Abstract
BackgroundThe mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system. However, the exact molecular mechanisms underlying microsatellite variation have not been fully elucidated. Here, we investigated mutation events in the hyper-variable trinucleotide microsatellite locus MML0050 located in the Rv0050 gene of W-Beijing and non-W-Beijing Mycobacterium tuberculosis strains in order to gain insight into the genomic structure and activity of repeated regions.ResultsSize analysis indicated the presence of five alleles that differed in length by three base pairs. Moreover, nucleotide gains occurred more frequently than loses in this trinucleotide microsatellite. Mutation frequency was not completely related with the total length, though the relative frequency in the longest allele was remarkably higher than that in the shortest. Sequence analysis was able to detect seven alleles and revealed that point mutations enhanced the level of locus variation. Introduction of an interruptive motif correlated with the total allele length and genetic lineage, rather than the length of the longest stretch of perfect repeats. Finally, the level of locus variation was drastically different between the two genetic lineages.ConclusionThe Rv0050 locus encodes the bifunctional penicillin-binding protein ponA1 and is essential to mycobacterial survival. Our investigations of this particularly dynamic genomic region provide insights into the overall mode of microsatellite evolution. Specifically, replication slippage was implicated in the mutational process of this microsatellite and a sequence-based genetic analysis was necessary to determine that point mutation events acted to maintain microsatellite size integrity while providing genomic diversity.
Highlights
The mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system
The detected mutations involved gains or losses of either three, six, or nine bps. These trinucleotide units are equivalent to one, two, or three repeats, respectively, suggesting that replication slippage is associated with the mutation events
Size analysis indicated the presence of five MML0050 alleles that differed in length by three base pairs, implicating replication slippage in the mutational process of this microsatellite
Summary
The mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system. The exact molecular mechanisms underlying microsatellite variation have not been fully elucidated. We investigated mutation events in the hyper-variable trinucleotide microsatellite locus MML0050 located in the Rv0050 gene of WBeijing and non-W-Beijing Mycobacterium tuberculosis strains in order to gain insight into the genomic structure and activity of repeated regions. Strand slippage during DNA replication can cause insertion and deletion of repeat units in newly-synthesized nucleotide chains. These events are the most common cause of expansion or contraction of microsatellites [2,3]. Recent studies have reported differences in rates and patterns of mutation among distinct loci and species; allele size, motif size, genetic lineage, G/C content, functional potential of the transcribed product, and effectiveness of mismatch repair enzymes might all act as mediators of the mutation patterns of such loci [4,5,6,7]
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