A hypertension-associated mitochondrial DNA mutation introduces an m1G37 modification into tRNAMet, altering its structure and function

Mi Zhou, Ling Xue, Yaru Chen, Haiying Li, Qiufen He, Bibin Wang, Feilong Meng, Meng Wang, Min-Xin Guan
2017 Journal of Biological Chemistry  
Defective nucleotide modifications of mitochondrial tRNAs have been associated with several human diseases, but their pathophysiology remains poorly understood. In this report, we investigated the pathogenic molecular mechanism underlying a hypertension-associated 4435A3 G mutation in mitochondrial tRNA Met . The m.4435A3 G mutation affected a highly conserved adenosine at position 37, 3 adjacent to the tRNA's anticodon, which is important for the fidelity of codon recognition and
more » ... We hypothesized that the m.4435A3 G mutation introduced an m 1 G37 modification of tRNA Met , altering its structure and function. Primer extension and methylation activity assays indeed confirmed that the m.4435A3 G mutation created a tRNA methyltransferase 5 (TRMT5)-catalyzed m 1 G37 modification of tRNA Met . We found that this mutation altered the tRNA Met structure, indicated by an increased melting temperature and electrophoretic mobility of the mutated tRNA compared with the wildtype molecule. We demonstrated that cybrid cell lines carrying the m.4435A3 G mutation exhibited significantly decreased efficiency in aminoacylation and steadystate levels of tRNA Met , as compared with those of control cybrids. The aberrant tRNA Met metabolism resulted in variable decreases in mitochondrial DNA (mtDNA)-encoded polypeptides in the mutant cybrids. Furthermore, we found that the m.4435A3 G mutation caused respiratory deficiency, markedly diminished mitochondrial ATP levels and membrane potential, and increased the production of reactive oxygen species in mutant cybrids. These results demonstrated that an aberrant m 1 G37 modification of mitochondrial tRNA Met affected the structure and function of its tRNA and consequently altered mito-chondrial function. Our findings provide critical insights into the pathophysiology of maternally inherited hypertension, which is manifested by the deficient tRNA nucleotide modification. Defects in nucleotide modifications of mitochondrial tRNAs have been associated with several clinical abnormalities including cancer, diabetes, neurological disorders, deafness, and hypertension (1-4). These nucleotide modifications of human 22 mitochondrial tRNAs encoded by its own genome were catalyzed by tRNA-modifying enzymes encoded by nuclear genome (5-8). To date, 15 types of modifications have been identified in 118 positions in different mammalian mitochondrial tRNAs (9, 10). These nucleotide modifications play a vital role in the structure and function of tRNAs (11) (12) (13) (14) . Core modifications including pseudouridinylation at position 55 at the T⌿C loop primarily contributed to structural stability of tRNAs and in some cases, may affect the aminoacylation (2, 3). These were exemplified by our recent discovery that the loss of pseudouridinylation at position 55 at the T⌿C loop of tRNA Glu caused the maternally inherited deafness and diabetes (4). Indeed, the anticodon loop modifications including nucleotides at positions 34 and 37 regulate the stabilization of anticodon structure, fidelity, and efficiency of translation (2, 3, 11, 15-17). The defective 5-taurinomethyluridine (m 5 U) at U34 of tRNA Leu(UUR) was associated with mitochondrial encephalopathy lactic acidosis and stroke-like episodes, whereas the lack of 5-taurinomethyl-2-thiouridine (m 5 s 2 U) at U34 of tRNA Lys was responsible for myoclonus epilepsy associated with ragged red fibers (MERRF) (18 -20). Furthermore, mutations in tRNA modifying enzymes TRMU, MTO1, GTPBP3, and NSUN3 involved in nucleotide modifications at position 34 of mitochondrial tRNAs have been associated several clinical phenotypes including deafness (21-25).
doi:10.1074/jbc.ra117.000317 pmid:29222331 fatcat:ebdwfh6v3bgjngvevp3eiclspe