Novel point mutations in mitochondrial 16S rRNA gene of Chinese hamster cells

To know the nature and mechanisms of spontaneous mutations in mitochondrial DNA (mtDNA), we determined, by direct cycle sequencing, the nucleotide seq uence of the 3' terminal region of the mitochondrial 16S rRNA gene from chl oramphenicol-resistant (CAP-R) mutants isolated in Chinese hamster V79 cell s. Four different: base substitutions were identified in common for the six CAP-R mutants. All mutations were heteroplasmic. One A to G transition was mapped at a site within the putative peptidyl transferase domain, the targ et region for chloramphenicol, and one C to A transition and two T to G tra nsversions were located within the two different; segments which form the s tems of the hairpin loop structures attached to this key domain in the pred icted secondary structure of 16S rRNA. The mutations detected in this study do not map to the same sites where CAP-R mutations were found previously i n mammalian cells. Allele specific-PCR analyses revealed that all four muta tions occurred on a single mutant-DNA molecule, but not on several ones ind ependently. Together with the other previous reports, our data suggest that spontaneous mtDNA mutations may not be caused exclusively by oxidative DNA damage at least in 16S rRNA gene.