Spectra of spontaneous mutations in Escherichia coli strains defective in mismatch correction: the nature of in vivo DNA replication errors.

Schaaper, Roel M.; Dunn, Ronnie L.

doi:10.1073/pnas.84.17.6220

Cited by 256 publications

(193 citation statements)

References 33 publications

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“…mutS::Tn10 Strain-As expected based on previous studies [31], complete disruption of MutS results in a strong mutator phenotype (Table 1). Sequence analysis of 216 independent lacI d mutants revealed that, in comparison to wild type E. coli, complete loss of MutS function resulted in a large increase of transition base substitutions (100-fold) and single-base insertiondeletions (45-fold), and a smaller increase (12-fold) of transversion base substitutions (Table 1).…”

Section: Contribution Of E Coli Muts Glu38 To Mmr In Vivosupporting

confidence: 84%

“…Sequence analysis of 216 independent lacI d mutants revealed that, in comparison to wild type E. coli, complete loss of MutS function resulted in a large increase of transition base substitutions (100-fold) and single-base insertiondeletions (45-fold), and a smaller increase (12-fold) of transversion base substitutions (Table 1). Single base mutations occurred throughout the N-terminal coding sequence of the lacI gene ( Figure 1A), and they were distributed in a non-random manner similar to that observed previously [31]. mutS(E38A/Q) Strains-E. coli strains in which MutS Glu38 has been replaced with either alanine or glutamine had lacI d mutant frequencies of 140 x 10 −8 and 68 x 10 −8 , respectively (Table 1).…”

Section: Contribution Of E Coli Muts Glu38 To Mmr In Vivosupporting

confidence: 76%

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Specialized mismatch repair function of Glu339 in the Phe-X-Glu motif of yeast Msh6

et al. 2007

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The major eukaryotic mismatch repair (MMR) pathway requires Msh2-Msh6, which, like E. coli MutS, binds to and participates in repair of the two most common replication errors, single base-base and single base insertion-deletion mismatches. For both types of mismatches, the side chain of E. coli Glu38 in a conserved Phe-X-Glu motif interacts with a mismatched base. The Oε of Glu38 forms a hydrogen bond with either the N7 of purines or the N3 of pyrimidines. We show here that changing E. coli Glu38 to alanine results in nearly complete loss of repair of both single base-base and single base deletion mismatches. In contrast, a yeast strain with alanine replacing homologous Glu339 in Msh6 has nearly normal repair for insertion-deletion and most base-base mismatches, but is defective in repairing base-base mismatches characteristic of oxidative stress, e.g., 8-oxo-G•A mismatches. The results suggest that bacterial MutS and yeast Msh2-Msh6 differ in how they recognize and/or process replication errors involving undamaged bases, and that Glu339 in Msh6 may have a specialized role in repairing mismatches containing oxidized bases.

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Section: Contribution Of E Coli Muts Glu38 To Mmr In Vivosupporting

confidence: 84%

Section: Contribution Of E Coli Muts Glu38 To Mmr In Vivosupporting

confidence: 76%

Specialized mismatch repair function of Glu339 in the Phe-X-Glu motif of yeast Msh6

et al. 2007

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“…In both cases the mismatched base pair had the canonical WatsonCrick geometry of the cognate base pair. One of the striking aspects of the MutL − data is that 79% of the A:T transitions occurred at sites with the sequence 5′ApC3′/3′TpG5′ (Table S2), a preference that has been observed in previous studies (22,63). We favor the hypothesis that these transitions are templated by the A for the following reason: because polymerase approaches the template base from the 3′ side, a C:G base pair would be established first and, because of its strong base-pairing and stacking interactions, could stabilize the A:C mispair.…”

Section: Discussionmentioning

confidence: 75%

Rate and molecular spectrum of spontaneous mutations in the bacteriumEscherichia colias determined by whole-genome sequencing

Lee¹,

Popodi

Tang³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

648

131

859

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Knowledge of the rate and nature of spontaneous mutation is fundamental to understanding evolutionary and molecular processes. In this report, we analyze spontaneous mutations accumulated over thousands of generations by wild-type Escherichia coli and a derivative defective in mismatch repair (MMR), the primary pathway for correcting replication errors. The major conclusions are (i) the mutation rate of a wild-type E. coli strain is ∼1 × 10 −3 per genome per generation; (ii) mutations in the wild-type strain have the expected mutational bias for G:C > A:T mutations, but the bias changes to A:T > G:C mutations in the absence of MMR; (iii) during replication, A:T > G:C transitions preferentially occur with A templating the lagging strand and T templating the leading strand, whereas G:C > A:T transitions preferentially occur with C templating the lagging strand and G templating the leading strand; (iv) there is a strong bias for transition mutations to occur at 5′ApC3′/3′TpG5′ sites (where bases 5′A and 3′T are mutated) and, to a lesser extent, at 5′GpC3′/3′CpG5′ sites (where bases 5′G and 3′C are mutated); (v) although the rate of small (≤4 nt) insertions and deletions is high at repeat sequences, these events occur at only 1/10th the genomic rate of base-pair substitutions. MMR activity is genetically regulated, and bacteria isolated from nature often lack MMR capacity, suggesting that modulation of MMR can be adaptive. Thus, comparing results from the wild-type and MMR-defective strains may lead to a deeper understanding of factors that determine mutation rates and spectra, how these factors may differ among organisms, and how they may be shaped by environmental conditions. evolution | mutation accumulation | neutral mutation | mutational hotspots | indels M utations are the source of variation upon which natural selection acts; thus, a complete understanding of evolutionary processes must include an accurate assessment of mutation rates and of the molecular spectrum of mutational events. In addition, we need to know whether, and how, intrinsic and extrinsic factors influence mutational processes. This understanding must be founded on baseline parameters established by analyzing mutations that accumulate in a neutral fashion, unbiased by selective pressures. Much of our knowledge of spontaneous mutation is based on mutations that occur in nonessential reporter genes during short-term laboratory culture of microorganisms (1). An alternative approach is to compare presumably neutral mutations that have accumulated over evolutionary time periods in diverged species (2). Both methods have substantial uncertainties. The experimental approach may use reporter loci that are not representative of the whole genome and necessarily incorporates assumptions about the expression and neutrality of the mutant phenotypes. The historical approach relies on estimated divergence times and the absence of selective pressure on synonymous sequence changes. High-throughput whole-genome sequencing allows some of these limitations to be...

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“…It is now widely accepted that transitions, in particular G:C to A:T, are predominant in cells defective in MMR. 16,56,57 On the other hand, A:T to C:G transversions predominate in proofreading-mutant mouse cells. 31 In E. coli dnaQ mutators, all types of base substitutions, in addition to frameshifts, are known to increase dramatically.…”

Section: Discussionmentioning

confidence: 99%

Concurrent genetic alterations in DNA polymerase proofreading and mismatch repair in human colorectal cancer

et al. 2010

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Genomic sequences encoding the 3¢ exonuclease (proofreading) domains of both replicative DNA polymerases, pol delta and pol epsilon, were explored simultaneously in human colorectal carcinomas including six established cell lines. Three unequivocal sequence alterations, including one previously reported, were found, and all these were considered as dysfunctional mutations in light of the local amino-acid sequences. In particular, the F367S mutation found in the POLE gene encoding the pol epsilon catalytic subunit, which includes the proofreading domain, is the first found in human diseases. Surprisingly, the tumours carrying these proofreading domain mutations were all defective in DNA mismatch repair (MMR). In addition to the two cell lines with acknowledged MMR gene mutations, the third tumour was also demonstrated to harbour a distinct mutation in MLH1, and indeed exhibited a microsatellite-unstable phenotype. These findings suggest that, in concert with MMR deficiency, defective polymerase proofreading may also contribute to the mutator phenotype observed in human colorectal cancer. Our observations may suggest previously unrecognised complexities in the molecular abnormalities underlying the mutator phenotype in human neoplasms. Keywords: polymerase delta; polymerase epsilon; proofreading domain; colorectal cancer INTRODUCTION Mutation rates on the genome are invariably regulated and typically suppressed to an extremely low level, such as 10 À10 per base replicated, in the organisms. 1 The high fidelity of DNA replication is achieved by several molecular mechanisms. The frequency of erroneous nucleotide incorporation is indeed extremely low compared with that expected from base-pairing energetics, and the vast reduction of the misincorporation frequency involves three steps of molecular events: (a) correct incorporation of complementary nucleotides by DNA polymerases, (b) removal of the newly added nucleotides, particularly incorrectly paired nucleotides, by the 3¢ exonuclease activities associated with the DNA polymerases (proofreading) and (c) postreplicative scanning of mispaired bases by DNA mismatch repair (MMR). 2,3 Various Escherichia coli (E. coli) mutator strains have been used to study these molecular mechanisms by which organisms maintain their mutation rates at very low levels. The dnaQ + (mutD + ) gene of E. coli encodes the epsilon subunit of the DNA polymerase III holoenzyme that is involved in 3¢ exonuclease proofreading activity, 4 and, therefore, the mutation frequencies in the dnaQ mutators are sometimes 1000-10 000 times higher than the wild-type levels. 4,5 In E. coli, MMR is chiefly directed by the proteins encoded by the three genes: mutS + , mutL + and mutH + . 6 The mutS or mutL mutators exhibit an approximately 100 times increase in the mutation frequency. 7 Thus, polymerase proofreading and MMR are the two major

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Spectra of spontaneous mutations in Escherichia coli strains defective in mismatch correction: the nature of in vivo DNA replication errors.

Cited by 256 publications

References 33 publications

Specialized mismatch repair function of Glu339 in the Phe-X-Glu motif of yeast Msh6

Specialized mismatch repair function of Glu339 in the Phe-X-Glu motif of yeast Msh6

Rate and molecular spectrum of spontaneous mutations in the bacteriumEscherichia colias determined by whole-genome sequencing

Concurrent genetic alterations in DNA polymerase proofreading and mismatch repair in human colorectal cancer

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