A deafness-associated mitochondrial DNA mutation altered the tRNASer(UCN) metabolism and mitochondrial function

Abstract Mutations in mitochondrial DNA (mtDNA) have been associated with deafness and their pathophysiology remains poorly understood. In this study, we investigated the pathogenic mechanism of deafness-associated 7505A > G variant in the mitochondrial tRNA Ser(UCN) . The m.7505A > G variant affected the highly conserved adenine at position 11 (A11), disrupted the highly conserved A11-U24 base-pairing of DHU stem of tRNA Ser(UCN) and introduced a tertiary base pairing (G11-C56) with the C56 in the TΨC loop. We therefore hypothesized that the m.7505A > G variant altered both structure and function of tRNA Ser(UCN) . We demonstrated that the m.7505A > G variant perturbed the conformation and stability of tRNA Ser(UCN) , as indicated by an increased melting temperature and electrophoretic mobility of the mutated tRNA compared with the wild type molecule. Using the cybrids constructed by transferring mitochondria from the Chinese family into mitochondrial DNA (mtDNA)-less cells, we demonstrated the m.7505A > G variant led to significantly decreased steady-state levels of tRNA Ser(UCN) in the mutant cybrids, as compared with those of control cybrids. The aberrant tRNA Ser(UCN) metabolism resulted in the variable decreases in mtDNA-encoded polypeptides in the mutant cybrids. Furthermore, we demonstrated that the m.7505A > G variant decreased the activities of mitochondrial respiratory complexes I, III and IV, markedly diminished mitochondrial ATP levels and membrane potential, and increased the production of reactive oxygen species in the mutant cybrids. These results demonstrated that the m.7505A > G variant affected both structure and function of tRNA Ser(UCN) and consequently altered mitochondrial function. Our findings highlighted critical insights into the pathophysiology of maternally inherited deafness, which is manifested by the aberrant tRNA metabolism.