Bacterial expression of cloned gene hMSH2 in E. coli, of mismatch repair system

Abstract Excessive recombination between repeated, interspersed, and diverged DNA sequences is a potential source of genomic instability. We have investigated the possibility that a mechanism exists to suppress genetic exchange between these quasi-homologous (homeologous) sequences. We examined the role of the general mismatch repair system of Escherichia coli because previous work has shown that the mismatch repair pathway functions as a barrier to interspecies recombination between E. coli and Salmonella typhimurium. The formation of large duplications by homeologous recombination in E. coli was increased some tenfold by mutations in the mutL and mutS genes that encode the mismatch recognition proteins. These findings indicate that the mismatch recognition proteins act to prevent excessive intrachromosomal exchanges. We conclude that mismatch repair proteins serve as general controllers of the fidelity of genetic inheritance, acting to suppress chromosomal rearrangements as well as point mutations.

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A GENETIC ANALYSIS OF PRIMARY PRODUCTS OF BACTERIOPHAGE LAMBDA RECOMBINATION

Genetics ◽

10.1093/genetics/112.3.409 ◽

1986 ◽

Vol 112 (3) ◽

pp. 409-420

Author(s):

Olivier Huisman ◽

Maurice S Fox

Keyword(s):

Genetic Analysis ◽

Mismatch Repair ◽

Visual Detection ◽

Bacteriophage Lambda ◽

Low Frequency ◽

Repair System ◽

Heteroduplex Dna ◽

E Coli ◽

Primary Products ◽

Mismatch Repair System

ABSTRACT Primary products of bacteriophage lambda recombination that display heterozygosity as a consequence of the presence of regions of heteroduplex DNA are rare in standard λ crosses. Phage manifesting heterozygosity at a given allele are evident when recombinants, emerging from a cross, are selected for an exchange in a neighboring interval. We show that the abundance of such heterozygotes can be increased 10- to 20-fold by selection on an E. coli indicator that is defective in methyl-directed mismatch repair (mutL). Thus, the activity of the methyl-directed mismatch repair system is, at least in part, responsible for the low frequency of detectably heterozygous phage emerging from a standard cross. In a mutL indicator, many primary products of recombination are replicated without the intervention of mismatch repair.—The products of a six-factor phage cross have been plated on a mutL indicator allowing visual detection of those phage products heterozygous for one of the allelic pairs, cI. By genetic analysis, we show that the heteroduplex regions of these primary products of recombination are on the average about 4 kb in length and can include as much as half of the lambda genome.

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Some Features of Base Pair Mismatch and Heterology Repair in Escherichia coli

Genetics ◽

10.1093/genetics/117.3.381 ◽

1987 ◽

Vol 117 (3) ◽

pp. 381-390

Author(s):

Susan Raposa ◽

Maurice S Fox

Keyword(s):

Escherichia Coli ◽

Mismatch Repair ◽

Base Pair ◽

Repair System ◽

Loop Structure ◽

E Coli ◽

Base Pair Mismatch ◽

Adenine Methylation ◽

Mismatch Repair System ◽

Lambda Dna

ABSTRACT We have used artificially constructed heteroallelic heteroduplex molecules of bacteriophage lambda DNA to transfect Escherichia coli, and E. coli mutants deficient in various functions involved in the adenine methylation-directed mismatch repair system, MutL, MutS, MutH, and UvrD (MutU). Analysis of the allele content of single infective centers shows that this repair system often acts on several mismatches, separated by as many as 2000 bp, on one of the strands of a heteroduplex molecule. When the methyl-directed mismatch repair system is disabled by mutH or uvrD mutations, localized mismatch repair becomes prominent. This prominent localized repair that can result in separation of very closely linked markers requires the functions MutL and MutS, is independent of adenine methylation, and appears to reflect another mechanism of mismatch repair. Heterology-containing heteroduplex molecules with a deletion in one strand often escape processing. However, when the heterology includes the stem and loop structure of a transposon, Tn10, the transposon is lost.

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Escherichia coli Strains (ndk) Lacking Nucleoside Diphosphate Kinase Are Powerful Mutators for Base Substitutions and Frameshifts in Mismatch-Repair-Deficient Strains

Genetics ◽

10.1093/genetics/162.1.5 ◽

2002 ◽

Vol 162 (1) ◽

pp. 5-13 ◽

Cited By ~ 1

Author(s):

Jeffrey H Miller ◽

Pauline Funchain ◽

Wendy Clendenin ◽

Tiffany Huang ◽

Anh Nguyen ◽

...

Keyword(s):

Escherichia Coli ◽

Mismatch Repair ◽

Nucleoside Diphosphate Kinase ◽

Base Substitution ◽

Repair System ◽

Ndp Kinase ◽

E Coli ◽

Base Substitutions ◽

Mismatch Repair System ◽

Substitution Mutations

Abstract Nucleoside diphosphate (NDP) kinase is one of the enzymes that maintains triphosphate pools. Escherichia coli strains (ndk) lacking this enzyme have been shown to be modest base substitution mutators, and two members of the human family of NDP kinases act as tumor suppressors. We show here that in E. coli strains lacking NDP kinase high levels of mispairs are generated, but most of these are corrected by the mismatch-repair system. Double mutants that are ndk mutS, lacking both the NDP kinase and mismatch repair, have levels of base substitutions 15-fold higher and levels of certain frameshifts up to 10-fold higher than those of the respective mutations in mutS strains that are NDP kinase proficient. A sequence analysis of the specificity of base substitution mutations generated in ndk and ndk mutS backgrounds as well as other experiments suggests that NDP kinase deficiency stimulates polymerase errors that lead to A:T → G:C transitions and that the editing capacity of cells may be affected, leading to additional uncorrected mispairs and to A:T → T:A transversions.

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Functional Specifics of the MutL Protein of the DNA Mismatch Repair System in Different Organisms

Russian Journal of Bioorganic Chemistry ◽

10.1134/s1068162020060217 ◽

2020 ◽

Vol 46 (6) ◽

pp. 875-890

Author(s):

M. V. Monakhova ◽

M. A. Milakina ◽

R. M. Trikin ◽

T. S. Oretskaya ◽

E. A. Kubareva

Keyword(s):

Mismatch Repair ◽

Dna Mismatch Repair ◽

Repair System ◽

Dna Mismatch ◽

Dna Mismatch Repair System ◽

Mismatch Repair System

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Poorly Repaired Mismatches in Heteroduplex DNA are Hyper-Recombinagenic in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/142.2.407 ◽

1996 ◽

Vol 142 (2) ◽

pp. 407-416 ◽

Cited By ~ 3

Author(s):

P Manivasakam ◽

Susan M Rosenberg ◽

P J Hastings

Keyword(s):

Saccharomyces Cerevisiae ◽

Mismatch Repair ◽

Genetic Markers ◽

Meiotic Recombination ◽

Repair System ◽

Normal Allele ◽

Heteroduplex Dna ◽

Mismatch Repair System ◽

Postmeiotic Segregation ◽

System Operating

Abstract In yeast meiotic recombination, alleles used as genetic markers fall into two classes as regards their fate when incorporated into heteroduplex DNA. Normal alleles are those that form heteroduplexes that are nearly always recognized and corrected by the mismatch repair system operating in meiosis. High PMS (postmeiotic segregation) alleles form heteroduplexes that are inefficiently mismatch repaired. We report that placing any of several high PMS alleles very close to normal alleles causes hyperrecombination between these markers. We propose that this hyperrecombination is caused by the high PMS allele blocking a mismatch repair tract initiated from the normal allele, thus preventing corepair of the two alleles, which would prevent formation of recombinants. The results of three point crosses involving two PMS alleles and a normal allele suggest that high PMS alleles placed between two alleles that are normally corepaired block that corepair.

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Saturation of DNA Mismatch Repair and Error Catastrophe by a Base Analogue in Escherichia coli

Genetics ◽

10.1093/genetics/161.4.1363 ◽

2002 ◽

Vol 161 (4) ◽

pp. 1363-1371

Author(s):

Kazuo Negishi ◽

David Loakes ◽

Roel M Schaaper

Keyword(s):

Escherichia Coli ◽

Mismatch Repair ◽

Dna Mismatch Repair ◽

Repair System ◽

Mutagenic Action ◽

Wild Type ◽

Dna Mismatch ◽

Cell Counts ◽

Error Catastrophe ◽

Mismatch Repair System

Abstract Deoxyribosyl-dihydropyrimido[4,5-c][1,2]oxazin-7-one (dP) is a potent mutagenic deoxycytidine-derived base analogue capable of pairing with both A and G, thereby causing G · C → A · T and A · T → G · C transition mutations. We have found that the Escherichia coli DNA mismatch-repair system can protect cells against this mutagenic action. At a low dose, dP is much more mutagenic in mismatch-repair-defective mutH, mutL, and mutS strains than in a wild-type strain. At higher doses, the difference between the wild-type and the mutator strains becomes small, indicative of saturation of mismatch repair. Introduction of a plasmid containing the E. coli mutL+ gene significantly reduces dP-induced mutagenesis. Together, the results indicate that the mismatch-repair system can remove dP-induced replication errors, but that its capacity to remove dP-containing mismatches can readily be saturated. When cells are cultured at high dP concentration, mutant frequencies reach exceptionally high levels and viable cell counts are reduced. The observations are consistent with a hypothesis in which dP-induced cell killing and growth impairment result from excess mutations (error catastrophe), as previously observed spontaneously in proofreading-deficient mutD (dnaQ) strains.

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