochondrial (mt)
DNA. Mutations in
mtDNA can cause fatal or severely debilitating disorders with limited treatment options. Clinical manifestations vary based on mutation type and heteroplasmy (that is, the relative levels of mutant and wild-type
mtDNA within each cell). Here we generated genetically corrected pluripotent stem cells (PSCs) from patients with
mtDNA disease. Multiple induced pluripotent stem (iPS) cell lines were derived from patients with common heteroplasmic mutations including 3243A>G, causing
mitochondrial encephalomyopathy and
stroke-like episodes (
MELAS), and 8993T>G and 13513G>A, implicated in
Leigh syndrome. Isogenic
MELAS and
Leigh syndrome iPS cell lines were generated containing exclusively wild-type or mutant
mtDNA through spontaneous segregation of heteroplasmic
mtDNA in proliferating fibroblasts. Furthermore, somatic cell nuclear transfer (SCNT) enabled replacement of mutant
mtDNA from homoplasmic 8993T>G fibroblasts to generate corrected Leigh-NT1 PSCs. Although Leigh-NT1 PSCs contained donor oocyte wild-type
mtDNA (human haplotype D4a) that differed from
Leigh syndrome patient haplotype (F1a) at a total of 47
nucleotide sites, Leigh-NT1 cells displayed transcriptomic profiles similar to those in embryo-derived PSCs carrying wild-type
mtDNA, indicative of normal nuclear-to-mitochondrial interactions. Moreover, genetically rescued patient PSCs displayed normal metabolic function compared to impaired oxygen consumption and
ATP production observed in mutant cells. We conclude that both reprogramming approaches offer complementary strategies for derivation of PSCs containing exclusively wild-type
mtDNA, through spontaneous segregation of heteroplasmic
mtDNA in individual iPS cell lines or mitochondrial replacement by SCNT in homoplasmic
mtDNA-based disease.