Disease-causing mitochondrial heteroplasmy segregated within induced pluripotent stem cell clones derived from a patient with MELAS.

Mitochondrial diseasesdisplay pathological phenotypes according to the mixture of mutant versus wild-type mitochondrial DNA (mtDNA), known as heteroplasmy. We herein examined the impact of nuclear reprogrammingand clonal isolation of induced pluripotent stem cells(iPSC) on mitochondrial heteroplasmy. Patient-derived dermal fibroblastswith a prototypical mitochondrial deficiency diagnosed as MELAS demonstrated mitochondrial dysfunction with reduced oxidative reserve due to heteroplasmyat position G13513A in the ND5 subunit of complex I. Bioengineered iPSC clones acquired pluripotency with multi-lineage differentiation capacity and demonstrated reduction in mitochondrial density and oxygen consumption distinguishing them from the somatic source. Consistent with the cellular mosaicismof the original patient-derived fibroblasts, the MELAS-iPSC clones contained a similar range of mtDNA heteroplasmyof the disease-causing mutation with identical profiles in the remaining mtDNA. High- heteroplasmyiPSC clones were used to demonstrate that extended stem cell passaging was sufficient to purge mutant mtDNA, resulting in isogenic iPSC subclones with various degrees of disease-causing genotypes. Upon comparative differentiation of iPSC clones, improved cardiogenic yield was associated with iPSC clones containing lower heteroplasmycompared to isogenic clones with high heteroplasmy. Thus, mtDNA heteroplasmic segregation within patient-derived stem cell linesenables direct comparison of genotype/phenotype relationships in progenitor cells and lineage-restricted progeny, and indicates that cell fate decisions are regulated as a function of mtDNA mutation load. The novel nuclear reprogramming-based model system introduces a disease-in-a-dish tool to examine the impact of mutant genotypes for MELAS patients in bioengineered tissues and a cellular probe for molecular features of individual mitochondrial diseases.