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Mismatch repair at stop codons is directed independent of GATC methylation on the Escherichia coli chromosome.

ABSTRACT: The mismatch repair system (MMR) corrects replication errors that escape proofreading. Previous studies on extrachromosomal DNA in Escherichia coli suggested that MMR uses hemimethylated GATC sites to identify the newly synthesized strand. In this work we asked how the distance of GATC sites and their methylation status affect the occurrence of single base substitutions on the E. coli chromosome. As a reporter system we used a lacZ gene containing an early TAA stop codon. We found that occurrence of point mutations at this stop codon is unaffected by GATC sites located more than 115 base pairs away. However, a GATC site located about 50 base pairs away resulted in a decreased mutation rate. This effect was independent of Dam methylation. The reversion rate of the stop codon increased only slightly in dam mutants compared to mutL and mutS mutants. We suggest that unlike on extrachromosomal DNA, GATC methylation is not the only strand discrimination signal for MMR on the E. coli chromosome.

SUBMITTER: Sneppen K

PROVIDER: S-EPMC5376664 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

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Mismatch repair at stop codons is directed independent of GATC methylation on the Escherichia coli chromosome.

Sneppen Kim K Semsey Szabolcs S

Scientific reports 20141205

The mismatch repair system (MMR) corrects replication errors that escape proofreading. Previous studies on extrachromosomal DNA in Escherichia coli suggested that MMR uses hemimethylated GATC sites to identify the newly synthesized strand. In this work we asked how the distance of GATC sites and their methylation status affect the occurrence of single base substitutions on the E. coli chromosome. As a reporter system we used a lacZ gene containing an early TAA stop codon. We found that occurrenc ...[more]

PMID: 25475788

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Project description:Methylated DNA adenines (6mA) are a critical epigenetic modification in bacteria that affect cell processes like replication, stress response, and pathogenesis. While much work has been done characterizing the influence of 6mA on specific loci, very few studies have examined the evolutionary dynamics of 6mA over long time scales. We used third-generation sequencing technology to analyze 6mA methylation across the Escherichia coli K-12 substr. MG1655 genome. 6mA levels were consistently high across GATC sites; however, we identified regions where 6mA is decreased, particularly in intergenic regions, especially around the -35 promoter element, and within cryptic prophages and IS elements. We further examined 6mA in WT and methyl-directed mismatch repair-deficient (MMR-) populations after 2,400 generations of experimental evolution. We find that, after evolution, MMR- populations acquire significantly more epimutations resulting in a genome-wide decrease in 6mA methylation. Here, clones from evolved MMR- populations display non-deterministic sets of epimutations, consistent with reduced selection on these modifications. Thus, we show that characterization of 6mA in bacterial populations is complementary to genetic sequencing and informative for molecular evolution. IMPORTANCE Methylation greatly influences the bacterial genome by guiding DNA repair and regulating pathogenic and stress-response phenotypes. But, the rate of epigenetic changes and their consequences on molecular phenotypes are underexplored. Through a detailed characterization of genome-wide adenine methylation in a commonly used laboratory strain of Escherichia coli, we reveal that mismatch repair deficient populations experience an increase in epimutations resulting in a genome-wide reduction of 6mA methylation in a manner consistent with genetic drift. Our findings highlight how methylation patterns evolve and the constraints on epigenetic evolution due to post-replicative DNA repair, contributing to a deeper understanding of bacterial genome evolution and how epimutations may introduce semi-permanent variation that can influence adaptation.

Dataset Information

Mismatch repair at stop codons is directed independent of GATC methylation on the Escherichia coli chromosome.

Publications

Mismatch repair at stop codons is directed independent of GATC methylation on the Escherichia coli chromosome.

Similar Datasets

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