scholarly journals Base Composition of Mononucleotide Runs Affects DNA Polymerase Slippage and Removal of Frameshift Intermediates by Mismatch Repair in Saccharomyces cerevisiae

2002 ◽  
Vol 22 (24) ◽  
pp. 8756-8762 ◽  
Author(s):  
Hana Gragg ◽  
Brian D. Harfe ◽  
Sue Jinks-Robertson
Keyword(s):  
Mismatch Repair ◽  
Dna Polymerase ◽  
Genome Stability ◽  
The Status ◽  
Nucleotide Insertion ◽  
Mismatch Recognition ◽  
Mutation Assay ◽  
Simple Repeats ◽  
Forward Mutation ◽  
Polymerase Slippage

ABSTRACT The postreplicative mismatch repair (MMR) system is important for removing mutational intermediates that are generated during DNA replication, especially those that arise as a result of DNA polymerase slippage in simple repeats. Here, we use a forward mutation assay to systematically examine the accumulation of frameshift mutations within mononucleotide runs of variable composition in wild-type and MMR-defective yeast strains. These studies demonstrate that (i) DNA polymerase slippage occurs more often in 10-cytosine/10-guanine (10C/10G) runs than in 10-adenine/10-thymine (10A/10T) runs, (ii) the MMR system removes frameshift intermediates in 10A/10T runs more efficiently than in 10C/10G runs, (iii) the MMR system removes −1 frameshift intermediates more efficiently than +1 intermediates in all 10-nucleotide runs, and (iv) the repair specificities of the Msh2p-Msh3p and Msh2p-Msh6p mismatch recognition complexes with respect to 1-nucleotide insertion/deletion loops vary dramatically as a function of run composition. These observations are relevant to issues of genome stability, with both the rates and types of mutations that accumulate in mononucleotide runs being influenced by the primary sequence of the run as well as by the status of the MMR system.

scholarly journals The TREX2 3′→ 5′ Exonuclease Physically Interacts with DNA Polymerase δ and Increases Its Accuracy

2002 ◽  
Vol 2 ◽  
pp. 275-281 ◽  
Author(s):  
Igor V. Shevelev ◽  
Kristijan Ramadan ◽  
Ulrich Hubscher
Keyword(s):  
Dna Replication ◽  
Dna Polymerase ◽  
Error Rates ◽  
High Fidelity ◽  
Replication Machinery ◽  
Dna Polymerase Δ ◽  
Pol I ◽  
Calf Thymus ◽  
Mutation Assay ◽  
Forward Mutation

Proofreading function by the 3′→ 5′ exonuclease of DNA polymerase δ (pol δ) is consistent with the observation that deficiency of the associated exonuclease can lead to a strong mutation phenotype, high error rates during DNA replication, and ultimately cancer. We have isolated pol δdfrom isotonic (pol δi) and detergent (pol δd) calf thymus extracts. Pol δdhad a 20-fold higher ratio of exonuclease to DNA polymerase than pol δi. This was due to the physical association of the TREX2 exonuclease to pol δd, which was missing from pol δi. Pol δdwas fivefold more accurate than pol δiunder error-prone conditions (1 μM dGTP and 20 dATP, dCTP, and dTTP) in a M13mp2 DNA forward mutation assay, and fourfold more accurate in an M13mp2T90 reversion assay. Under error-free conditions (20 μM each of the four dNTPs), however, both polymerases showed equal fidelity. Our data suggested that autonomous 3′→ 5′ exonucleases, such as TREX2, through its association with pol I can guarantee high fidelity under difficult conditions in the cell (e.g., imbalance of dNTPs) and can add to the accuracy of the DNA replication machinery, thus preventing mutagenesis.

scholarly journals Role of DNA polymerase β oxidized nucleotide insertion in DNA ligation failure

2017 ◽  
Vol 58 (5) ◽  
pp. 603-607 ◽  
Author(s):  
Melike Çağlayan ◽  
Samuel H Wilson
Keyword(s):  
Dna Polymerase ◽  
Genome Stability ◽  
Excision Repair ◽  
Cancer Therapeutics ◽  
Repair System ◽  
Dna Polymerase Β ◽  
Dna Base Excision Repair ◽  
Dna Base ◽  
Nucleotide Insertion ◽  
Base Excision

Abstract Production of reactive oxygen and nitrogen species (ROS), such as hydrogen peroxide, superoxide and hydroxyl radicals, has been linked to cancer, and these oxidative molecules can damage DNA. Base excision repair (BER), a major repair system maintaining genome stability over a lifespan, has an important role in repairing oxidatively induced DNA damage. Failure of BER leads to toxic consequences in ROS-exposed cells, and ultimately can contribute to the pathobiology of disease. In our previous report, we demonstrated that oxidized nucleotide insertion by DNA polymerase β (pol β) impairs BER due to ligation failure and leads to formation of a cytotoxic repair intermediate. Biochemical and cytotoxic effects of ligation failure could mediate genome stability and influence cancer therapeutics. In this review, we discuss the importance of coordination between pol β and DNA ligase I during BER, and how this could be a fundamental mechanism underlying human diseases such as cancer and neurodegeneration. A summary of this work was presented in a symposium at the International Congress of Radiation Research 2015 in Kyoto, Japan.

scholarly journals Mismatch Repair: From Preserving Genome Stability to Enabling Mutation Studies in Real-Time Single Cells

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1535
Author(s):  
Marina Elez
Keyword(s):  
Real Time ◽  
Mismatch Repair ◽  
Genetic Information ◽  
Genome Stability ◽  
Direct Evidence ◽  
Single Cells ◽  
Living Cells ◽  
Spontaneous Mutations ◽  
Mismatch Recognition

Mismatch Repair (MMR) is an important and conserved keeper of the maintenance of genetic information. Miroslav Radman’s contributions to the field of MMR are multiple and tremendous. One of the most notable was to provide, along with Bob Wagner and Matthew Meselson, the first direct evidence for the existence of the methyl-directed MMR. The purpose of this review is to outline several aspects and biological implications of MMR that his work has helped unveil, including the role of MMR during replication and recombination editing, and the current understanding of its mechanism. The review also summarizes recent discoveries related to the visualization of MMR components and discusses how it has helped shape our understanding of the coupling of mismatch recognition to replication. Finally, the author explains how visualization of MMR components has paved the way to the study of spontaneous mutations in living cells in real time.

scholarly journals The Polymerase η Translesion Synthesis DNA Polymerase Acts Independently of the Mismatch Repair System To Limit Mutagenesis Caused by 7,8-Dihydro-8-Oxoguanine in Yeast

2009 ◽  
Vol 29 (19) ◽  
pp. 5316-5326 ◽  
Author(s):  
Sarah V. Mudrak ◽  
Caroline Welz-Voegele ◽  
Sue Jinks-Robertson
Keyword(s):  
Dna Replication ◽  
Mismatch Repair ◽  
Dna Polymerase ◽  
Translesion Synthesis ◽  
Nuclear Antigen ◽  
Repair System ◽  
New Paradigm ◽  
Mismatch Repair System ◽  
Mutation Assay ◽  
Proliferating Cell

ABSTRACT Reactive oxygen species are ubiquitous mutagens that have been linked to both disease and aging. The most studied oxidative lesion is 7,8-dihydro-8-oxoguanine (GO), which is often miscoded during DNA replication, resulting specifically in GC → TA transversions. In yeast, the mismatch repair (MMR) system repairs GO·A mismatches generated during DNA replication, and the polymerase η (Polη) translesion synthesis DNA polymerase additionally promotes error-free bypass of GO lesions. It has been suggested that Polη limits GO-associated mutagenesis exclusively through its participation in the filling of MMR-generated gaps that contain GO lesions. In the experiments reported here, the SUP4-o forward-mutation assay was used to monitor GC → TA mutation rates in strains defective in MMR (Msh2 or Msh6) and/or in Polη activity. The results clearly demonstrate that Polη can function independently of the MMR system to prevent GO-associated mutations, presumably through preferential insertion of cytosine opposite replication-blocking GO lesions. Furthermore, the Polη-dependent bypass of GO lesions is more efficient on the lagging strand of replication and requires an interaction with proliferating cell nuclear antigen. These studies establish a new paradigm for the prevention of GO-associated mutagenesis in eukaryotes.

Sequence Composition and Context Effects on the Generation and Repair of Frameshift Intermediates in Mononucleotide Runs in Saccharomyces cerevisiae

Genetics ◽  
2000 ◽  
Vol 156 (2) ◽  
pp. 571-578
Author(s):  
Brian D Harfe ◽  
Sue Jinks-Robertson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mismatch Repair ◽  
Dna Polymerase ◽  
Context Effects ◽  
Complete Elimination ◽  
Sequence Context ◽  
Frameshift Mutations ◽  
Sequence Composition ◽  
Overall Efficiency ◽  
Polymerase Slippage

Abstract DNA polymerase slippage occurs frequently in tracts of a tandemly repeated nucleotide, and such slippage events can be genetically detected as frameshift mutations. In long mononucleotide runs, most frameshift intermediates are repaired by the postreplicative mismatch repair (MMR) machinery, rather than by the exonucleolytic proofreading activity of DNA polymerase. Although mononucleotide runs are hotspots for polymerase slippage events, it is not known whether the composition of a run and the surrounding context affect the frequency of slippage or the efficiency of MMR. To address these issues, 10-nucleotide (10N) runs were inserted into the yeast LYS2 gene to create +1 frameshift alleles. Slippage events within these runs were detected as Lys+ revertants. 10G or 10C runs were found to be more unstable than 10A or 10T runs, but neither the frequency of polymerase slippage nor the overall efficiency of MMR was greatly influenced by sequence context. Although complete elimination of MMR activity (msh2 mutants) affected all runs similarly, analyses of reversion rates in msh3 and msh6 mutants revealed distinct specificities of the yeast Msh2p-Msh3p and Msh2p-Msh6p mismatch binding complexes in the repair of frameshift intermediates in different sequence contexts.

scholarly journals Mismatch Repair Status Characterization in Oncologic Pathology: Taking Stock of the Real-World Possibilities

2021 ◽  
Vol 2 (2) ◽  
pp. 93-100
Author(s):  
Roberto Piciotti ◽  
Konstantinos Venetis ◽  
Elham Sajjadi ◽  
Nicola Fusco
Keyword(s):  
Mismatch Repair ◽  
Dna Polymerase ◽  
Genome Stability ◽  
Testing Methods ◽  
Mmr Genes ◽  
Status Assessment ◽  
Tailored Approach ◽  
Mismatch Repair Status ◽  
Tumor Types

The mismatch repair (MMR) system has a key role in supporting the DNA polymerase proofreading function and in maintaining genome stability. Alterations in the MMR genes are driving events of tumorigenesis, tumor progression, and resistance to therapy. These genetic scars may occur in either hereditary or sporadic settings, with different frequencies across tumor types. Appropriate characterization of the MMR status is a crucial task in oncologic pathology because it allows for both the tailored clinical management of cancer patients and surveillance of individuals at risk. The currently available MMR testing methods have specific strengths and weaknesses, and their application across different tumor types would require a tailored approach. This article highlights the indications and challenges in MMR status assessment for molecular pathologists, focusing on the possible strategies to overcome analytical and pre-analytical issues.

scholarly journals Mitotic crossovers between diverged sequences are regulated by mismatch repair proteins in Saccaromyces cerevisiae.

1996 ◽  
Vol 16 (3) ◽  
pp. 1085-1093 ◽  
Author(s):  
A Datta ◽  
A Adjiri ◽  
L New ◽  
G F Crouse ◽  
S Jinks Robertson
Keyword(s):  
Mismatch Repair ◽  
Genome Stability ◽  
Multiple Roles ◽  
Homologous Sequences ◽  
Mismatch Repair Proteins ◽  
Repair Systems ◽  
The Stability ◽  
Saccaromyces Cerevisiae ◽  
Simple Repeats ◽  
Repair Genes

Mismatch repair systems correct replication- and recombination-associated mispaired bases and influence the stability of simple repeats. These systems thus serve multiple roles in maintaining genetic stability in eukaryotes, and human mismatch repair defects have been associated with hereditary predisposition to cancer. In prokaryotes, mismatch repair systems also have been shown to limit recombination between diverged (homologous) sequences. We have developed a unique intron-based assay system to examine the effects of yeast mismatch repair genes (PMS1, MSH2, and MSH3) on crossovers between homologous sequences. We find that the apparent antirecombination effects of mismatch repair proteins in mitosis are related to the degree of substrate divergence. Defects in mismatch repair can elevate homologous recombination between 91% homologous substrates as much as 100-fold while having only modest effects on recombination between 77% homologous substrates. These observations have implications for genome stability and general mechanisms of recombination in eukaryotes.

scholarly journals Pol32, a Subunit of Saccharomyces cerevisiae DNA Polymerase δ, Suppresses Genomic Deletions and Is Involved in the Mutagenic Bypass Pathway

Genetics ◽  
2002 ◽  
Vol 160 (4) ◽  
pp. 1409-1422 ◽  
Author(s):  
Meng-Er Huang ◽  
Anne-Gaëlle Rio ◽  
Marie-Dominique Galibert ◽  
Francis Galibert
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Polymerase ◽  
Genome Stability ◽  
Double Mutant ◽  
Direct Repeats ◽  
Genomic Deletions ◽  
Mutator Effect ◽  
A Subunit ◽  
Mutation Spectra ◽  
Forward Mutation

Abstract The Pol32 subunit of S. cerevisiae DNA polymerase (Pol) δ plays an important role in replication and mutagenesis. Here, by measuring the CAN1 forward mutation rate, we found that either POL32 or REV3 (which encodes the Pol ζ catalytic subunit) inactivation produces overlapping antimutator effects against rad mutators belonging to three epistasis groups. In contrast, the msh2Δ pol32Δ double mutant exhibits a synergistic mutator phenotype. Canr mutation spectrum analysis of pol32Δ strains revealed a substantial increase in the frequency of deletions and duplications (primarily deletions) of sequences flanked by short direct repeats, which appears to be RAD52 and RAD10 independent. To better understand the pol32Δ and rev3Δ antimutator effects in rad backgrounds and the pol32Δ mutator effect in a msh2Δ background, we determined Canr mutation spectra for rad5Δ, rad5Δ pol32Δ, rad5Δ rev3Δ, msh2Δ, msh2Δ pol32Δ, and msh2Δ rev3Δ strains. Both rad5Δ pol32Δ and rad5Δ rev3Δ mutants exhibit a reduction in frameshifts and base substitutions, attributable to antimutator effects conferred by the pol32Δ and rev3Δ mutations. In contrast, an increase in these two types of alterations is attributable to a synergistic mutator effect between the pol32Δ and msh2Δ mutations. Taken together, these observations indicate that Pol32 is important in ensuring genome stability and in mutagenesis.

Sign in / Sign up

Export Citation Format

Share Document