A hypertension-associated mitochondrial DNA mutation alters the tertiary interaction and function of tRNALeu(UUR)

Several mitochondrial tRNA mutations have been associated with hypertension, but their pathophysiology remains poorly understood. In this report, we identified a novel homoplasmic 3253TC mutation in the mitochondrial tRNA(Leu(UUR)) gene in a Han Chinese family with maternally inherited hypertension. The m.3253TC mutation affected a highly conserved uridine at position 22 at the D-stem of tRNA(Leu(UUR)), introducing a G-C base pairing (G13-C22) at the D-stem and a tertiary base pairing (C22-G46) between the D-stem and the variable loop. We therefore hypothesized that the m.3253TC mutation altered both the structure and function of tRNA(Leu(UUR)). Using cytoplasmic hybrid (cybrid) cell lines derived from this Chinese family, we demonstrated that the m.3253TC mutation perturbed the conformation and stability of tRNA(Leu(UUR)), as suggested by faster electrophoretic mobility of mutated tRNA relative to the wild-type molecule. Northern blot analysis revealed an approximate to 45% decrease in the steady-state level of tRNA(Leu(UUR)) in the mutant cell lines carrying the m.3253TC mutation, as compared with control cell lines. Moreover, an approximate to 35% reduction in aminoacylation efficiency of tRNA(Leu(UUR)) was observed in the m.3253TC mutant cells. These alterations in tRNA(Leu(UUR)) metabolism impaired mitochondrial translation, especially for those polypeptides with a high proportion of Leu(UUR) codons, such as ND6. Furthermore, we demonstrated that the m.3253TC mutation decreased the activities of mitochondrial complexes I and V, markedly diminished mitochondrial ATP levels and membrane potential, and increased the production of reactive oxygen species in the cells. In conclusion, our findings may provide new insights into the pathophysiology of maternally inherited hypertension.