Biochemical characterization of the deafness-associated mitochondrial tRNASer(UCN) A7445G mutation in osteosarcoma cell cybrids

The deafness-associated A7445G mutation in the precursor of mitochondrial tRNASer(UCN) has been identified in several pedigrees from different ethnic backgrounds. To determine the role of nuclear background in the biochemical manifestation associated with the A7445G mutation, we performed a biochemical characterization of this mutation using cybrids constructed by transferring mitochondria from lymphoblastoid cell lines derived from a New Zealand family into human osteosarcoma mtDNA-less (ρ0) cells. Compared with three control cybrids, three cybrids derived from an affected matrilineal relative carrying the homoplasmic A7445G mutation exhibited ∼38-57% decrease in the steady-state level of tRNASer(UCN), which is less reduced levels than in lymphoblastoid cells in the previous study. Furthermore, ∼22% reduction in the level of aminoacylation of tRNASer(UCN) was observed in the mutant cybrid cells. Interestingly, ∼60-63% decrease of steady-state level of ND6 gene, which belongs to the same precursor as that of tRNASer(UCN), in cybrid cell lines carrying the A7445G mutation, is more than that observed in lymphoblastoid cells. These observations strongly point out a mechanistic link between the processing defect of the tRNASer(UCN) precursor and decreased stability of ND6 mRNA precursor. These results also imply the influence of nuclear background on the biochemical phenotype associated with the A7445G mutation.