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The EMBO Journal Nov 1990Molecular evidence for intrachromosomal recombination between closely linked DNA repeats within the plant genome is presented. The non-overlapping complementary deletion...
Molecular evidence for intrachromosomal recombination between closely linked DNA repeats within the plant genome is presented. The non-overlapping complementary deletion derivatives of the selectable neomycin phosphotransferase gene (nptII), when intact conferring kanamycin resistance, were inserted into the genome of Nicotiana tabacum. The functional marker gene was restored with frequencies between 10(-4) and 10(-6) per proliferating cell clone. Prolonged tissue culture prior to kanamycin selection did not increase the number of recombinant kanamycin-resistant (KanR) cell clones. DNA analysis of KanR clones derived from cells carrying multiple tandem recombination units suggested that these units have a tendency to undergo concerted recombination. Recovery and analysis of kanamycin-sensitive seedlings with patches of KanR cells provided direct evidence for mitotic recombination in plant tissue.
Topics: Base Sequence; Blotting, Southern; Cell Line; Chromosomes; Cloning, Molecular; In Vitro Techniques; Kanamycin Kinase; Meiosis; Mitosis; Molecular Sequence Data; Phosphotransferases; Plants, Toxic; Recombination, Genetic; Restriction Mapping; Nicotiana
PubMed: 2170114
DOI: 10.1002/j.1460-2075.1990.tb07551.x -
DNA Repair Sep 2009Genome integrity is frequently challenged by DNA lesions from both endogenous and exogenous sources. A single DNA double-strand break (DSB) is lethal if unrepaired and... (Review)
Review
Genome integrity is frequently challenged by DNA lesions from both endogenous and exogenous sources. A single DNA double-strand break (DSB) is lethal if unrepaired and may lead to loss of heterozygosity, mutations, deletions, genomic rearrangements and chromosome loss if repaired improperly. Such genetic alterations are the main causes of cancer and other genetic diseases. Consequently, DNA double-strand break repair (DSBR) is an important process in all living organisms. DSBR is also the driving mechanism in most strategies of gene targeting, which has applications in both genetic and clinical research. Here we review the cell biological response to DSBs in mitotically growing cells with an emphasis on homologous recombination pathways in yeast Saccharomyces cerevisiae and in mammalian cells.
Topics: Animals; DNA Breaks, Double-Stranded; DNA Damage; DNA Repair; Humans; Nuclear Proteins; Recombination, Genetic; Signal Transduction
PubMed: 19473884
DOI: 10.1016/j.dnarep.2009.04.007 -
Nucleic Acids Research 2006Homologous recombination (HR) is a ubiquitous cellular pathway that mediates transfer of genetic information between homologous or near homologous (homeologous) DNA... (Review)
Review
Homologous recombination (HR) is a ubiquitous cellular pathway that mediates transfer of genetic information between homologous or near homologous (homeologous) DNA sequences. During meiosis it ensures proper chromosome segregation in the first division. Moreover, HR is critical for the tolerance and repair of DNA damage, as well as in the recovery of stalled and broken replication forks. Together these functions preserve genomic stability and assure high fidelity transmission of the genetic material in the mitotic and meiotic cell divisions. This review will focus on the Rad54 protein, a member of the Snf2-family of SF2 helicases, which translocates on dsDNA but does not display strand displacement activity typical for a helicase. A wealth of genetic, cytological, biochemical and structural data suggests that Rad54 is a core factor of HR, possibly acting at multiple stages during HR in concert with the central homologous pairing protein Rad51.
Topics: Adenosine Triphosphatases; DNA Helicases; DNA Repair; DNA Repair Enzymes; DNA-Binding Proteins; Models, Biological; Rad52 DNA Repair and Recombination Protein; Recombination, Genetic; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Transcription Factors
PubMed: 16935872
DOI: 10.1093/nar/gkl481 -
Mechanisms of Ageing and Development Sep 2010Werner syndrome (WS) is a rare, segmental progeroid syndrome caused by defects in the WRN gene, which encodes a RecQ helicase. WRN has roles in many aspects of DNA...
Werner syndrome (WS) is a rare, segmental progeroid syndrome caused by defects in the WRN gene, which encodes a RecQ helicase. WRN has roles in many aspects of DNA metabolism including DNA repair and recombination. In this study, we exploited two different recombination assays previously used to describe a role for the structure-specific endonuclease ERCC1-XPF in mitotic and targeted homologous recombination. We constructed Chinese hamster ovary (CHO) cell lines isogenic with the cell lines used in these previous studies by depleting WRN using shRNA vectors. When intrachromosomal, mitotic recombination was assayed in WRN-depleted CHO cells, a hyperrecombination phenotype was observed, and a small number of aberrant recombinants were generated. Targeted homologous recombination was also examined in WRN-depleted CHO cells using a plasmid-chromosome targeting assay. In these experiments, loss of WRN resulted in a significant decrease in nonhomologous integration events and ablation of recombinants that required random integration of the corrected targeting vector. Aberrant recombinants were also recovered, but only from WRN-depleted cells. The pleiotropic recombination phenotypes conferred by WRN depletion, reflected in distinct homologous and nonhomologous recombination pathways, suggest a role for WRN in processing specific types of homologous recombination intermediates as well as an important function in nonhomologous recombination.
Topics: Animals; CHO Cells; Chromosomes; Cricetinae; Cricetulus; DNA-Binding Proteins; Endonucleases; Exodeoxyribonucleases; Humans; Mice; Mitosis; Phenotype; RNA, Small Interfering; RecQ Helicases; Recombination, Genetic; Werner Syndrome Helicase
PubMed: 20708636
DOI: 10.1016/j.mad.2010.08.001 -
Genetics Apr 1980Semi-dominant mutants displaying greatly elevated (up to 200-fold above control) levels of spontaneous mitotic recombination have been isolated in a disomic haploid...
Semi-dominant mutants displaying greatly elevated (up to 200-fold above control) levels of spontaneous mitotic recombination have been isolated in a disomic haploid strain of yeast heteroallelic at the arg4 locus. They are designated by the symbol MIC. The mutants variously exhibit associated sensitivity to UV and ionizing radiation and to methyl methanesulfonate, enhanced UV-induced mitotic recombination, and enhanced spontaneous forward mutation rates. Possible enzyme defects and involvement in repair and editing of DNA are discussed. The mutants are expected to simplify the analysis of recombination pathways in yeast.
Topics: DNA; Gene Conversion; Methyl Methanesulfonate; Mitosis; Mutation; Radiation Tolerance; Recombination, Genetic; Saccharomyces cerevisiae
PubMed: 7002715
DOI: 10.1093/genetics/94.4.825 -
Journal of Cell Science Jun 2012BRCT-containing protein 1 (Brc1) is a multi-BRCT (BRCA1 carboxyl terminus) domain protein in Schizosaccharomyces pombe that is required for resistance to chronic...
BRCT-containing protein 1 (Brc1) is a multi-BRCT (BRCA1 carboxyl terminus) domain protein in Schizosaccharomyces pombe that is required for resistance to chronic replicative stress, but whether this reflects a repair or replication defect is unknown and the subject of this study. We show that brc1Δ cells are significantly delayed in recovery from replication pausing, though this does not activate a DNA damage checkpoint. DNA repair and recombination protein Rad52 is a homologous recombination protein that loads the Rad51 recombinase at resected double-stranded DNA (dsDNA) breaks and is also recruited to stalled replication forks, where it may stabilize structures through its strand annealing activity. Rad52 is required for the viability of brc1Δ cells, and brc1Δ cells accumulate Rad52 foci late in S phase that are potentiated by replication stress. However, these foci contain the single-stranded DNA (ssDNA) binding protein RPA, but not Rad51 or γH2A. Further, these foci are not associated with increased recombination between repeated sequences, or increased post-replication repair. Thus, these Rad52 foci do not represent sites of recombination. Following the initiation of DNA replication, the induction of these foci by replication stress is suppressed by defects in origin recognition complex (ORC) function, which is accompanied by loss of viability and severe mitotic defects. This suggests that cells lacking Brc1 undergo an ORC-dependent rescue of replication stress, presumably through the firing of dormant origins, and this generates RPA-coated ssDNA and recruits Rad52. However, as Rad51 is not recruited, and the checkpoint effector kinase Chk1 is not activated, these structures must not contain the unprotected primer ends found at sites of DNA damage that are required for recombination and checkpoint activation.
Topics: DNA Replication; DNA, Ribosomal; Hydroxyurea; Mutagenesis; Mutation; Origin Recognition Complex; Rad52 DNA Repair and Recombination Protein; Recombination, Genetic; Replication Protein A; S Phase; Schizosaccharomyces; Schizosaccharomyces pombe Proteins; Signal Transduction; Stress, Physiological
PubMed: 22366461
DOI: 10.1242/jcs.103119 -
Cancer Science Dec 2014Double-strand breaks (DSBs) are one of the severest types of DNA damage. Unrepaired DSBs easily induce cell death and chromosome aberrations. To maintain genomic... (Review)
Review
Double-strand breaks (DSBs) are one of the severest types of DNA damage. Unrepaired DSBs easily induce cell death and chromosome aberrations. To maintain genomic stability, cells have checkpoint and DSB repair systems to respond to DNA damage throughout most of the cell cycle. The failure of this process often results in apoptosis or genomic instability, such as aneuploidy, deletion, or translocation. Therefore, DSB repair is essential for maintenance of genomic stability. During mitosis, however, cells seem to suppress the DNA damage response and proceed to the next G1 phase, even if there are unrepaired DSBs. The biological significance of this suppression is not known. In this review, we summarize recent studies of mitotic DSB repair and discuss the mechanisms of suppression of DSB repair during mitosis. DSB repair, which maintains genomic integrity in other phases of the cell cycle, is rather toxic to cells during mitosis, often resulting in chromosome missegregation and aberration. Cells have multiple safeguards to prevent genomic instability during mitosis: inhibition of 53BP1 or BRCA1 localization to DSB sites, which is important to promote non-homologous end joining or homologous recombination, respectively, and also modulation of the non-homologous end joining core complex to inhibit DSB repair. We discuss how DSBs during mitosis are toxic and the multiple safeguard systems that suppress genomic instability.
Topics: Animals; Cell Cycle Proteins; Chromosome Aberrations; DNA Breaks, Double-Stranded; DNA Repair; Genomic Instability; Humans; Mitosis; Recombination, Genetic
PubMed: 25287622
DOI: 10.1111/cas.12551 -
Genetics May 2008In wild-type diploid cells, heteroallelic recombination between his4A and his4C alleles leads mostly to His+ gene conversions that have a parental configuration of... (Comparative Study)
Comparative Study
In wild-type diploid cells, heteroallelic recombination between his4A and his4C alleles leads mostly to His+ gene conversions that have a parental configuration of flanking markers, but approximately 22% of recombinants have associated reciprocal crossovers. In rad52 strains, gene conversion is reduced 75-fold and the majority of His+ recombinants are crossover associated, with the largest class being half-crossovers in which the other participating chromatid is lost. We report that UV irradiating rad52 cells results in an increase in overall recombination frequency, comparable to increases induced in wild-type (WT) cells, and surprisingly results in a pattern of recombination products quite similar to RAD52 cells: gene conversion without exchange is favored, and the number of 2n - 1 events is markedly reduced. Both spontaneous and UV-induced RAD52-independent recombination depends strongly on Rad50, whereas rad50 has no effect in cells restored to RAD52. The high level of noncrossover gene conversion outcomes in UV-induced rad52 cells depends on Rad51, but not on Rad59. Those outcomes also rely on the UV-inducible kinase Dun1 and Dun1's target, the repressor Crt1, whereas gene conversion events arising spontaneously depend on Rad59 and Crt1. Thus, there are at least two Rad52-independent recombination pathways in budding yeast.
Topics: Mitosis; Rad52 DNA Repair and Recombination Protein; Recombination, Genetic; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Ultraviolet Rays
PubMed: 18458103
DOI: 10.1534/genetics.108.087189 -
Nucleic Acids Research May 2019The post-replicative mismatch repair (MMR) system has anti-recombination activity that limits interactions between diverged sequences by recognizing mismatches in...
The post-replicative mismatch repair (MMR) system has anti-recombination activity that limits interactions between diverged sequences by recognizing mismatches in strand-exchange intermediates. In contrast to their equivalent roles during replication-error repair, mismatch recognition is more important for anti-recombination than subsequent mismatch processing. To obtain insight into this difference, ectopic substrates with 2% sequence divergence were used to examine mitotic recombination outcome (crossover or noncrossover; CO and NCO, respectively) and to infer molecular intermediates formed during double-strand break repair in Saccharomyces cerevisiae. Experiments were performed in an MMR-proficient strain, a strain with compromised mismatch-recognition activity (msh6Δ) and a strain that retained mismatch-recognition activity but was unable to process mismatches (mlh1Δ). While the loss of either mismatch binding or processing elevated the NCO frequency to a similar extent, CO events increased only when mismatch binding was compromised. The molecular features of NCOs, however, were altered in fundamentally different ways depending on whether mismatch binding or processing was eliminated. These data suggest a model in which mismatch recognition reverses strand-exchange intermediates prior to the initiation of end extension, while subsequent mismatch processing that is linked to end extension specifically destroys NCO intermediates that contain conflicting strand-discrimination signals for mismatch removal.
Topics: Base Pair Mismatch; Crossing Over, Genetic; DNA Breaks, Double-Stranded; DNA Mismatch Repair; DNA Repair; DNA Replication; DNA-Binding Proteins; Mitosis; MutL Protein Homolog 1; Nucleic Acid Heteroduplexes; Recombination, Genetic; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 30809658
DOI: 10.1093/nar/gkz126 -
Environmental Health Perspectives Dec 1996An overall hypothesis for benzene-induced leukemia is proposed. Key components of the hypothesis include a) activation of benzene in the liver to phenolic metabolites;... (Review)
Review
An overall hypothesis for benzene-induced leukemia is proposed. Key components of the hypothesis include a) activation of benzene in the liver to phenolic metabolites; b) transport of these metabolites to the bone marrow and conversion to semiquinone radicals and quinones via peroxidase enzymes; c) generation of active oxygen species via redox cycling; d) damage to tubulin, histone proteins, topoisomerase II, other DNA associated proteins, and DNA itself; and e) consequent damage including DNA strand breakage, mitotic recombination, chromosome translocations, and aneuploidy. If these effects take place in stem or early progenitor cells a leukemic clone with selective advantage to grow may arise, as a result of protooncogene activation, gene fusion, and suppressor gene inactivation. Epigenetic effects of benzene metabolites on the bone marrow stroma, and perhaps the stem cell itself, may then foster development and survival of the leukemic clone. Evidence for this hypothesis is mounting with the recent demonstration that benzene induces gene-duplicating mutations in human bone marrow and chromosome-specific aneuploidy and translocations in peripheral blood cells. If this hypothesis is correct, it also potentially implicates phenolic and quinonoid compounds in the induction of "spontaneous" leukemia in man.
Topics: Aneuploidy; Animals; Benzene; Carcinogens; Humans; Leukemia; Liver; Models, Biological; Phenols; Recombination, Genetic; Translocation, Genetic
PubMed: 9118896
DOI: 10.1289/ehp.961041219