Yeast models of human MTO1 variants confirm two diagnoses of mitochondrial modopathy

Introduction: MTO1 is a nuclear gene encoding an enzyme that catalyzes the formation of 5-taurinomethyluridine at position 34 in the anticodon of mt-tRNAs. Pathogenic variants in this gene have been identified in patients with primary mitochondrial diseases associated with the lack of modification of tRNAs, which have been called modopathies (Chujo and Tomizawa, 2021). Here, we analyze four variants that were found in two unrelated patients in compound heterozygosity to confirm pathogenicity: patient 1, harbouring T352M and V272G MTO1 variants, with dystonia, bilateral thalamic changes, high lactate, cardiomyopathy, developmental impairment and nystagmus, whilst patient 2, harbouring A428T and G98S MTO1 variants, with cardiomyopathy, lactic acidosis, myopathy, developmental delay and seizures. MTO1 variants were modeled in Saccharomyces cerevisiae. Material and methods: Spot assay analyses were performed on media supplemented with non-fermentable carbon sources. Oxygen Consumption Rate was measured using an Oxygraph plus. In vivo mitochondrial protein synthesis was carried out for 15 minutes. All the experiments were performed in a mto1Δ strain transformed with wt and mutant mto1 alleles (Ghezzi et al, 2012). Results and discussion: Spot assay: strains equivalent to V272G resulted in a slight decrease of oxidative growth, whereas strains equivalent to T352M and G98S strongly impaired growth. Oxygen Consumption Rate: for the T352M and G98S equivalent strains, measurement revealed a ~70% decrease compared to wt, while A428T and V272G strains result in a 40% and 50% decrease, respectively. In vivo mitochondrial protein synthesis showed a reduction Cox1, Cox2 and Cob protein synthesis in the G98S, V272G or T352M strains. Conclusions: All variants studied affect the oxidative phenotype, though at different extents, and both patients harbour one very detrimental variant in trans with a milder allele. Our data suggest a loss of mitochondrial complex III and IV subunits, which we are evaluating further.