Abstract
Nuclear modifier gene(s) was proposed to modulate the phenotypic expression of mitochondrial DNA mutation(s). Our previous investigations revealed that a nuclear modifier allele (A10S) in TRMU (methylaminomethyl-2-thiouridylate-methyltransferase) related to tRNA modification interacts with 12S rRNA 1555A→G mutation to cause deafness. The A10S mutation resided at a highly conserved residue of the N-terminal sequence. It was hypothesized that the A10S mutation altered the structure and function of TRMU, thereby causing mitochondrial dysfunction. Using molecular dynamics simulations, we showed that the A10S mutation introduced the Ser10 dynamic electrostatic interaction with the Lys106 residue of helix 4 within the catalytic domain of TRMU. The Western blotting analysis displayed the reduced levels of TRMU in mutant cells carrying the A10S mutation. The thermal shift assay revealed the Tm value of mutant TRMU protein, lower than that of the wild-type counterpart. The A10S mutation caused marked decreases in 2-thiouridine modification of U34 of tRNALys, tRNAGlu and tRNAGln However, the A10S mutation mildly increased the aminoacylated efficiency of tRNAs. The altered 2-thiouridine modification worsened the impairment of mitochondrial translation associated with the m.1555A→G mutation. The defective translation resulted in the reduced activities of mitochondrial respiration chains. The respiratory deficiency caused the reduction of mitochondrial ATP production and elevated the production of reactive oxidative species. As a result, mutated TRMU worsened mitochondrial dysfunctions associated with m.1555A→G mutation, exceeding the threshold for expressing a deafness phenotype. Our findings provided new insights into the pathophysiology of maternally inherited deafness that was manifested by interaction between mtDNA mutation and nuclear modifier gene.
Highlights
Nuclear modifier gene(s) was proposed to modulate the phenotypic expression of mitochondrial DNA mutation(s)
We showed that MTO1, MSS1 (GTPBP3), or MTO2 (TRMU) genes involved in the biosynthesis of the hypermodified nucleoside 5-methyl-aminomethyl-2thio-uridine of several mitochondrial tRNAs were the potential modifier genes for the phenotypic expression of deafness-associated 12S rRNA 1555A3 G mutation [25,26,27,28,29,30]
To further investigate the effect of the TRMU A10S mutation on mitochondrial function, we examined for the levels of tRNA modification, aminoacylation of tRNAs, translation, the rates of respiration, and the production of ATP and reactive oxygen species (ROS), through use of lymphoblastoid mutant cell lines derived from Arab-Israeli control subjects and from members of an Arab-Israeli family (two subjects carrying only m.1555A3 G mutation (F12H and F6D), two individuals (F20C and F8A) harboring both m.1555A3 G and heterozygous TRMU A10S mutations, and two individuals (F20A and F20D) carrying both m.1555A3 G and homozygous TRMU A10S mutations) [25, 40]
Summary
Nuclear modifier gene(s) was proposed to modulate the phenotypic expression of mitochondrial DNA mutation(s). We showed that MTO1, MSS1 (GTPBP3), or MTO2 (TRMU) genes involved in the biosynthesis of the hypermodified nucleoside 5-methyl-aminomethyl-2thio-uridine of several mitochondrial tRNAs were the potential modifier genes for the phenotypic expression of deafness-associated 12S rRNA 1555A3 G mutation [25,26,27,28,29,30] These modified uridines at the wobble positions of tRNALys, tRNAGlu, and tRNAGln have a pivotal role in the structure and function of tRNAs, including structural stabilization, aminoacylation, and codon recognition at the decoding site of small rRNA [31,32,33]. To further investigate the effect of the TRMU A10S mutation on mitochondrial function, we examined for the levels of tRNA modification, aminoacylation of tRNAs, translation, the rates of respiration, and the production of ATP and reactive oxygen species (ROS), through use of lymphoblastoid mutant cell lines derived from Arab-Israeli control subjects and from members of an Arab-Israeli family (two subjects carrying only m.1555A3 G mutation (F12H and F6D), two individuals (F20C and F8A) harboring both m.1555A3 G and heterozygous TRMU A10S mutations, and two individuals (F20A and F20D) carrying both m.1555A3 G and homozygous TRMU A10S mutations) [25, 40]
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