Deleterious mutations in RRM2B, a gene responsible for maintenance of mtDNA, result in severe reduction of mtDNA content in skeletal muscle

Mutations in POLG1, DGUOK, TK2, SUCLG1, SUCLA2, PEO1, MPV17 and TYMP can cause mtDNA depletion. Recently, deleterious mutations in RRM2B encoding the p53 inducible ribonucleotide reductase subunit (p53R2) were shown to be associated with severe mtDNA depletion. We report on a patient with severe hypotonia, deafness, blindness, hyperammonemia and increased lactate. The metabolic disorder progressed rapidly and patient died at the age of two months. Oxidative phosphorylation (OXPHOS) enzyme activities were measured using spectrophotometrical analysis. Functional integrity of the five complexes was evaluated using blue native polyacrylamide gel electrophoresis (BN-PAGE) followed by in-gel activity staining. Western blotting was performed to investigate residual protein amounts in the OXPHOS complexes. The amount of mtDNA was evaluated using qPCR. Direct PCR nucleotide sequencing analysis screening of all coding exons and part of flanking introns of RRM2B was used. Severe reduction of activities from partially mtDNA encoded OXPHOS complexes was demonstrated in skeletal muscle using spectrophotometrical analysis and BN-PAGE. Western blotting using antibodies against selective OXPHOS subunits showed that the nuclearly encoded complex II was preserved. The amount of mtDNA in muscle was 3.1 %. Sequencing of RRM2B showed that the patient harboured a homozygous 3-bp in-frame deletion (nt 253-255delGAG, p.Glu85del ) mutation in exon 3. Apart from mutations in enzymes involved in salvage pathway of dNTP’s, also deficient enzymes involved in de novo synthesis can cause mtDNA depletion. In patients with myopathy, primary lactic acidosis and severe mtDNA depletion, screening of RRM2B gene is highly recommended.