The 7472insC mitochondrial DNA mutation impairs the synthesis and extent of aminoacylation of tRNA^Ser(UCN) but not its structure or rate of turnover

The 7472insC mitochondrial DNA mutation in the tRNA^Ser(UCN) gene is associated with sensorineural deafness combined, in some patients, with a wider neurological syndrome. In cultured cybrid cells it causes a 70% decrease in tRNA^Ser(UCN) abundance and mild respiratory impairment, previously suggested to be due to decreased tRNA stability. When mitochondrial transcription was blocked by ethidium bromide treatment, the half-life of the mutant tRNA was not significantly different from that of wild-type tRNA^Ser(UCN). Over-expression of mitochondrial translational elongation factor EF-Tu also had no effect on the mutant phenotype. However, during recovery from prolonged ethidium bromide treatment, the synthesis of the mutant tRNA^Ser(UCN) was specifically impaired, without polarity effects on downstream tRNAs of the light strand transcription unit. We infer that the mutation acts posttranscriptionally to decrease tRNA^Ser(UCN) abundance by affecting its synthesis rather than its stability. The extent of aminoacylation of the mutant tRNA was also decreased by ∼25%. In contrast, the mutation had no detectable effect on tRNA^Ser(UCN) base modification or structure other than the insertion of an extra guanosine templated by the mutation, which was structurally protected from nuclease digestion like the surrounding nucleotides. These findings indicate a common molecular process underlying sensorineural deafness caused by mitochondrial tRNA^Ser(UCN) mutations.