scholarly journals Overexpression of MutS impairs DNA mismatch repair and causes cell division defect in E.coli

2020 ◽  
Author(s):  
Rajesh V Iyer ◽  
Shivranjani C Moharir ◽  
Satish Kumar
Keyword(s):  
Cell Division ◽  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Fold Increase ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Dna Mismatch ◽  
Dna Mismatches ◽  
Post Transcriptional Regulation

MutS and its homologues, from prokaryotes to humans, recognize and bind to DNA mismatches generated during DNA replication, initiate DNA mismatch repair and ensures 100-200 fold increase in replication fidelity. In E.coli, through post transcriptional regulation, at least three mechanisms mediate decline of MutS intracellular concentrations during stress conditions. To understand the significance of this multifold regulation, we overexpressed MutS in E.coli and found that it led to impairment of DNA mismatch repair as reflected by preferential accumulation of transition mutations in spontaneous base pair substitution spectrum. This phenomenon was dependent on MutS-mismatch affinity and interaction. Higher MutS overexpression levels promoted DNA double strand breaks, inhibited cell division and resultantly caused a manifold increase in E.coli cell length. This cell division defect involved a novel MutS-FtsZ interaction and impediment of FtsZ ring function. Our findings may have relevance for cancers where mismatch proteins are known to be overexpressed.

scholarly journals DNA mismatch repair-induced double-strand breaks

DNA Repair ◽  
2008 ◽  
Vol 7 (1) ◽  
pp. 48-56 ◽  
Author(s):  
Anetta Nowosielska ◽  
M.G. Marinus
Keyword(s):  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Double Strand Breaks ◽  
Strand Breaks ◽  
Dna Mismatch

scholarly journals Bi-directional routing of DNA mismatch repair protein human exonuclease 1 to replication foci and DNA double strand breaks

DNA Repair ◽  
2011 ◽  
Vol 10 (1) ◽  
pp. 73-86 ◽  
Author(s):  
Sascha E. Liberti ◽  
Sofie D. Andersen ◽  
Jing Wang ◽  
Alfred May ◽  
Simona Miron ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Dna Mismatch ◽  
Repair Protein ◽  
Directional Routing ◽  
Replication Foci ◽  
Exonuclease 1 ◽  
Mismatch Repair Protein

scholarly journals Genetic evidence for the involvement of mismatch repair proteins, PMS2 and MLH3, in a late step of homologous recombination

2020 ◽  
Vol 295 (51) ◽  
pp. 17460-17475
Author(s):  
Md Maminur Rahman ◽  
Mohiuddin Mohiuddin ◽  
Islam Shamima Keka ◽  
Kousei Yamada ◽  
Masataka Tsuda ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Mismatch Repair ◽  
Ectopic Expression ◽  
Point Mutations ◽  
Sister Chromatid Exchanges ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Γ Irradiation ◽  
Strand Breaks ◽  
Homologous Sequences

Homologous recombination (HR) repairs DNA double-strand breaks using intact homologous sequences as template DNA. Broken DNA and intact homologous sequences form joint molecules (JMs), including Holliday junctions (HJs), as HR intermediates. HJs are resolved to form crossover and noncrossover products. A mismatch repair factor, MLH3 endonuclease, produces the majority of crossovers during meiotic HR, but it remains elusive whether mismatch repair factors promote HR in nonmeiotic cells. We disrupted genes encoding the MLH3 and PMS2 endonucleases in the human B cell line, TK6, generating null MLH3−/− and PMS2−/− mutant cells. We also inserted point mutations into the endonuclease motif of MLH3 and PMS2 genes, generating endonuclease death MLH3DN/DN and PMS2EK/EK cells. MLH3−/− and MLH3DN/DN cells showed a very similar phenotype, a 2.5-fold decrease in the frequency of heteroallelic HR-dependent repair of restriction enzyme–induced double-strand breaks. PMS2−/− and PMS2EK/EK cells showed a phenotype very similar to that of the MLH3 mutants. These data indicate that MLH3 and PMS2 promote HR as an endonuclease. The MLH3DN/DN and PMS2EK/EK mutations had an additive effect on the heteroallelic HR. MLH3DN/DN/PMS2EK/EK cells showed normal kinetics of γ-irradiation–induced Rad51 foci but a significant delay in the resolution of Rad51 foci and a 3-fold decrease in the number of cisplatin-induced sister chromatid exchanges. The ectopic expression of the Gen1 HJ re-solvase partially reversed the defective heteroallelic HR of MLH3DN/DN/PMS2EK/EK cells. Taken together, we propose that MLH3 and PMS2 promote HR as endonucleases, most likely by processing JMs in mammalian somatic cells.

scholarly journals WRN helicase promotes repair of DNA double-strand breaks caused by aberrant mismatch repair of chromium-DNA adducts

Cell Cycle ◽  
2009 ◽  
Vol 8 (17) ◽  
pp. 2769-2778 ◽  
Author(s):  
Alma Zecevic ◽  
Haley Menard ◽  
Volkan Gurel ◽  
Elizabeth Hagan ◽  
Rosamaria DeCaro ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Dna Adducts ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Wrn Helicase

scholarly journals Stability across the Whole Nuclear Genome in the Presence and Absence of DNA Mismatch Repair

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1224
Author(s):  
Scott Alexander Lujan ◽  
Thomas A. Kunkel
Keyword(s):  
Cell Division ◽  
Cell Line ◽  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Nuclear Genome ◽  
Mutation Accumulation ◽  
Mutation Rates ◽  
Wild Type ◽  
Dna Mismatch ◽  
Chicken Cell

We describe the contribution of DNA mismatch repair (MMR) to the stability of the eukaryotic nuclear genome as determined by whole-genome sequencing. To date, wild-type nuclear genome mutation rates are known for over 40 eukaryotic species, while measurements in mismatch repair-defective organisms are fewer in number and are concentrated on Saccharomyces cerevisiae and human tumors. Well-studied organisms include Drosophila melanogaster and Mus musculus, while less genetically tractable species include great apes and long-lived trees. A variety of techniques have been developed to gather mutation rates, either per generation or per cell division. Generational rates are described through whole-organism mutation accumulation experiments and through offspring–parent sequencing, or they have been identified by descent. Rates per somatic cell division have been estimated from cell line mutation accumulation experiments, from systemic variant allele frequencies, and from widely spaced samples with known cell divisions per unit of tissue growth. The latter methods are also used to estimate generational mutation rates for large organisms that lack dedicated germlines, such as trees and hyphal fungi. Mechanistic studies involving genetic manipulation of MMR genes prior to mutation rate determination are thus far confined to yeast, Arabidopsis thaliana, Caenorhabditis elegans, and one chicken cell line. A great deal of work in wild-type organisms has begun to establish a sound baseline, but far more work is needed to uncover the variety of MMR across eukaryotes. Nonetheless, the few MMR studies reported to date indicate that MMR contributes 100-fold or more to genome stability, and they have uncovered insights that would have been impossible to obtain using reporter gene assays.

scholarly journals Crosstalk between repair pathways elicits Double-Strand Breaks in alkylated DNA: implications for the action of temozolomide

2020 ◽  
Author(s):  
Robert P. Fuchs ◽  
Asako Isogawa ◽  
Joao A. Paulo ◽  
Kazumitsu Onizuka ◽  
Tatsuro Takahashi ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Futile Cycle ◽  
Repair Synthesis ◽  
Strand Breaks ◽  
Egg Extracts ◽  
Established Cell ◽  
Cycle Dependent ◽  
Xenopus Egg Extracts

AbstractTemozolomide, a DNA methylating agent, is the primary chemotherapeutic drug used in glioblastoma treatment. TMZ induces mostly N-alkylation adducts (N7-methylguanine and N3-methyladenine) and some O6-methylguanine (O6mG). Current models propose that during DNA replication, thymine is incorporated across from O6mG, promoting a futile cycle of mismatch repair (MMR) that leads to DNA double strand breaks (DSBs). To revisit the mechanism of O6mG processing, we reacted plasmid DNA with N-Methyl-N-nitrosourea (MNU), a temozolomide mimic, and incubated it in Xenopus egg extracts. We show that in this system, mismatch repair (MMR) proteins are enriched on MNU-treated DNA and we observe robust, MMR-dependent, repair synthesis. Our evidence also suggests that MMR, initiated at O6mG:C sites, is strongly stimulated in cis by repair processing of other lesions, such as N-alkylation adducts. Importantly, MNU-treated plasmids display DSBs in extracts, the frequency of which increased linearly with the square of alkylation dose. We suggest that DSBs result from two independent repair processes, one involving MMR at O6mG:C sites and the other involving BER acting at a nearby N-alkylation adducts. We propose a new, replication-independent mechanism of action of TMZ, that operates in addition to the well-established cell cycle dependent mode of action.

scholarly journals Crosstalk between repair pathways elicits Double Strand Breaks in alkylated DNA and implications for the action of temozolomide

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Robert P Fuchs ◽  
Asako Isogawa ◽  
Joao A Paulo ◽  
Kazumitsu Onizuka ◽  
Tatsuro Takahashi ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Futile Cycle ◽  
Repair Synthesis ◽  
Strand Breaks ◽  
Independent Mechanism ◽  
Xenopus Egg ◽  
Cycle Dependent ◽  
Mmr Proteins

Temozolomide (TMZ), a DNA methylating agent, is the primary chemotherapeutic drug used in glioblastoma treatment. TMZ induces mostly N-alkylation adducts (N7-methylguanine and N3-methyladenine) and some O6-methylguanine (O6mG). Current models propose that during DNA replication, thymine is incorporated across from O6mG, promoting a futile cycle of mismatch repair (MMR) that leads to DNA double strand breaks (DSBs). To revisit the mechanism of O6mG processing, we reacted plasmid DNA with N-Methyl-N-nitrosourea (MNU), a temozolomide mimic, and incubated it in Xenopus egg-derived extracts. We show that in this system, mismatch repair (MMR) proteins are enriched on MNU-treated DNA and we observe robust, MMR-dependent, repair synthesis. Our evidence also suggests that MMR, initiated at O6mG:C sites, is strongly stimulated in cis by repair processing of other lesions, such as N-alkylation adducts. Importantly, MNU-treated plasmids display DSBs in extracts, the frequency of which increased linearly with the square of alkylation dose. We suggest that DSBs result from two independent repair processes, one involving MMR at O6mG:C sites and the other involving BER acting at a nearby N-alkylation adducts. We propose a new, replication-independent mechanism of action of TMZ, that operates in addition to the well-studied cell cycle dependent mode of action.

scholarly journals Recombination Is Essential for Viability of anEscherichia coli dam (DNA Adenine Methyltransferase) Mutant

2000 ◽  
Vol 182 (2) ◽  
pp. 463-468 ◽  
Author(s):  
M. G. Marinus
Keyword(s):  
Double Strand Breaks ◽  
Holliday Junction ◽  
Gene Products ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
Strand Breaks ◽  
Dna Mismatch ◽  
Dna Damaging Agents ◽  
Ultimate Source ◽  
Dna Adenine Methyltransferase

ABSTRACT Double mutants of Escherichia coli dam (DNA adenine methyltransferase) strains with ruvA, ruvB, orruvC could not be constructed, whereas damderivatives with recD, recF, recJ, and recR were viable. The ruv gene products are required for Holliday junction translocation and resolution of recombination intermediates. A dam recG (Holliday junction translocation) mutant strain was isolated but at a very much lower frequency than expected. The inviability of a dam lexA(Ind−) host was abrogated by the simultaneous presence of plasmids encoding both recA and ruvAB. This result indicates that of more than 20 SOS genes, only recAand ruvAB need to be derepressed to allow fordam mutant survival. The presence of mutS ormutL mutations allowed the construction of dam lexA (Ind−) derivatives. The requirement forrecA, recB, recC, ruvA,ruvB, ruvC, and possibly recG gene expression indicates that recombination is essential for viability ofdam bacteria probably to repair DNA double-strand breaks. The effect of mutS and mutL mutations indicates that DNA mismatch repair is the ultimate source of most of these DNA breaks. The requirement for recombination also suggests an explanation for the sensitivity of dam cells to certain DNA-damaging agents.

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