The pathogenicity scoring system for mitochondrial tRNA mutations revisited

Abstract

Confirming the pathogenicity of mitochondrial tRNA point mutations is one of the classical challenges in the field of mitochondrial medicine. In addition to genetic and functional studies, the evaluation of a genetic change using a pathogenicity scoring system is extremely useful to discriminate between disease-causing mutations from neutral polymorphisms. The pathogenicity scoring system is very robust for confirming pathogenicity, especially of mutations that show impaired activity in functional studies. However, mutations giving normal results using the same functional approaches are disregarded, and this compromises the power of the system to rule out pathogenicity. We propose to include a new criterion in the pathogenicity scoring systems regarding mutations which fail to show any mitochondrial defect in functional studies. To evaluate this proposal we characterized two mutations, m.8296A>G and m.8347A>G, in the mitochondrial tRNA(L) (ys) gene (MT-TK) using trans-mitochondrial cybrid analysis. m.8347A>G mutation severely impairs oxidative phosphorylation, suggesting that it is highly pathogenic. By contrast, the behavior of cybrids homoplasmic for the m.8296A>G mutation is similar to cybrids containing wild-type mitochondrial DNA (mtDNA). The results indicate that including not only positive but also negative outcomes of functional studies in the scoring system is critical for facilitating the diagnosis of this complex group of diseases.

Keywords: MT-TK; Mitochondrial tRNA; pathogenicity; scoring system; trans-mitochondrial cybrid study.

Figure 1

Mitochondrial membrane potentials, lactate production, and H₂O₂ production in the cell lines used in this study. (A) Mitochondrial membrane potential (n = 4). (B) Lactate production (n = 4). (c) H₂O₂ production (n = 4) Student's t-test, *P < 0.05, ***P < 0.001 8347: cybrid cell line carrying the m8347A>G mutation. 8296: cybrid cell line carrying the m8296A>G mutation. Control: cybrid cell line carrying a haplogroup B-matched mtDNA obtained from a healthy donor.