Sequence Analysis Of The Complete Mitochondrial Genome And TRMU Gene Of Two Pedigrees With Hearing Impairment Associated With The 12S RRNA A827G Mutation

Objective: To understand the potential role of the mitochondrial haplogroups and nuclear modifier gene TRMU in the phenotypic expression of the 12S rRNA A827G mutation in two families with non-syndromic hearing loss.Methods: Family members of two Chinese pedigrees with non-syndromic hearing loss and mitochondrial 12S rRNA A827G mutation were involved in this study. Genomic DNA was isolated from whole blood of participants using the Puregene DNA Isolation Kits. Firstly, the subject's DNA fragments spanning the entire mitochondrial 12S rRNA gene were amplified by PCR and sequenced to identify A827G mutation. The whole mitochondrial genomes and coding sequence of TRMU gene of the probands from two families who carry the A827G mutation were then PCR amplified using specific primers. Each fragment was purified and subsequently analyzed by direct sequencing in an Applied Biosystems 3730 automated DNA sequencer. The resultant sequence data were compared with the standard sequence to identify base variations.Results: PCR amplifications were successfully conducted in all the subjects. Mitochondrial 12S rRNA A827G mutation was detected in maternal members which was absent for the spouses of the two family. Sequence analysis of the complete mitochondrial genomes in the probands showed the distinct sets of mtDNA polymorphism: 17 other base variations in family A and 24 other nucleotide changes in family B respectively, in addition to the identical 12S rRNA A827G mutation. These variants were further evaluated by phylogenetic analysis with mtDNAs from other organisms, and none of the sites of these variants are evolutionarily conserved. In addition, sequence analysis of TRMU gene reveals no meaningful mutations.Conclusions: The mitochondrial 12S rRNA A to G mutation at position 827 is the main molecular mechanism responsible for the non-syndromic hearing loss in the two pedigrees. Both mitochondrial haplogroups and TRMU gene are unlikely to affect the phenotypic expression of the 12S rRNA A827G mutation in these two families.