A Hypertension-Associated tRNA^Ala Mutation Alters tRNA Metabolism and Mitochondrial Function

Abstract

In this report, we investigated the pathophysiology of a novel hypertension-associated mitochondrial tRNA^Ala 5655A → G (m.5655A → G) mutation. The destabilization of a highly conserved base pairing (A1-U72) at the aminoacyl acceptor stem by an m.5655A → G mutation altered the tRNA^Ala function. An in vitro processing analysis showed that the m.5655A → G mutation reduced the efficiency of tRNA^Ala precursor 5′ end cleavage catalyzed by RNase P. By using cybrids constructed by transferring mitochondria from lymphoblastoid cell lines derived from a Chinese family into mitochondrial DNA (mtDNA)-less (ρ^o) cells, we showed a 41% reduction in the steady-state level of tRNA^Ala in mutant cybrids. The mutation caused an improperly aminoacylated tRNA^Ala, as suggested by aberrantly aminoacylated tRNA^Ala and slower electrophoretic mobility of mutated tRNA. A failure in tRNA^Ala metabolism contributed to variable reductions in six mtDNA-encoded polypeptides in mutant cells, ranging from 21% to 37.5%, with an average of a 29.1% reduction, compared to levels of the controls. The impaired translation caused reduced activities of mitochondrial respiration chains. Furthermore, marked decreases in the levels of mitochondrial ATP and membrane potential were observed in mutant cells. These caused increases in the production of reactive oxygen species in the mutant cybrids. The data provide evidence for the association of the tRNA^Ala 5655A → G mutation with hypertension.

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.00199-16.

ACKNOWLEDGMENTS

We thank the members of Chinese families for their participation in this investigation.

This study was supported by grants from the National Basic Research Priorities Program of China (2014CB541700) and National Natural Science Foundation of China (81330024) to M.-X.G., grants (31371270 and 31401070) from the National Natural Science Foundation of China to P.J. and L.X., and a general financial grant from the China Postdoctoral Science Foundation to M.W. (2014M551726).