Mutator action by
            <i>Escherichia coli</i>
            strains carrying
            <i>dnaE</i>
            mutations

Sevastopoulos, C. G.; Glaser, Donald A.

doi:10.1073/pnas.74.9.3947

Cited by 34 publications

(10 citation statements)

References 19 publications

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“…The in vivo experiments have suggested that indeed HE is highly accurate, as expected for the major replicating enzyme (Schaaper, 1993a). The importance of Pol III for controlling the mutation rate is also evidenced by the properties of several E. coli mutator mutants (which have enhanced mutation rates) carrying a defect in the dnaE gene encoding the Pol III a subunit (Sevastopoulos & Glaser, 1977;Fijalkowska et al, 1993;Oller et al, 1993;Vandewiele et al, 2002). Strong mutators have also been found residing in the dnaQ gene, encoding the e proofreading subunit (Fijalkowska & Schaaper, 1996;Taft-Benz & Schaaper, 1998), establishing the importance of the proofreading process for the chromosomal replication process.…”

Section: Dna Pol III Hementioning

confidence: 99%

DNA replication fidelity inEscherichia coli: a multi-DNA polymerase affair

2012

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High accuracy (fidelity) of DNA replication is important for cells to preserve the genetic identity and to prevent the accumulation of deleterious mutations. The error rate during DNA replication is as low as 10−9 to 10−11 errors per base pair. How this low level is achieved is an issue of major interest. This review is concerned with the mechanisms underlying the fidelity of the chromosomal replication in the model system Escherichia coli by DNA polymerase III holoenzyme, with further emphasis on participation of the other, accessory DNA polymerases, of which E. coli contains four (Pols I, II, IV, and V). Detailed genetic analysis of mutation rates revealed that (1) Pol II has an important role as a back‐up proofreader for Pol III, (2) Pols IV and V do not normally contribute significantly to replication fidelity, but can readily do so under conditions of elevated expression, (3) participation of Pols IV and V, in contrast to that of Pol II, is specific to the lagging strand, and (4) Pol I also makes a lagging‐strand‐specific fidelity contribution, limited, however, to the faithful filling of the Okazaki fragment gaps. The fidelity role of the Pol III τ subunit is also reviewed.

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Section: Dna Pol III Hementioning

confidence: 99%

DNA replication fidelity inEscherichia coli: a multi-DNA polymerase affair

2012

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“…In Escherichia coli, mutator mutations mapping in the dnaE gene presumably affect nucleotide selection by DNA polymerase III (28). Mutations mapping in the dnaQ gene (also called mutD) affect the proofreading 3'->5' exonuclease function of the DNA polymerase III holoenzyme (6,8), whereas mutH, mutL, mutS, and mutU mutations cause defects in methyl-directed postreplicative mismatch correction (for reviews, see references 21 and 24).…”

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confidence: 99%

Saturation of mismatch repair in the mutD5 mutator strain of Escherichia coli

1989

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The mutD (dnaQ) gene of Escherichia coli codes for the proofreading activity of DNA polymerase III. The very strong mutator phenotype of mutD5 strains seems to indicate that their postreplicational mismatch repair activity is also impaired. We show that the mismatch repair system of mutDS strains is functional but saturated, presumably by the excess of DNA replication errors, since it is recovered by inhibiting chromosomal DNA replication. This recovery depends on de novo protein synthesis.The mechanisms ensuring accuracy of DNA replication have been elucidated by study of mutator mutants affecting specific accuracy functions (for a review, see reference 4). In Escherichia coli, mutator mutations mapping in the dnaE gene presumably affect nucleotide selection by DNA polymerase III (28). Mutations mapping in the dnaQ gene (also called mutD) affect the proofreading 3'->5' exonuclease function of the DNA polymerase III holoenzyme (6, 8), whereas mutH, mutL, mutS, and mutU mutations cause defects in methyl-directed postreplicative mismatch correction (for reviews, see references 21 and 24). mutD5 and dnaQ49 mutants are the most potent mutator strains known. All classes of transition, transversion, and frameshift mutations are increased up to 105-fold when mutD5 strains are grown in rich medium (5, 10, 11). The mutD5 gene of E. coli encodes the epsilon subunit, which carries out the 3'-*5' proofreading exonuclease function of the DNA polymerase III holoenzyme (6,8). However, the high-mutation-rate effect of mutD5 strains cannot be explained only by the defect in exonuclease activity, as this mutation rate is comparable to the error rate of in vitro replication with a polymerase devoid of proofreading activity (9, 16). The strong mutator phenotype of mutD5 suggested that mismatch repair functions may be impaired as well. This prediction was supported by results from DNA heteroduplex transfection experiments (23,25,26). One hypothesis is that the high error rate in DNA replication saturates mismatch repair in mutD5 mutants (23). Therefore, we have tested the mismatch repair capacity of mutD5 strains under conditions in which chromosomal DNA replication was or was not allowed. Mismatch repair activity was determined by the extent of pure infective centers derived from infection with packaged hemimethylated heteroduplexes of lambda DNA. Under these conditions, bacteria proficient in mismatch repair produced essentially pure infective centers, whereas bacteria deficient in mismatch repair produced mostly mixed infective centers.To evaluate mismatch repair activity, pure hemimethy-* Corresponding author. t Present address: Institute of Molecular Biology, University of Oregon, Eugene, OR 97403. lated heteroduplexes of lambda DNA containing an A. C or a G T mismatch were artificially constructed and introduced into mutD5 and other appropriate strains. Hemimethylated heteroduplexes of lambda DNA with defined mismatches were prepared as previously described (19) by reannealing the separated DNA strands of a lambda mutant carry...

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“…All of the mutator mutations so far described in Escherichia coli increase the frequency of point mutations (base-pair substitutions and frame-shift mutations). Thermosensitive mutations in dnaE cause erratic functioning of DNA polymerase III (dnaE gene product), which includes incorrect nucleotides (1). Mutations in dnaQ (mutD) decrease the proofreading activity of the DNA polymerase III holoenzyme (2), and mutations in mutH, mutL, mutS, and mutU affect the methyl-directed postreplicative mismatch correction (for review, see ref.…”

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confidence: 99%

Mutations in the bglY gene increase the frequency of spontaneous deletions in Escherichia coli K-12.

Lejeune¹,

Danchin²

1990

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

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A strong mutator effect has been observed in Escherichia coli K-12 strains mutated in the bglY gene (27 min). The frequency of point mutations is not modified in bglY mutant strains. In contrast, a strong increase in spontaneous generation of large deletions has been observed in these strains, both for chromosomal markers (10-fold increase of galETKchU deletions and 100-fold increase ofptsl-cysK deletions) and for plasmid DNA (100-fold increase of large deletions in the region located upstream of the chloramphenicol-resistance gene in plasmid pGR71). bgIY mutations are recessive and can be complemented by a DNA fragment of 900 base pairs assumed to contain the entire bglY wild-type gene. This mutator effect is recA-independent.deletions extending from chiA to the closely located galETK operon. Aliquots of exponential cultures were plated onto McConkey medium containing sodium chlorate (0.2%) and galactose (0.2%). The plates were incubated 24 hr anaerobically and 6-10 hr aerobically. In these conditions, chiA mutants (chlorate resistant) appeared as red colonies, and chlA-galETK double mutants appeared as white colonies. To investigate whether these double mutants resulted from a large deletion or from two independent point mutations, the absence of the pgl gene (located between chiA and galETK) can be easily verified: when grown on LB plates containing maltose (0.2%), pgl mutant clones have the property to turn blue after treatment with iodine (9).The sequence of nucleotides within a gene can be mutationally altered by base-pair substitutions, frame-shift mutations, insertions of transposable elements, and inversions or deletions of large DNA fragments. Mutation rates are genetically controlled, and mutations that lead to appreciable alteration in the rate of mutations of other genes are known as mutator and antimutator mutations. All of the mutator mutations so far described in Escherichia coli increase the frequency of point mutations (base-pair substitutions and frame-shift mutations). Thermosensitive mutations in dnaE cause erratic functioning of DNA polymerase III (dnaE gene product), which includes incorrect nucleotides (1). Mutations in dnaQ (mutD) decrease the proofreading activity of the DNA polymerase III holoenzyme (2), and mutations in mutH, mutL, mutS, and mutU affect the methyl-directed postreplicative mismatch correction (for review, see ref.3).We have described (4) a method to produce large deletions at 27 min on the E. coli chromosome. In the present communication, we report that some of these mutant strains exhibit a strong mutator phenotype that can be related to a vastly increased frequency of spontaneous deletions. Genetic analysis of the mutants indicates that the absence of the bglY gene is responsible for this effect. MATERIALS AND METHODSBacterial Strains. The bacteria used in this work were all derivatives of E. coli K-12. Their source and characteristics are listed in Table 1. Genetic Methods. Phage P1 vir was used for transductions carried out by the procedure described by Mil...

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Mutator action by Escherichia coli strains carrying dnaE mutations

Cited by 34 publications

References 19 publications

DNA replication fidelity inEscherichia coli: a multi-DNA polymerase affair

DNA replication fidelity inEscherichia coli: a multi-DNA polymerase affair

Saturation of mismatch repair in the mutD5 mutator strain of Escherichia coli

Mutations in the bglY gene increase the frequency of spontaneous deletions in Escherichia coli K-12.

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