Overexpression of vsr in Escherichia coli is mutagenic

Doiron, Kathy M.J; Viau, Stéphane; Koutroumanis, Maria; Cupples, Claire G.

doi:10.1128/jb.178.14.4294-4296.1996

Cited by 31 publications

(24 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, this hypothesis is counterintuitive given the decided preference of Vsr for T/G mismatches occurring at sites of Dcm methylation (6). It also contradicts previous evidence from our lab which shows that VSP repair is dominant over MMR at C(T/G)AGG sites (4). Nevertheless, we explored the role of MMR in the reversal of deamination damage further by measuring CCAGG-to-CTAGG mutations in other MMR Ϫ backgrounds.…”

Section: Discussioncontrasting

confidence: 46%

Very-Short-Patch Repair in Escherichia coli Requires the dam Adenine Methylase

Bell

Cupples

2001

J Bacteriol

View full text Add to dashboard Cite

Strains of Escherichia coli which lack the dam-encoded adenine methylase are mutators due to a reduction in the efficiency of postreplication mismatch repair. In this study, we show that Dam ؊ strains are also defective in very-short-patch repair, the system which corrects T/G mismatches arising from the deamination of 5-methylcytosine. This defect is associated with decreased levels of Vsr, the endonuclease which initiates short-patch repair. We also show that production of the dcm-encoded cytosine methylase is unaffected in Dam ؊ strains. Since the dcm and vsr genes are cotranscribed, the regulation of Vsr by Dam is probably posttranscriptional.The methylation of GATC sites in the Escherichia coli K-12 genome by the dam-encoded adenine methylase is crucial for efficient methyl-directed mismatch repair (MMR) (reviewed in reference 19). MMR functions most effectively during the time period between the synthesis of a new strand of DNA and its methylation by Dam. The transient undermethylation of GATC's targets MMR to the new strand of the DNA, preserving the sequence of the template strand. The lesion itself, a mispaired or unpaired base, is recognized by MutS. In conjunction with MutL, this protein activates MutH, an endonuclease which cleaves the unmethylated strand of hemimethylated GATC sites. The repair process is completed by removal and resynthesis of the DNA between the nick and the errant base. dam strains, like mut strains, are mutators (16), characterized by an increased incidence of transition and frameshift mutations. Increased production of Dam is also mutagenic, presumably due to premature methylation of the newly synthesized DNA strand (9).In contrast to MMR, the very-short-patch (VSP) repair system of E. coli is thought to be independent of adenine methylation. VSP repair corrects T/G mismatches caused by deamination of 5-methylcytosine to thymine (reviewed in reference 12). Repair is initiated by Vsr, an endonuclease which cleaves 5Ј of the mismatched T (8). Strains lacking VSP repair have a high frequency of C-to-T mutations, primarily at CCWGG sites (W ϭ A or T). The site specificity is due to the fact that Dcm, the sole cytosine methylase of E. coli K-12, methylates the second C of this sequence. VSP repair is reduced in mutS and mutL strains but is unaffected in mutH cells (10,11,24). The independence of MutH, combined with the fact that fully methylated and unmethylated C(T/G)AGG heteroduplexes are repaired as efficiently as hemimethylated DNA, suggested that Dam methylation is not important for VSP repair. However, the extent of VSP repair in a dam background has never been tested explicitly.In this study, we used a Lac reversion assay to compare the frequency of CCAGG-to-CTAGG mutations in dam strains with that in mutS, mutL, and mutH strains. Mutation is increased far more in the dam strain than in any of the mut strains. Furthermore, the majority of mutations in the dam strain are dependent on the presence of the Dcm methylase and thus result from lack of VSP repair not from a defect in...

show abstract

Section: Discussioncontrasting

confidence: 46%

Very-Short-Patch Repair in Escherichia coli Requires the dam Adenine Methylase

Bell

Cupples

2001

J Bacteriol

View full text Add to dashboard Cite

show abstract

“…Twelve of these were not previously known to cause a mutator phenotype when overexpressed. One gene previously shown to increase mutagenesis when overexpressed (8) was not detected in our study. A striking finding from this work is that overexpression of the emrR gene, involved in regulating the EmrAB multidrug resistance pump (25; emr signifying E. coli multidrug resistance [24]), leads to greatly increased mutagenesis (45).…”

contrasting

confidence: 41%

Mutator Effects in Escherichia coli Caused by the Expression of Specific Foreign Genes

2005

View full text Add to dashboard Cite

Certain genes from Lactococcus lactis and Pseudomonas aeruginosa, including the nfxB gene, generate a mutator phenotype in Escherichia coli. The results of this study, together with those of a previous study, support conservation of regulatory sequences in E. coli and P. aeruginosa and suggest that some efflux pumps prevent mutagenicity by exporting mutagenic products of metabolism.Understanding mutational pathways and the repair systems that operate on them has proved to be important in elucidating the nature of a number of human diseases, including cancer (for instance, for reviews, see references 17 and 22). The discovery of genes involved in mutagenesis and repair has frequently depended on mutational analysis, and often this involves looking for genes that when inactivated result in a mutator (increased mutagenesis) phenotype (for reviews, see references 29 and 30). For example, in Escherichia coli the inactivation of either the dam, mutS, mutL, mutH, or uvrD gene leads to loss of the mismatch repair system (13,26,34) and loss of the mutY or mutM gene leads to reduced capacity to repair oxidative damage to DNA (5,27,35). The human counterparts to mutS, mutL, mutY, and mutM (ogg-1) have been characterized (2, 38, 44; for reviews, see references 9, 21, and 32), and loss of the mismatch repair system or the MutY repair capacity leads to increased cancer susceptibilities in humans (1,20). Defining additional genes involved in mutagenesis is therefore of great interest. New approaches may lead to novel mutational pathways. Recently, using a random shotgun cloning approach, we have identified 15 genes in E. coli that when cloned onto a multicopy plasmid result in a mutator phenotype (45). Twelve of these were not previously known to cause a mutator phenotype when overexpressed. One gene previously shown to increase mutagenesis when overexpressed (8) was not detected in our study. A striking finding from this work is that overexpression of the emrR gene, involved in regulating the EmrAB multidrug resistance pump (25; emr signifying E. coli multidrug resistance [24]), leads to greatly increased mutagenesis (45). We have extended this approach by looking for genes from foreign DNA that result in increased mutagenesis when cloned into E. coli on a multicopy plasmid. This analysis might not only identify potential mutational pathways in E. coli by finding genes whose counterparts did not show up in the original study but can also allow the dissection of mutagenic and repair pathways in microorganisms that do not have developed genetic systems that provide mutator screens. In the work reported here, we identify a set of genes from Lactococcus lactis that result in a mutagenic response in E. coli. We also find that only one gene from the opportunistic pathogen Pseudomonas aeruginosa generates a detectable mutagenic effect when cloned onto a specific multicopy plasmid in E. coli. This gene, the nfxB gene (nfx signifying resistance to norfloxacin [19]), is a regulator of one of the P. aeruginosa multidrug resistance pumps...

show abstract

“…These results suggest that MutL and MutS serve distinct functions in promoting VSP repair and that the absence of MutL in cells cannot be compensated for by increasing amounts of MutS. Furthermore, just as overproduction of Vsr disrupts MMR (3,16), overproduction of MutS disrupts VSP repair, suggesting that both proteins must be present in cells at optimal levels for both mismatch repair processes to minimize mutations. Dominant-negative mutS mutations differ with respect to their effect on VSP repair.…”

mentioning

confidence: 88%

Interaction of MutS and Vsr: Some Dominant-Negative mutS Mutations That Disable Methyladenine-Directed Mismatch Repair Are Active in Very-Short-Patch Repair

2001

View full text Add to dashboard Cite

In Escherichia coli and related bacteria, the very-short-patch (VSP) repair pathway uses an endonuclease, Vsr, to correct T ⅐ G mismatches that result from the deamination of 5-methylcytosines in DNA to C ⅐ G. The products of mutS and mutL, which are required for adenine methylation-directed mismatch repair (MMR), enhance VSP repair. Multicopy plasmids carrying mutS alleles that are dominant negative for MMR were tested for their effects on VSP repair. Some mutS mutations (class I) did not lower VSP repair in a mutS

show abstract

Overexpression of vsr in Escherichia coli is mutagenic

Cited by 31 publications

References 20 publications

Very-Short-Patch Repair in Escherichia coli Requires the dam Adenine Methylase

Very-Short-Patch Repair in Escherichia coli Requires the dam Adenine Methylase

Mutator Effects in Escherichia coli Caused by the Expression of Specific Foreign Genes

Interaction of MutS and Vsr: Some Dominant-Negative mutS Mutations That Disable Methyladenine-Directed Mismatch Repair Are Active in Very-Short-Patch Repair

Contact Info

Product

Resources

About