Molecular Mechanism Of Nuclear Modifier MTO2/TRMU In Regulation Aminoglycoside Antibiotic Sensitity Of Yeast Carrying P^R₄₅₄Mutation

Mitochondrial DNA (mtDNA) mutation is one of the most important factors leading to sensorineural hearing loss. Mitochondrial12S rRNA A1555G mutation is associated with both aminoglycoside-induced and non-syndromic deafness. However, the phenotypic expression of population carrying A1555G mutation is often found to be pleiotropic, ranging from severe deafness to normal hearing. This indicates that modifier factors including nuclear genes, mitochondrial haplotype and ototoxic drugs could affect the phenotypic manifestation of A1555G mutation. Using yeast Saccharomyces cerevisiae as a model system, we investigated the crosstalk among mitochondrial PR454mutation (corresponding to human A1555G mutation), nuclear modifier MTO2/TRMU and aminoglycosides, as well as their combined effects on mitochondrial and cellular function.Our previous work found that G28T mutation (A1OS) in TRMU gene could suppress aminoglycoside antibiotic sensitivity in lymphocytes carrying mitochondrial A1555G mutation. Yeast model was constructed by deleting TRMU homolog MT02gene from a strain carrying mitochondrial15S rRNA PR454mutation. We analyzed the effects of aminoglycosides on yeast carrying PR454mutation, and the role of nuclear modifier MTO2was also investigated. Phenotypic expression indicated that deletion of nuclear gene MT02suppressed aminoglycoside antibiotic sensitivity in-yeast carrying PR454mutation, and the underlying molecular mechanism was further analyzed. Biochemical function of cell and mitochondria including respiratory rate, mitochondrial membrane potential, antibiotic intake, expression level of drug resistance gene and energy metabolism were tested. Results showed that mitochondrial function in PR454mutant was impaired by aminoglycosides. Noticeably, deletion of MTO2from PR454mutant strain enhanced the expression levels of HXK2, PFK1and PYK1which regulating the glycolytic pathway. We proposed this independence of mitochondrial function and accelerated glycolysis made this strain less sensitive to aminoglycosides.Yeast strains expressing human TRMU gene were obtained by transforming TRMU cDNA into mto2(PR). Human TRMU gene was specifically expressed in yeast mitochondria, and trmu could be functionally complementary with yeast mto2p. Interestingly, yeast transformed with TRMU A1OS mutant cDNA was less sensitive to neomycin when compared with the TRMU wild-type transformant. However, functional analysis showed the mechanism that TRMU gene regulating neomycin sensitivity was different from yeast MTO2, and mitochondrial dysfunction had not been observed in yeast transformed with TRMU AlOS mutant cDNA. We also analyzed this difference and suggested a potential model to explain the phenomenon.Current study discovered a novel function of MT02/TRMU gene in regulating aminoglycoside sensitivity caused by mitochondrial (A1555G) mutation, and the crosstalk among nuclear modifier gene, mtDNA mutation and environmental factors was elucidated. Meanwhile, these results provided new insights for the study of human nuclear modifier TRMU.