The Identification of Nucleotide Variations in Mitochondrial tRNA of Serine, Asparagine, Aspartic acid, Cysteine and Tyrosine Genes in Non-Dystrophic Myotonia Patients

Introduction: The nondystrophic myotonias are a heterogeneous set of rare diseases that demonstrate clinicalmyotonia, electrical myotonia, or both. Myotonia is an intrinsic disorder of muscle caused by defects in either ionchannel or muscle membrane function.Based on researches the association of mutations in mitochondrial tRNA genes has been discovered for many ofdiseases such as seizure, cardiac dysfunction, and epilepsy. tRNAs have important functions within numerousprotein-RNA complexes and therefore minimal changes in their precise three dimensional folding could defectefficiency of their assembly and complex bindings. The aim of this research is to identification of nucleotidevariations in mitochondrial tRNAAsn, Tyr, Asp, Ser, Cys genes in Iranian patients with non-dystrophic mytonia.Methods: After PCR of mitochondrial tRNA of Serine, Asparagine, Aspartic acid, Cysteine and Tyrosine genesfrom 28 Iranian patient’s blood samples, all PCR products were run on SSCP gel and samples which showedconsiderable shifts was sent for DNA sequencing.Results: We found a novel homoplasmic mutation in the mtDNA tRNAcys gene. The 5794A> G mutationchanges the nucleotide flanking the anticodon loop. The mutation is located in a conserved region, and none ofthe databases has ever been reported until now.To visualize the second structure of the mutated tRNA, we use the website mfold (unafold.rna.albany.edu/mfold)for predicting the secondary structure of RNA and DNA, mainly by using thermodynamic methods andRNAfold (rna.tbi.univie.ac.at/cgi bin/RNAWebSuite/RNAfold.cg) that predicts secondary structures of singlestranded RNA or DNA sequences for partition function calculations and minimum free energy (MFE). Ourresults showed that this mutation alters the minimum free energy (MFE) from -17.60 kcal/mol to -18.10 kcal/mol.Conclusion: The tRNACys change was the only mutation affecting a conserved position in the entire mtDNAsequence of the proband. The mutation can alter the secondary, and possibly tertiary, structure of the anticodontRNA loop. Based on our results we suggest that T5794C can be potentially a pathogenic mutation.

Keywords:

Non-dystrophic Myotonia , Mitochondrial tRNA , PCR-SSCP