MohammedIsmail, W. | bioRxiv Science

bioRxiv2019

The symmetrical pattern of base-pair substitutions rates across the chromosome in Escherichia coli has multiple causes

Mutation accumulation experiments followed by whole-genome sequencing have revealed that for several bacterial species the rate of base-pair substitutions is not constant across the chromosome but varies in a wave-like pattern symmetrical about the origin of replication. The experiments reported here demonstrate that in Escherichia coli several interacting factors determine the wave. Perturbing replication timing, progression, or the structure of the terminus disrupts the pattern. Biases in error-correction by proofreading and mismatch repair are major factors. The activities of the nucleoid binding proteins, HU and Fis, are important, suggesting that mutation rates increase when highly structured DNA is replicated. These factors should apply to most bacterial, and possibly eukaryotic, genomes, and imply that different areas of the genome evolve at different rates.

microbiology↗

bioRxiv2018

Determinants of base-pair substitution patterns revealed by whole-genome sequencing of DNA mismatch repair defective Escherichia coli

Mismatch repair (MMR) is a major contributor to replication fidelity, but its impact varies with sequence context and the nature of the mismatch. Mutation accumulation experiments followed by whole-genome sequencing of MMR-defective E. coli strains yielded {approx}30,000 base-pair substitutions, revealing mutational patterns across the entire chromosome. The base-pair substitution spectrum was dominated by A:T > G:C transitions, which occurred predominantly at the center base of 5'NAC3'+5'GTN3' triplets. Surprisingly, growth on minimal medium or at low temperature attenuated these mutations. Mononucleotide runs were also hotspots for base-pair substitutions, and the rate at which these occurred increased with run length. Comparison with {approx}2000 base-pair substitutions accumulated in MMR-proficient strains revealed that both kinds of hotspots appeared in the wild-type spectrum and so are likely to be sites of frequent replication errors. In MMR-defective strains transitions were strand biased, occurring twice as often when A and C rather than T and G were on the lagging-strand template. Loss of nucleotide diphosphate kinase increases the cellular concentration of dCTP, which resulted in increased rates of mutations due to misinsertion of C opposite A and T. In an mmr ndk double mutant strain, these mutations were more frequent when the template A and T were on the leading strand, suggesting that lagging-strand synthesis was more error-prone or less well corrected by proofreading than was leading strand synthesis.

genetics↗