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Tissue-specific mitochondrial heteroplasmy at position 16,093 within the same individual.

ABSTRACT: Human mitochondrial DNA (mtDNA) research has entered a massively parallel sequencing (MPS) era, providing deep insight into mtDNA genomics and molecular diagnostics. Analysis can simultaneously include coding and control regions, many samples can be studied in parallel, and even minor heteroplasmic changes can be detected. We investigated heteroplasmy using 16 different tissues from three unrelated males aged 40-54 years at the time of death. mtDNA was enriched using two independent overlapping long-range PCR amplicons and analysed by employing illumina paired-end sequencing. Point mutation heteroplasmy at position 16,093 (m.16093T > C) in the non-coding regulatory region showed great variability among one of the studied individuals; heteroplasmy extended from 5.1 % in red bone marrow to 62.0 % in the bladder. Red (5.1 %) and yellow bone marrow (8.9 %) clustered into one group and two arteries and two aortas from different locations into another (31.2-50.9 %), giving an ontogenetic explanation for the formation of somatic mitochondrial heteroplasmy. Our results demonstrate that multi-tissue screening using MPS provides surprising data even when there is a limited number (3) of study subjects and they give reason to speculate that mtDNA heteroplasmic frequency, distribution, and even its possible role in complex diseases or phenotypes seem to be underestimated.

SUBMITTER: Krjutskov K

PROVIDER: S-EPMC3895442 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Mitochondrial diseases display pathological phenotypes according to the mixture of mutant versus wild-type mitochondrial DNA (mtDNA), known as heteroplasmy. We herein examined the impact of nuclear reprogramming and clonal isolation of induced pluripotent stem cells (iPSC) on mitochondrial heteroplasmy. Patient-derived dermal fibroblasts with a prototypical mitochondrial deficiency diagnosed as mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS) demonstrated mitochondrial dysfunction with reduced oxidative reserve due to heteroplasmy at position G13513A in the ND5 subunit of complex I. Bioengineered iPSC clones acquired pluripotency with multilineage differentiation capacity and demonstrated reduction in mitochondrial density and oxygen consumption distinguishing them from the somatic source. Consistent with the cellular mosaicism of the original patient-derived fibroblasts, the MELAS-iPSC clones contained a similar range of mtDNA heteroplasmy of the disease-causing mutation with identical profiles in the remaining mtDNA. High-heteroplasmy iPSC 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. On comparative differentiation of iPSC clones, improved cardiogenic yield was associated with iPSC clones containing lower heteroplasmy compared with isogenic clones with high heteroplasmy. Thus, mtDNA heteroplasmic segregation within patient-derived stem cell lines enables 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.

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Tissue-specific mitochondrial heteroplasmy at position 16,093 within the same individual.

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