Point mutations in mitochondrial tRNA genes are responsible for individual subgroups of mitochondrial encephalomyopathies. We have recently reported that point mutations in the tRNA(Leu) (UUR) and tRNA(Lys) genes cause a defect in the normal modification at the first nucleotide of the anticodon. As part of a systematic analysis of pathogenic mutant mitochondrial tRNAs, we purified tRNA(Ile) with a point mutation at nucleotide 4269 to determine its nucleotide sequence, including modified nucleotides. We found that, instead of causing a defect in the post-transcriptional modification, a pathogenic point mutation in the mitochondrial tRNA(Ile) reduced the stability of the mutant tRNA molecule, resulting in a low steady-state level of aminoacyl-tRNA. The reduced stability was confirmed by examining the life-span of the mutant tRNA(Ile) both in vitro and in vivo, as well as by monitoring its melting profile. Our finding indicates that the mutant tRNA(Ile) itself is intrinsically unstable.
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