Mismatch repair and the fidelity of genetic recombination

Two modes of mismatch repair are known to operate in bacteria: long-patch mismatch repair and very short patch mismatch repair. Very short patch mismatch repair systems act on a specific mismatch by conserving only one base pair. Therefore, when very short patch mismatch repair acts on heteroduplex recombination intermediates, it hyper-recombines specific markers by creating patchwork sequences, i.e., apparent multiple exchange events, on the repaired strand. Long-patch mismatch repair is antirecombinagenic, apparently by decomposing heteroduplex DNA or aborting its formation whenever well-recognized mismatches are formed by strand exchange between nonidentical parental sequences. It is postulated here that mismatch-stimulated antirecombination by long-patch mismatch repair is a "proofreading" system assuring high fidelity of homologous recombination. This accounts for chromosomal stability in eucaryotes (i.e., the rare occurrence of chromosomal aberrations and mitotic recombination versus the high frequency of precise sister chromatid exchange), suggests a role for diverged repetitive and other noncoding sequences as chromosomal antirecombination elements, and provides a molecular mechanism for speciation without the necessity of geographical separation.     

Mismatch repair and homeologous recombination

DNA mismatch repair influences the outcome of recombination events between diverging DNA sequences. Here we discuss how mismatch repair proteins are active in different homologous recombination subpathways and specific reaction steps, resulting in differential modulation of these recombination events, with a focus on the mechanism of heteroduplex rejection during the inhibition of recombination between slightly diverged (homeologous) DNA sequences.     

Mismatch repair and recombination in E. coli

The involvement of the E. coli methyl-directed and very short patch (vsp) mismatch repair systems in bacteriophage lambda recombination has been studied. Genetic crosses and heteroduplex transfection experiments were performed using lambda phages with sequenced mutations in the cl gene. The results indicate that methyl-directed repair does operate during bacteriophage lambda recombination but generally does not contribute to the formation of recombinants involving close markers. Vsp repair apparently acts during bacteriophage lambda recombination to produce recombinants involving close markers because its action does not involve extensive excision tracts. Marker-specific hyperrecombination and the apparent clustering of genetic exchanges in bacteriophage lambda recombination can be accounted for by the action of the vsp repair system.     

Mismatch repair proteins and mitotic genome stability

Mismatch repair (MMR) proteins play a critical role in maintaining the mitotic stability of eukaryotic genomes. MMR proteins repair errors made during DNA replication and in their absence, mutations accumulate at elevated rates. In addition, MMR proteins inhibit recombination between non-identical DNA sequences, and hence prevent genome rearrangements resulting from interactions between repetitive elements. This review provides an overview of the anti-mutator and anti-recombination functions of MMR proteins in the yeast Saccharomyces cerevisiae.     

Rab proteins: The key regulators of intracellular vesicle transport

Vesicular/membrane trafficking essentially regulates the compartmentalization and abundance of proteins within the cells and contributes in many signalling pathways. This membrane transport in eukaryotic cells is a complex process regulated by a large and diverse array of proteins. A large group of monomeric small GTPases; the Rabs are essential components of this membrane trafficking route. Most of the Rabs are ubiquitously expressed proteins and have been implicated in vesicle formation, vesicle motility/delivery along cytoskeleton elements and docking/fusion at target membranes through the recruitment of effectors. Functional impairments of Rabs affecting transport pathways manifest different diseases. Rab functions are accompanied by cyclical activation and inactivation of GTP-bound and GDP-bound forms between the cytosol and membranes which is regulated by upstream regulators. Rab proteins are characterized by their distinct sub-cellular localization and regulate a wide variety of endocytic, transcytic and exocytic transport pathways. Mutations of Rabs affect cell growth, motility and other biological processes.     

Mismatch repair

Specific repair systems are activated in response to the DNA damage. Mismatch repair protects the genome of prokaryotic and eukaryotic cells from lesions that appear during process of DNA replication or are induced by mutagenic factors. The methyl directed mismatch repair distinguishes the new strand from the old strand by the hemi-methylated state of the DNA and controls the fidelity of genetic information after homologous recombination. The very short patch repair restores the mismatches at the sites with nucleotide sequence CC(W/T)GG. The "8-oxoG" pathway is independent of the hemi-methylated state of the DNA, and removes the oxidated nucleotides from the genome of prokaryotes and eukaryotes. Mutations in genes of mismatch repair enhance the process of mutagenesis in prokaryotic cell, and are the reason for the development of the colon cancer in humans. The mechanisms of mismatch repair and the role of defective repair proteins in mutagenesis and carcinogenesis are discussed in this review.     

Meiotic recombination intermediates and mismatch repair proteins

Mismatch repair proteins are a highly diverse group of proteins that interact with numerous DNA structures during DNA repair and replication. Here we review data for the role of Msh4, Msh5, Mlh1, Mlh3 and Exo1 in crossing over. Based on the paradigm of interactions developed from studies of mismatch repair, we propose models for the mechanism of crossover implementation by Msh4/Msh5 and Mlh1/Mlh3.     

Mismatch repair

Specific repair systems are activated in response to DNA lesions. Mismatch repair protects the genome of prokaryotic and eukaryotic cells from errors arising during replication or induced by mutagenic factors. The mismatch repair system distinguishes between the newly synthesized and pattern DNA strands by the extent of methylation and checks the accuracy of genetic information after homologous recombination. Very short-patch repair corrects mismatches in CC(A/T)GG sites. The 8-oxoguanine system is independent of DNA hemimethylation and removes oxidized bases from prokaryotic and eukaryotic genomes. Mutations of repair genes increase mutagenesis in prokaryotic cells and cause colorectal cancer in humans. The review considers the repair mechanisms and the role of repair defects in mutagenesis and carcinogenesis.     

Mismatch repair converts AID-instigated nicks to double-strand breaks for antibody class-switch recombination

Mismatch repair (MMR) proteins are important for antibody class-switch recombination (CSR), but their roles are unknown. We propose a model for the function of MMR in CSR in which MMR proteins convert single-strand nicks instigated by activation-induced cytidine deaminase (AID) into the double-strand breaks (DSBs) that are required for CSR. This model does not invoke any novel functions for MMR but simply posits that, owing to numerous single-strand nicks in the switch (S) regions of both DNA strands, when MMR proteins are recruited by U:G mismatches, they excise one strand of DNA and soon reach a nick on the opposite strand. This halts excision activity and creates a DSB. This model explains why B cells that lack either S mu and MSH2 or UNG and MSH2 cannot undergo CSR.     

Mismatch repair of heteroduplex DNA intermediates of extrachromosomal recombination in mammalian cells.

Previous work indicated that extrachromosomal recombination in mammalian cells could be explained by the single-strand annealing (SSA) model. This model predicts that extrachromosomal recombination leads to nonconservative crossover products and that heteroduplex DNA (hDNA) is formed by annealing of complementary single strands. Mismatched bases in hDNA may subsequently be repaired to wild-type or mutant sequences, or they may remain unrepaired and segregate following DNA replication. We describe a system to examine the formation and mismatch repair of hDNA in recombination intermediates. Our results are consistent with extrachromosomal recombination occurring via SSA and producing crossover recombinant products. As predicted by the SSA model, hDNA was present in double-strand break-induced recombination intermediates. By placing either silent or frameshift mutations in the predicted hDNA region, we have shown that mismatches are efficiently repaired prior to DNA replication.     

Mismatch repair proteins are activators of toxic responses to chromium-DNA damage

Chromium(VI) is a toxic and carcinogenic metal that causes the formation of DNA phosphate-based adducts. Cr-DNA adducts are genotoxic in human cells, although they do not block replication in vitro. Here, we report that induction of cytotoxicity in Cr(VI)-treated human colon cells and mouse embryonic fibroblasts requires the presence of all major mismatch repair (MMR) proteins. Cr-DNA adducts lost their ability to block replication of Cr-modified plasmids in human colon cells lacking MLH1 protein. The presence of functional mismatch repair caused induction of p53-independent apoptosis associated with activation of caspases 2 and 7. Processing of Cr-DNA damage by mismatch repair resulted in the extensive formation of gamma-H2AX foci in G(2) phase, indicating generation of double-stranded breaks as secondary toxic lesions. Induction of gamma-H2AX foci was observed at 6 to 12 h postexposure, which was followed by activation of apoptosis in the absence of significant G(2) arrest. Our results demonstrate that mismatch repair system triggers toxic responses to Cr-DNA backbone modifications through stress mechanisms that are significantly different from those for other forms of DNA damage. Selection for Cr(VI) resistant, MMR-deficient cells may explain the very high frequency of lung cancers with microsatellite instability among chromate workers.     

Mismatch repair factor MSH2-MSH3 binds and alters the conformation of branched DNA structures predicted to form during genetic recombination

Genetic studies in Saccharomyces cerevisiae predict that the mismatch repair (MMR) factor MSH2-MSH3 binds and stabilizes branched recombination intermediates that form during single strand annealing and gene conversion. To test this model, we constructed a series of DNA substrates that are predicted to form during these recombination events. We show in an electrophoretic mobility shift assay that S. cerevisiae MSH2-MSH3 specifically binds branched DNA substrates containing 3' single-stranded DNA and that ATP stimulates its release from these substrates. Chemical footprinting analyses indicate that MSH2-MSH3 specifically binds at the double-strand/single-strand junction of branched substrates, alters its conformation and opens up the junction. Therefore, MSH2-MSH3 binding to its substrates creates a unique nucleoprotein structure that may signal downstream steps in repair that include interactions with MMR and nucleotide excision repair factors.     

TRAF proteins as key regulators of plant development and stress responses

Tumor necrosis factor receptor-associated factor (TRAF) proteins are conserved in higher eukaryotes and play key roles in transducing cellular signals across different organelles. They are characterized by their C-terminal region (TRAF-C domain) containing seven to eight anti-parallel β-sheets, also known as the meprin and TRAF-C homology (MATH) domain. Over the past few decades, significant progress has been made toward understanding the diverse roles of TRAF proteins in mammals and plants. Compared to other eukaryotic species, the Arabidopsis thaliana and rice (Oryza sativa) genomes encode many more TRAF/MATH domain-containing proteins; these plant proteins cluster into five classes: TRAF/MATH-only, MATH-BPM, MATH-UBP (ubiquitin protease), Seven in absentia (SINA), and MATH-Filament and MATH-PEARLI-4 proteins, suggesting parallel evolution of TRAF proteins in plants. Increasing evidence now indicates that plant TRAF proteins form central signaling networks essential for multiple biological processes, such as vegetative and reproductive development, autophagosome formation, plant immunity, symbiosis, phytohormone signaling, and abiotic stress responses. Here, we summarize recent advances and highlight future prospects for understanding on the molecular mechanisms by which TRAF proteins act in plant development and stress responses.     

Mismatch repair and nucleotide excision repair proteins cooperate in the recognition of DNA interstrand crosslinks

DNA interstrand crosslinks (ICLs) are among the most cytotoxic types of DNA damage, thus ICL-inducing agents such as psoralen, are clinically useful chemotherapeutics. Psoralen-modified triplex-forming oligonucleotides (TFOs) have been used to target ICLs to specific genomic sites to increase the selectivity of these agents. However, how TFO-directed psoralen ICLs (Tdp-ICLs) are recognized and processed in human cells is unclear. Previously, we reported that two essential nucleotide excision repair (NER) protein complexes, XPA–RPA and XPC–RAD23B, recognized ICLs in vitro, and that cells deficient in the DNA mismatch repair (MMR) complex MutSβ were sensitive to psoralen ICLs. To further investigate the role of MutSβ in ICL repair and the potential interaction between proteins from the MMR and NER pathways on these lesions, we performed electrophoretic mobility-shift assays and chromatin immunoprecipitation analysis of MutSβ and NER proteins with Tdp-ICLs. We found that MutSβ bound to Tdp-ICLs with high affinity and specificity in vitro and in vivo, and that MutSβ interacted with XPA–RPA or XPC–RAD23B in recognizing Tdp-ICLs. These data suggest that proteins from the MMR and NER pathways interact in the recognition of ICLs, and provide a mechanistic link by which proteins from multiple repair pathways contribute to ICL repair.     

Mismatch repair proteins collaborate with methyltransferases in the repair of O(6)-methylguanine

DNA repair is essential for combatting the adverse effects of damage to the genome. One example of base damage is O(6)-methylguanine (O(6)mG), which stably pairs with thymine during replication and thereby creates a promutagenic O(6)mG:T mismatch. This mismatch has also been linked with cellular toxicity. Therefore, in the absence of repair, O(6)mG:T mismatches can lead to cell death or result in G:C-->A:T transition mutations upon the next round of replication. Cysteine thiolate residues on the Ada and Ogt methyltransferase (MTase) proteins directly reverse the O(6)mG base damage to yield guanine. When a cytosine is opposite the lesion, MTase repair restores a normal G:C pairing. However, if replication past the lesion has produced an O(6)mG:T mismatch, MTase conversion to a G:T mispair must still undergo correction to avoid mutation. Two mismatch repair pathways in E. coli that convert G:T mispairs to native G:C pairings are methyl-directed mismatch repair (MMR) and very short patch repair (VSPR). This work examined the possible roles that proteins in these pathways play in coordination with the canonical MTase repair of O(6)mG:T mismatches. The possibility of this repair network was analyzed by probing the efficiency of MTase repair of a single O(6)mG residue in cells deficient in individual mismatch repair proteins (Dam, MutH, MutS, MutL, or Vsr). We found that MTase repair in cells deficient in Dam or MutH showed wild-type levels of MTase repair. In contrast, cells lacking any of the VSPR proteins MutS, MutL, or Vsr showed a decrease in repair of O(6)mG by the Ada and Ogt MTases. Evidence is presented that the VSPR pathway positively influences MTase repair of O(6)mG:T mismatches, and assists the efficiency of restoring these mismatches to native G:C base pairs.     

幽门螺杆菌对胃癌前期阶段胃上皮DNA同源重组修复关键蛋白的影响

目的 通过检测胃癌前期阶段幽门螺杆菌（Helicobacter pylori,H. pylori）阳性和阴性患者胃黏膜组织中DNA损伤标志物H2AX及同源重组（homologous recombination,HR）修复关键蛋白MRE11、Rad51、CtIP表达水平,评价H. pylori感染在胃癌前期阶段对HR精确修复的影响。方法 选择2017年3月至9月行胃镜及病理检测的165例H. pylori阳性和阴性患者,取胃黏膜上皮组织,石蜡包埋切片,行HE染色,根据世界卫生组织标准和更新的悉尼标准,划分病理类型。然后应用免疫组织化学染色方法检测H. pylori和DNA损伤标记蛋白及HR修复关键蛋白表达水平,并行统计学分析。结果 胃黏膜上皮细胞中H2AX的表达,在CSG、CAG和IM阶段,H. pylori阳性组表达高于阴性组（Mann-Whitney U=1116.5,P=0.001;Mann-Whitney U=185.0,P=0.018;Mann-Whitney U=214.5,P=0.041）,在Dys阶段,H. pylori阳性组和阴性组差异无统计学意义（Mann-Whitney U=35.5,P=0.964）;MRE11的表达,在CSG、CAG阶段,H. pylori阳性组表达高于阴性组（Mann-Whitney U=1117.0,P=0.001;Mann-Whitney U=201.0,P=0.002）,在IM、Dys阶段,H. pylori阳性组和阴性组差异无统计学意义（Mann-Whitney U=171.0,P=0.568;Mann-Whitney U=41.5,P=0.616）;Rad51的表达,在CSG、IM阶段,H. pylori阳性组表达低于阴性组（Mann-Whitney U=490.0,P=0.002;Mann-Whitney U=73.0,P=0.007）,在CAG、Dys阶段,H. pylori阳性组和阴性组差异无统计学意义（Mann-Whitney U=101.0,P=0.404;Mann-Whitney U=24.0,P=0.291）;CtIP的表达,在CSG、IM阶段,H. pylori阳性组表达低于阴性组（Mann-Whitney U=593.0,P=0.044;Mann-Whitney U=58.5,P=0.001）,在CAG、Dys阶段,H. pylori阳性组和阴性组差异无统计学意义（Mann-Whitney U=84.0,P=0.136;Mann-Whitney U=18.5,P=0.102）。结论 在胃癌前期发展阶段,H. pylori感染导致人体胃上皮细胞DNA损伤,却抑制部分HR修复通道关键蛋白表达,从而可能抑制精确的HR修复,增加细胞恶变几率。