Mitochondrial DNA (mtDNA) maintenance is critical for oxidative phosphorylation (OXPHOS) since some subunits of the respiratory chain complexes are mitochondrially encoded. Pathological mutations in nuclear genes involved in the mtDNA metabolism may result in a quantitative decrease in mtDNA levels, referred to as mtDNA depletion, or in qualitative defects in mtDNA, especially in multiple deletions. Since, in the last decade, most of the novel mutations have been identified through whole-exome sequencing, it is crucial to confirm the pathogenicity by functional analysis in the appropriate model systems. Among these, the yeast Saccharomyces cerevisiae has proved to be a good model for studying mutations associated with mtDNA instability. This review focuses on the use of yeast for evaluating the pathogenicity of mutations in six genes, MPV17/SYM1, MRM2/MRM2, OPA1/MGM1, POLG/MIP1, RRM2B/RNR2, and SLC25A4/AAC2, all associated with mtDNA depletion or multiple deletions. We highlight the techniques used to construct a specific model and to measure the mtDNA instability as well as the main results obtained. We then report the contribution that yeast has given in understanding the pathogenic mechanisms of the mutant variants, in finding the genetic suppressors of the mitochondrial defects and in the discovery of molecules able to improve the mtDNA stability.

Maintaining protein lipoylation is vital for cell metabolism. The H-protein encoded by GCSH has a dual role in protein lipoylation required for bioenergetic enzymes including pyruvate dehydrogenase and 2-ketoglutarate dehydrogenase, and in the one-carbon metabolism through its involvement in glycine cleavage enzyme system, intersecting two vital roles for cell survival. Here we report six patients with biallelic pathogenic variants in GCSH and a broad clinical spectrum ranging from neonatal fatal glycine encephalopathy to an attenuated phenotype of developmental delay, behavioral problems, limited epilepsy, and variable movement problems. The mutational spectrum includes one insertion c.293-2_293–1insT, one deletion c.122_(228 + 1_229–1) del, one duplication of exons 4 and 5, one nonsense variant p.Gln76*and four missense p.His57Arg, p.Pro115Leu, and p.Thr148Pro and the previously described p.Met1?. Via functional studies in patient’s fibroblasts, molecular modelling, expression analysis in GCSH knock-down COS7 cells and yeast, and in vitro protein studies, we demonstrate for the first time that most variants identified in our cohort produced a hypomorphic effect on both mitochondrial activities, protein lipoylation and glycine metabolism, causing combined deficiency whereas some missense variants affect primarily one function only. The clinical features of the patients reflect the impact of the GCSH changes on any of the two functions analyzed. Our analysis illustrates the complex interplay of functional and clinical impact when pathogenic variants affect a multifunctional protein involved in two metabolic pathways and emphasizes the value of the functional assays to select the treatment and investigate new personalized options.

In eukaryotes, mitochondrial RNAs (mt-tRNAs and mt-rRNAs) are subject to specific nucleotide modifications, which are critical for distinct functions linked to the synthesis of mitochondrial proteins encoded by mitochondrial genes, and thus for oxidative phosphorylation. In recent years, mutations in genes encoding for mt-RNAs modifying enzymes have been identified as being causative of primary mitochondrial diseases, which have been called modopathies. These latter pathologies can be caused by mutations in genes involved in the modification either of tRNAs or of rRNAs, resulting in the absence of/decrease in a specific nucleotide modification and thus on the impairment of the efficiency or the accuracy of the mitochondrial protein synthesis. Most of these mutations are sporadic or private, thus it is fundamental that their pathogenicity is confirmed through the use of a model system. This review will focus on the activity of genes that, when mutated, are associated with modopathies, on the molecular mechanisms through which the enzymes introduce the nucleotide modifications, on the pathological phenotypes associated with mutations in these genes and on the contribution of the yeast Saccharomyces cerevisiae to confirming the pathogenicity of novel mutations and, in some cases, for defining the molecular defects.

No abstract

No abstract