Mutator Phenotype Resulting from DNA Polymerase IV Overproduction in
            <i>Escherichia coli</i>
            : Preferential Mutagenesis on the Lagging Strand

Kuban, Wojciech; Banach-Orlowska, Magdalena; Bialoskorska, Malgorzata; Lipowska, Aleksandra; Schaaper, Roel M.; Jonczyk, Piotr; Fijałkowska, Iwona J.

doi:10.1128/jb.187.19.6862-6866.2005

Cited by 48 publications

(78 citation statements)

References 39 publications

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“…For example, the presence of a copy of the dinB gene on the FЈ episome in addition to the copy on the chromosome results in 4-fold more Pol IV and a 2-to 3-fold increase in mutation frequencies (22,36). The presence of the dinB gene on a multicopy plasmid results in 10-to 20-fold more Pol IV (36,73) and, depending on the mutational target, 5-to 200-fold increases in mutation frequencies (37,39,63,65,70,73,75). These observations strongly suggest that the mutagenic activity of Pol IV normally is tightly regulated in growing cells but that even a modest increase in abundance allows Pol IV to, at least partially, escape this regulation.…”

mentioning

confidence: 57%

The SMC-Like Protein Complex SbcCD Enhances DNA Polymerase IV-Dependent Spontaneous Mutation inEscherichia coli

Storvik

Foster

2011

J Bacteriol

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In Escherichia coli, RpoS, the general stress response sigma factor, regulates the activity of the specialized DNA polymerase DNA polymerase IV (Pol IV) both in stationary-phase and in exponential-phase cells. In Escherichia coli and its relatives, DNA damage results in the induction of about 40 genes as part of the SOS response. Three of these genes encode specialized DNA polymerases: DNA polymerase II (Pol II; encoded by the polB gene), DNA Pol IV (encoded by the dinB gene), and DNA Pol V (encoded by the umuDC genes) (11,35). Pol IV and Pol V are members of the Y family of DNA polymerases that are widely distributed in all domains of life. Y family polymerases can replicate damaged DNA, a process called translesion synthesis, but they replicate undamaged DNA with fidelities that are typically orders of magnitude lower than those achieved by replicative DNA polymerases (20,48

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mentioning

confidence: 57%

The SMC-Like Protein Complex SbcCD Enhances DNA Polymerase IV-Dependent Spontaneous Mutation inEscherichia coli

Storvik

Foster

2011

J Bacteriol

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“…Recently, Kuban et al reported that dinB was found to make more errors on the lagging strand during synthesis than on the leading strand [36]. In our [G/C] 10 vector, the mononucleotide G repeat is located on the leading strand of synthesis.…”

Section: Pol IV and Microsatellite Mutagenesismentioning

confidence: 89%

Escherichia coli DNA polymerase IV contributes to spontaneous mutagenesis at coding sequences but not microsatellite alleles

Jacob

Eckert

2007

Mutation Research/Fundamental and Molecular Mechanisms of Mutag

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Slipped strand mispairing during DNA synthesis is one proposed mechanism for microsatellite or short tandem repeat (STR) mutation. However, the DNA polymerase(s) responsible for STR mutagenesis have not been determined. In this study, we investigated the effect of the Escherichia coli dinB gene product (Pol IV) on mononucleotide and dinucleotide repeat stability, using an HSVtk gene episomal reporter system for microsatellite mutations. For the control vector (HSV-tk gene only) we observed a statistically significant 3.5-fold lower median mutation frequency in dinB -than dinB + cells (p<0.001, Wilcoxon Mann Whitney Test). For vectors containing an in-frame mononucleotide allele ([G/C] 10 ) or either of two dinucleotide alleles ([GT/CA] 10 and [TC/AG] 11 ) we observed no statistically significant difference in the overall HSV-tk mutation frequency observed between dinB + and dinB -strains. To determine if a mutational bias exists for mutations made by Pol IV, mutational spectra were generated for each STR vector and strain. No statistically significant differences between strains were observed for either the proportion of mutational events at the STR or STR specificity among the three vectors. However, the specificity of mutational events at the STR alleles in each strain varied in a statistically significant manner as a consequence of microsatellite sequence. Our results indicate that while Pol IV contributes to spontaneous mutations within the HSV-tk coding sequence, Pol IV does not play a significant role in spontaneous mutagenesis at [G/ C] 10 , [GT/CA] 10 , or [TC/AG] 11 microsatellite alleles. Our data demonstrate that in a wild type genetic background, the major factor influencing microsatellite mutagenesis is the allelic sequence composition.

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“…Alternatively, the differential specificities could reflect a more complicated mode of mutation production in ⌬holE versus ⌬holE::hot strains. For example, accessory DNA polymerases, such as Pol II or Pol IV, have been shown to be involved in the production of mutations, particularly when Pol III has difficulty extending certain terminal mispairs (1,17). Possibly, HE without and HE containing Hot behave differently with respect to this phenomenon of polymerase trafficking.…”

Section: Discussionmentioning

confidence: 99%

Mutator and Antimutator Effects of the Bacteriophage P1hotGene Product

Chikova

Schaaper

2006

J Bacteriol

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The Hot (homolog of theta) protein of bacteriophage P1 can substitute for the Escherichia coli DNA polymerase III subunit, as evidenced by its stabilizing effect on certain dnaQ mutants that carry an unstable polymerase III proofreading subunit (antimutator effect). Here, we show that Hot can also cause an increase in the mutability of various E. coli strains (mutator effect). The hot mutator effect differs from the one caused by the lack of . Experiments using chimeric /Hot proteins containing various domains of Hot and along with a series of point mutants show that both N-and C-terminal parts of each protein are important for stabilizing the subunit. In contrast, the N-terminal part of Hot appears uniquely responsible for its mutator activity.The Escherichia coli chromosome is replicated with high efficiency and fidelity by DNA polymerase (Pol) III holoenzyme (HE), a large dimeric multisubunit enzyme complex that simultaneously copies the leading and lagging strands at the replication fork (for reviews, see references 15 and 24). HE contains two core polymerase subassemblies, each composed of three separate subunits, ␣, ε, and , which are tightly bound in the linear order ␣-ε-. The ␣ subunit (dnaE gene product) (135 kDa) is the polymerase, while the ε subunit (dnaQ gene product) (28.5 kDa) is the 3Ј35Ј exonucleolytic activity that functions as a proofreader for replication errors.The precise function of the subunit (holE gene product) (8 kDa) within the Pol III core is less clear. binds tightly to the ε subunit and does not seem to interact with the ␣ subunit (2, 32). Strains lacking (⌬holE strains) are viable, indicating that the subunit is not essential (31). On the other hand, our laboratory has shown that such deletion strains possess a mutator phenotype in mismatch repair-defective strains, suggesting that may have a positive effect on the accuracy of DNA replication, likely through its effect on the ε proofreading activity (33). This fidelity role is consistent with the increase in the ε exonuclease activity observed in an in vitro exonuclease assay in the presence of (32). In addition, large effects of the deletion of were observed in strains containing an impaired or unstable ε subunit (33). For example, the mutability of the temperature-sensitive dnaQ49 mutator strain was increased nearly 1,000-fold upon the loss of (⌬holE strain). Based on these and other results, it was postulated (33) that the subunit fulfills a general stabilizing role for the intrinsically unstable ε subunit (7, 10, 12).appears to be well preserved throughout the enterobacteria, suggestive of a meaningful role for the protein. Homologs of have also been found, surprisingly, to be encoded by two conjugative plasmids as well as by bacteriophage P1 (3). In experiments with this P1 homolog produced by the P1 hot gene (homolog of theta) (22), we showed that the resulting Hot protein can substitute for in certain dnaQ mutators such as dnaQ49, as it was capable of reducing the high mutability of a dnaQ49 ⌬holE strain (3). In fact, for d...

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Mutator Phenotype Resulting from DNA Polymerase IV Overproduction in Escherichia coli : Preferential Mutagenesis on the Lagging Strand

Cited by 48 publications

References 39 publications

The SMC-Like Protein Complex SbcCD Enhances DNA Polymerase IV-Dependent Spontaneous Mutation inEscherichia coli

The SMC-Like Protein Complex SbcCD Enhances DNA Polymerase IV-Dependent Spontaneous Mutation inEscherichia coli

Escherichia coli DNA polymerase IV contributes to spontaneous mutagenesis at coding sequences but not microsatellite alleles

Mutator and Antimutator Effects of the Bacteriophage P1hotGene Product

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