Genomes of all living organisms are constantly threatened by endogenous and exogenous agents that challenge the chemical integrity of DNA. Most bacteria have evolved a coordinated response to DNA damage. InEscherichia coli,this inducible system is termed the SOS response. The SOS global regulatory network consists of multiple factors promoting the integrity of DNA as well as error‐prone factors allowing for survival and continuous replication upon extensive DNA damage at the cost of elevated mutagenesis. Due to its mutagenic potential, the SOS response is subject to elaborate regulatory control involving not only transcriptional derepression, but also post‐translational activation, and inhibition. This review summarizes current knowledge about the molecular mechanism of the SOS response induction and progression and its consequences for genome stability. Environ. Mol. Mutagen. 60:368–384, 2019. © 2018 The Authors.Environmental and Molecular Mutagenesispublished by Wiley Periodicals, Inc. on behalf of Environmental Mutagen Society.

We have investigated the question whether during chromosomal DNA replication in Escherichia coli the two DNA strands may be replicated with differential accuracy. This possibility of differential replication fidelity arises from the distinct modes of replication in the two strands, one strand (the leading strand) being synthesized continuously, the other (the lagging strand) discontinuously in the form of short Okazaki fragments. We have constructed a series of lacZ strains in which the lac operon is inserted into the bacterial chromosome in the two possible orientations with regard to the chromosomal replication origin oriC . Measurement of lac reversion frequencies for the two orientations, under conditions in which mutations reflect replication errors, revealed distinct differences in mutability between the two orientations. As gene inversion causes a switching of leading and lagging strands, these findings indicate that leading and lagging strand replication have differential fidelity. Analysis of the possible mispairs underlying each specific base pair substitution suggests that the lagging strand replication on the E. coli chromosome may be more accurate than leading strand replication.

This review is concerned with the mechanisms underlying the fidelity of the chromosomal replication in the model system Escherichia coli by DNA polymerase III h

The Escherichia coli dnaQ gene encodes the proofreading 3' exonuclease (epsilon subunit) of DNA polymerase III holoenzyme and is a critical determinant of chromosomal replication fidelity. We constructed by site-specific mutagenesis a mutant, dnaQ926, by changing two conserved amino acid residues (Asp-12-->Ala and Glu-14-->Ala) in the Exo I motif, which, by analogy to other proofreading exonucleases, is essential for the catalytic activity. When residing on a plasmid, dnaQ926 confers a strong, dominant mutator phenotype, suggesting that the protein, although deficient in exonuclease activity, still binds to the polymerase subunit (alpha subunit or dnaE gene product). When dnaQ926 was transferred to the chromosome, replacing the wild-type gene, the cells became inviable. However, viable dnaQ926 strains could be obtained if they contained one of the dnaE alleles previously characterized in our laboratory as antimutator alleles or if it carried a multicopy plasmid containing the E. coli mutL+ gene. These results suggest that loss of proofreading exonuclease activity in dnaQ926 is lethal due to excessive error rates (error catastrophe). Error catastrophe results from both the loss of proofreading and the subsequent saturation of DNA mismatch repair. The probability of lethality by excessive mutation is supported by calculations estimating the number of inactivating mutations in essential genes per chromosome replication.

To better understand the mechanisms of SOS mutagenesis in the bacterium Escherichia coli, we have undertaken a genetic analysis of the SOS mutator activity. The SOS mutator activity results from constitutive expression of the SOS system in strains carrying a constitutively activated RecA protein (RecA730). We show that the SOS mutator activity is not enhanced in strains containing deficiencies in the uvrABC nucleotide excision-repair system or the xth and nfo base excision-repair systems. Further, recA730-induced errors are shown to be corrected by the MutHLS-dependent mismatch-repair system as efficiently as the corresponding errors in the rec+ background. These results suggest that the SOS mutator activity does not reflect mutagenesis at so-called cryptic lesions but instead represents an amplification of normally occurring DNA polymerase errors. Analysis of the base-pair-substitution mutations induced by recA730 in a mismatch repair-deficient background shows that both transition and transversion errors are amplified, although the effect is much larger for transversions than for transitions. Analysis of the mutator effect in various dnaE strains, including dnaE antimutators, as well as in proofreading-deficient dnaQ (mutD) strains suggests that in recA730 strains, two types of replication errors occur in parallel: (i) normal replication errors that are subject to both exonucleolytic proofreading and dnaE antimutator effects and (ii) recA730-specific errors that are not susceptible to either proofreading or dnaE antimutator effects. The combined data are consistent with a model suggesting that in recA730 cells error-prone replication complexes are assembled at sites where DNA polymerization is temporarily stalled, most likely when a normal polymerase insertion error has created a poorly extendable terminal mismatch. The modified complex forces extension of the mismatch largely at the exclusion of proofreading and polymerase dissociation pathways. SOS mutagenesis targeted at replication-blocking DNA lesions likely proceeds in the same manner.

We have analyzed the spectra of UV-induced mutations in the lacI gene of a wild-type and an mfd strain of Escherichia coli. mfd strains have been recently proposed to be deficient in a factor coupling DNA repair and transcription. Analysis of UV-induced mutations occurring at adjacent pyrimidines showed that mutations in the wild-type strain arose largely from the nontranscribed strand but arose predominantly from the transcribed strand in the mfd strain. The overall strand switch was 14-fold. One mutation, G.C-->A.T in the lacI initiation codon, showed a > 300-fold shift. No effect was observed for mutations at non-pyrimidine-pyrimidine sequences. These results provide in vivo evidence for a key role of the mfd gene in controlling the strandedness of mutagenesis and support the proposed role of the mfd gene product in directing DNA excision repair to the transcribed strand of a damaged gene.