Which Double-strand Repair Mechanism Is Most Active During The G1 Phase?
Cell Cycle. Author manuscript; bachelor in PMC 2009 Sep 29.
Published in final edited form equally:
PMCID: PMC2754209
NIHMSID: NIHMS111601
DNA repair by nonhomologous end joining and homologous recombination during cell bicycle in man cells
Abstract
DNA double-strand breaks (DSBs) are dangerous lesions that tin can lead to potentially oncogenic genomic rearrangements or cell decease. The two major pathways for repair of DSBs are nonhomologous end joining (NHEJ) and homologous recombination (Hr). NHEJ is an intrinsically fault-prone pathway while Hr results in authentic repair. To understand the origin of genomic instability in human being cells information technology is important to know the contribution of each DSB repair pathway. Studies of rodent cells and man cancer jail cell lines have shown that the choice between NHEJ or HR pathways depends on jail cell cycle stage. Surprisingly, cell cycle regulation of DSB repair has not been examined in normal human being cells with intact jail cell bicycle checkpoints. Hither we measured the efficiency of NHEJ and HR at different prison cell cycle stages in hTERT-immortalized diploid human fibroblasts. We utilized cells with chromosomally-integrated fluorescent reporter cassettes, in which a unique DSB is introduced by a rare-cut endonuclease. Nosotros show that NHEJ is active throughout the jail cell bike, and its activity increases as cells progress from G1 to G2/M (G1<S<G2/M). Hour is nigh absent-minded in G1, most agile in the South phase, and declines in G2/M. Thus, in G2/1000 NHEJ is elevated, while Hour is on decline. This is in dissimilarity to a full general belief that NHEJ is nearly agile in G1, while 60 minutes is active in Southward, G2 and M. The overall efficiency of NHEJ was higher than Hr at all prison cell wheel stages. We conclude that human somatic cells utilize error-prone NHEJ as the major DSB repair pathway at all jail cell cycle stages, while Hr is used, primarily, in the Due south stage.
Keywords: Cell cycle, Dna repair, normal human being fibroblasts, nonhomologous terminate joining, homologous recombination
INTRODUCTION
Deoxyribonucleic acid double-strand breaks (DSBs), in which both strands in the double helix are severed, are the virtually dangerous type of DNA lesion. If left unrepaired, or repaired incorrectly, DSBs may outcome in massive loss of genetic data, genomic rearrangements, or cell expiry. DSBs are repaired by 2 major mechanisms: not-homologous stop joining (NHEJ) and homologous recombination (HR) i. The two pathways differ in their fidelity and template requirements. NHEJ modifies the broken Dna ends, and ligates them together with fiddling or no homology, generating deletions or insertions two. In dissimilarity, HR uses an undamaged DNA template on the sis chromatid or homologous chromosome to repair the suspension, leading to the reconstitution of the original sequence three. Thus, the option of DSB repair pathway determines the fidelity of repair, which in turn may influence the rates of aging and tumorigenesis iv – vi.
Both DSB repair pathways play important roles in mammalian DSB repair 7 , eight. The verbal mechanism by which the choice between the two DSB repair pathways is made remains unclear. In role, the choice of repair pathway is determined by cell bicycle stage. The dependence of DSB repair on the cell bike was first shown past analyzing the sensitivity of craven DT40 cells, scarce in NHEJ or HR factors, to ionizing radiation ix. NHEJ mutants were highly sensitive in G1 and early on S, while HR mutants were sensitive but in S/G2. Studies in hamster CHO prison cell lines containing mutations in DSB repair genes showed that NHEJ-defective cells take reduced repair at all cell cycle stages, while HR-defective cells have a small defect in G1, and greater impairment in S/G2/M 10 , 11. Thus, NHEJ is a major DSB repair pathway in G1 stage, while both NHEJ and 60 minutes may be agile in S/G2/G. The studies of mutant cell lines practise not let for comparison of the contributions of each pathway, because diverse mutations inactivate NHEJ and HR to a different extent. Furthermore, the effect of jail cell cycle phase on DSB repair has not been analyzed in primary human being cells, which maintain intact prison cell bicycle checkpoints.
We recently adult sensitive fluorescent reporter assays to examine NHEJ and Hour in hTERT-immortalized diploid human cells. These cells retain all characteristics of untransformed primary cells 12, including intact cell cycle checkpoints. Nosotros show that NHEJ is active throughout the cell wheel, but is the highest in G2/M. HR is nearly absent in G1, most active in the South phase, and is low in G2/M. Thus, in normal human fibroblasts NHEJ is the major DSB repair pathway, while 60 minutes primarily repairs Dna breaks that occur during replication.
RESULTS
The efficiency of NHEJ and HR during jail cell bicycle has not been examined in normal human being cells. The utilise of normal cells is of import because normal cells maintain an intact jail cell cycle command appliance, which may exist involved in regulating Deoxyribonucleic acid repair. To examine the contribution of NHEJ and HR in DSB repair during the prison cell bike we used hTERT-immortalized normal human fibroblasts (HCA2-hTERT) containing chromosomally integrated GFP-based NHEJ and Hour reporter constructs. The construction of NHEJ and Hour reporter prison cell lines has been described previously 13 , fourteen. Hither we used two NHEJ (I9a and S13a) and two Hr (H15c and H32c) prison cell lines for assay of repair during the cell bicycle.
The reporter cassette for detecting NHEJ 15 contains a GFP gene with an artificially engineered iii kb intron from the Pem1 gene (GFP-Pem1). The Pem1 intron contains an adenoviral exon flanked by 18 bp recognition sequences for I-SceI endonuclease (Figure 1A). I-SceI is used to generate site-specific DSBs in vivo. In the NHEJ-I9a clone (Effigy 1A) I-SceI sites are in inverted orientation, which generate incompatible ends (Figure 1C), and in NHEJ-S13a (Figure 1A) I-SceI sites are in a direct orientation, which generate compatible DNA ends (Figure 1D). An un-rearranged NHEJ cassette is GFP negative since the adenoviral exon disrupts the GFP ORF. Upon induction of DSBs by the expression of I-SceI 16, the adenoviral exon is removed and NHEJ restores role of the GFP gene. This reporter can detect a wide spectrum of NHEJ events since the intron tin tolerate deletions and insertions. The HR reporter (Figure 1B) is congenital using the same GFP-Pem1 gene every bit the NHEJ reporter 14. In the Hr reporter, the first exon of GFP-Pem1 contains a 22 bp deletion combined with the insertion of three restriction sites, I-SceI-HindIII-I-SceI, which are used for inducing DSBs. The deletion ensures that GFP cannot be reconstituted past an NHEJ outcome. The 2 I-SceI sites are in an inverted orientation, and then that I-SceI digestion leaves incompatible ends (Figure 1C). The start re-create of GFP-Pem1 is followed by a promoter-less/ATG-less first exon and intron of GFP-Pem1. The intact construct is GFP-negative. Upon induction of a DSB by I-SceI digestion the functional GFP factor will be reconstituted by intramolecular or intermolecular gene conversion between the two mutated copies of the showtime GFP-Pem1 exon. Since the second copy of the GFP factor lacks a promoter, the first ATG codon, and the second exon, crossing over or single-strand annealing volition not restore the GFP activity. This design allows for the exclusive detection of gene conversion, which is the predominant Hour pathway in mammalian cells 17. The H15c and H32c prison cell lines used in this study are 2 independent integrants of the HR reporter. Reconstitution of the functional GFP gene past both NHEJ or Hour has been confirmed past sequencing reporter cassettes from GFP-positive clones 13.
Reporter constructs for analysis of NHEJ and HR repair. (A) Reporter cassette for detection of NHEJ. The cassette consists of a GFP factor under a CMV promoter with an engineered intron from the rat Pem1 factor, interrupted by an adenoviral exon (Advert). The adenoviral exon is flanked by I-SceI recognition sites in inverted orientation (cell line I9a) or in direct orientation (cell line S13a) for induction of DSBs. In this construct the GFP gene is inactive; nonetheless upon consecration of a DSB and successful NHEJ the construct becomes GFP+. SD, splice donor; SA, splice acceptor; shaded squares, polyadenylation sites. (B) Reporter cassette for detection of Hr (cell lines H15c and H32c). The cassette consists of ii mutated copies of GFP-Pem1. In the start copy of GFP-Pem1 the outset GFP exon contains a deletion of 22 nt and an insertion of two I-SceI recognition sites in inverted orientation. The 22 nt deletion ensures that GFP cannot exist reconstituted by a NHEJ issue. The 2nd copy of GFP-Pem1 lacks a promoter, the first ATG, and the second exon of GFP. Upon induction of DSBs by I-SceI, gene conversion events reconstitute an active GFP cistron. (C) Incompatible DNA ends generated by digestion of two I-SceI sites in inverted orientation as in the lines NHEJ-I9a, Hour-H15c, and 60 minutes-H32c. (D) Compatible DNA ends generated by digestion of two I-SceI sites in direct orientation as in the line NHEJ-S13a.
To written report DSB repair during jail cell wheel we outset determined the treatments that arrest HCA2-hTERT cells at various prison cell cycle stages. Normal cells are sensitive to contact inhibition, and arrest in G1 stage at confluence (Figure 2A). Treatment with a DNA polymerse α inhibitor, aphidicolin 18, arrested HCA2-hTERT in S stage (Figure 2A). Colchicine, which prevents microtubule polymerization 19, arrested the cells in G2/Chiliad stage (Figure 2A). Jail cell bike distribution in the treated cells was verified by propidium iodide staining and flow cytometry every 24-hour interval for 7 days following treatment. Drug-treated cells entered prison cell cycle arrest 2 days subsequently handling, and remained arrested for at least seven days. No cell decease was observed in either confluent or drug-treated cells. Confluent cells were in G1 stage afterwards reaching confluence, and could be maintained in G1 indefinitely with regular media changes. Figure 2A shows cell bicycle distribution on 24-hour interval vii subsequently reaching confluence and day 4 after drug treatment. This is the fourth dimension point when DSB repair was taking identify (discussed below).
DSB repair pathways during cell cycle. (A) Prison cell bicycle distribution of HCA2-hTERT cells following indicated treatments. Number of cells is plotted against Deoxyribonucleic acid content determined by PI staining. Cell cycle distribution was analyzed every day for vii sequent days. The images show confluent cells on day 7 after they reached confluence, and drug treated cells on solar day 4 after handling, which is the time point when DSB repair took place. (B) Representative FACS traces for the analysis of NHEJ and HR. Cells were co-transfected with 5 µg of plasmid encoding I-SceI, and 0.1 µg of a DsRed plasmid, later on they entered growth abort as shown in panel A. Cells were analyzed by menstruum cytometry 4 days after transfection, using green-versus-red fluorescent plot every bit described previously fifteen. Green fluorescence is plotted on the 10-axis and cerise fluorescence is plotted on the y-axis. The narrow triangular area on the diagonal corresponds to autofluorescent cells. Typically xx,000 cells were analyzed in each sample. In the treatments where the numbers of GFP+ cells were low, 40,000 cells were scored. (C, D) Frequency of NHEJ and HR at various cell wheel stages analyzed in two independent NHEJ and HR reporter cell lines. Each cell line contains a single integrated re-create of an NHEJ or 60 minutes reporter cassette. The ratio of GFP+/DsRed+ cells is used every bit a measure out of repair efficiency. NHEJ and HR are shown on different scales due to the big differences in repair frequency. The experiments were repeated at least four times and error bars are s. d. Stars (*) point the statistically pregnant differences (P<0.05, t-test).
To clarify NHEJ and Hr G1 arrested cells were co-transfected with 5 µg of I-SceI-expressing plasmid 20 to induce DSBs and 0.ane µg DsRed plasmid to normalize for the transfection efficiency. G1-arrested cells were transfected on twenty-four hours vi after cells reached confluence, and drug-treated cells were transfected on day 3 subsequently drug treatment. In HCA2 fibroblasts I-SceI expression reaches a maximum during the start 24 hours after transfection, and and then progressively declines 14. Therefore, the bulk of DSBs were induced on mean solar day 7 of confluence and day four after drug treatment. Repair of I-SceI-induced breaks results in the appearance of GFP+ cells. To quantify NHEJ and HR events cells were analyzed by menstruation cytometry 4 days post-transfection to allow for maximum GFP expression in drug-treated cells. Our airplane pilot experiments showed no decline in GFP or DsRed indicate during the first 4 days after transfection. GFP and DsRed fluorescence was analyzed using the green-versus-red plot (Figure 2B) every bit described previously 15. To normalize for the efficiency of transfection, the ratio of GFP+ to DsRed+ cells was used as a measure of NHEJ and Hr efficiency.
In G1 phase NHEJ was active while HR was about completely repressed (Figure 2C, D). As the HR reporter contains a duplication of the GFP gene, HR can potentially occur intrachromosomally. The near absence of 60 minutes in G1 indicates that HR does not occur when just an intrachromosomal template is available.
In S stage the frequency of NHEJ was increased by approximately 1.v to 3 fold relative to the G1 stage cells (Figure 2C). The frequency of HR in S increased xx to 27 fold relative to G1 phase (Figure 2d). Despite the sharp increase in HR frequency in S phase it remained lower than S-phase NHEJ.
Interestingly, in G2/One thousand NHEJ frequency was further elevated 1.5 to iv fold relative to the Southward phase (Figure 2C). NHEJ of both uniform and incompatible ends was elevated in G2/M, but the rise in compatible-finish NHEJ (line S13a) was more dramatic. The frequency of 60 minutes declined in G2/M by 5 fold relative to Hr in S phase (Figure 1D). The increment of NHEJ and refuse of Hr in G2/M is an unexpected finding, since HR is believed to be efficient in G2/Thousand.
In summary, our results indicate that NHEJ is the major DSB repair pathway at all cell bicycle stages, and is most agile in G2/Yard phase. Hour operates predominantly in South-phase, but even in S phase Hr may be less efficient than NHEJ.
DISCUSSION
Here we report the commencement direct comparison of NHEJ and 60 minutes in normal man cells. We use sensitive fluorescent reporter assays that allow for a straight comparison of the efficiencies of NHEJ and HR events upon induction of chromosomal DSBs with a rare-cutting endonuclease. Fluorescent assays score DSB repair events in thousands of cells, and are highly quantitative.
Our data shows that NHEJ is active throughout the jail cell cycle. Our distinct finding is that NHEJ efficiency is G1<Due south<G2/Grand. In earlier studies the roles of NHEJ and HR during the jail cell cycle were analyzed by assaying the sensitivity of synchronized NHEJ or 60 minutes mutant cells to ionizing radiation ix – 11 , 21 – 23. Most of the NHEJ-deficient cell lines are hypersensitive in G1 phase, leading to determination that NHEJ acts primarily in G1. NHEJ has been proposed to act also in S-phase 24 , 25, or in all cell cycle stages ten , 11. It was also shown that fidelity of NHEJ is higher in G2 than G1 26. We show that the efficiency of NHEJ in G2/K is 4–6 fold higher than in G1. Thus, NHEJ plays the major role in DSB repair in homo cells and its activity is increased in South and G2/M stages despite availability of Hr. Our results likewise suggest that G2/M phase of the jail cell cycle is characterized by the highest efficiency of Dna intermission repair.
Nosotros evidence that HR is extremely low in G1, is most efficient in S, and is decreased in G2/M. The prison cell survival studies uniformly concord that Hr is not utilized in G1 phase 9 – eleven , 27. Ectopic and allelic sequences may potentially exist used equally templates for repair, only in organisms with large repetitive genomes this may generate chromosomal rearrangements and a loss of heterozygosity. Indeed, studies in both plants and animals show that homologous and ectopic sequences are used at very low frequencies 17 , 28 , 29. Theoretically, HR may occur in Due south, G2 and Thousand since sister chromatids are nowadays. Our results indicate that HR is used more frequently in S than in G2/Thou. It was shown that Hour is important for repairing lesions resulting from replication block 10 , xxx , 31. Thus, a possible explanation for decreased HR in G2/M is that a primary function of 60 minutes in mammalian cells is to repair Dna damage associated with Dna replication, and HR is active when Deoxyribonucleic acid replication mechanism is present.
Since our assay straight measures NHEJ and Hour using the aforementioned type of reporter we can evaluate the contribution of NHEJ and Hr at each cell cycle stage. The comparison between NHEJ and 60 minutes is summarized in a model (Figure iii) in which NHEJ contributes to DSB repair at all cell cycle phases, but is most agile in G2/M, while Hour contributes primarily in S phase, and modestly in G2/1000. Importantly, fifty-fifty in the Due south phase when HR is the nigh active, NHEJ is more efficient than HR. Our information is consequent with NHEJ being the major DSB repair pathway during all cell bicycle stages in human somatic cells. The reason for preferential use of NHEJ by human being cells is likely to be the repetitive nature of the human being genome 5. If an incorrect template were to be used for repair, an HR upshot could result in a gross genomic rearrangement. Organisms with highly repetitive genomes may therefore favor NHEJ, every bit a pocket-size deletion associated with an NHEJ event is less deleterious than an aberrant recombination event. This strategy will protect immature organisms from genomic rearrangements; however, over fourth dimension mutations introduced by NHEJ will accumulate in the genome and contribute to a functional decline associated with aging.
Model for the contribution of NHEJ and HR to repair of DSBs during cell cycle. NHEJ is active during all stages of cell cycle, simply its efficiency is the highest during G2/M. 60 minutes is active primarily in the Southward-phase and has lower activity in G2/M. The length of each cell wheel stage is proportional to the number of cells at a corresponding stage in unsynchronized culture of normal fibroblasts in Figure 2A.
MATERIALS AND METHODS
Jail cell lines and culture conditions
HCA2 are human neonatal foreskin fibroblasts isolated by the laboratory of O.M. Pereira-Smith (UTHSCSA, Houston, TX). Construction of NHEJ-I9a, NHEJ-S13a, Hour-H15c, and Hr-H32c reporter cell lines is described in 14. Cells were cultured at 37° in a 5% CO2, 3% O2 incubator, in EMEM media supplemented with 15% fetal bovine serum, 100 units/ml penicillin and 100 μµ/ml streptomycin.
Cell wheel arrest
In guild to arrest the cells at diverse stages of the jail cell cycle, several growth atmospheric condition, and drug treatments were used. For G1 arrest, cells were plated on 100 mm plates at a concentration of 5×x5 and allowed to grow to confluence and kept there for 6 days before transfection. Following transfection, the confluent cells were replated on threescore mm plates to maintain a land of contact inhibition and therefore G1 arrest. S-stage arrest was induced by the addition of one µg/ml aphidicolin, and G2/M arrest was induced by the improver of 0.one mg/ml colchicine.
Transfections
The transfections were performed using an Amaxa Nucleofector; program U-20. In each transfection 2×ten6 cells were transfected with 0.1 µg of a DsRed expressing plasmid, and 5 µg of an I-SceI expressing plasmid.
FACS assay
For the analysis of NHEJ and 60 minutes cells were harvested, resuspended in ~1ml 1×PBS, put on ice, and run on a FACS Calibur instrument. GFP and DsRed fluorescence was analyzed using the red-versus-light-green plot as described previously 15. Cell cycle distribution was analyzed by PI staining. Information was analyzed using CellQuest software.
Acknowledgements
We thank Dara Brown, A'Shantee O'Steen, and Anna Sokolov for help with construction of HR reporter cell lines. This work was supported by grants from US National Found of Health AG027237 (5.Thou.), American Federation for Aging Research (5.M.), the Komen Foundation (V.G.), and Ellison Medical Foundation (5.G. and A.Southward.).
Abbreviations
| DSB | double-strand break |
| HR | homologous recombination |
| NHEJ | nonhomologous terminate joining |
| hTERT | human telomerase contrary transcriptase |
| GFP | green fluorescent protein |
| DsRed | red fluorescent protein |
REFERENCES
1. Haber JE. Partners and pathways repairing a double-strand break. Trends Genet. 2000;xvi:259–264. [PubMed] [Google Scholar]
2. Lieber MR. The machinery of human nonhomologous DNA stop joining. J Biol Chem. 2008;283:1–5. [PubMed] [Google Scholar]
3. Thompson LH, Schild D. Homologous recombinational repair of Dna ensures mammalian chromosome stability. Mutat Res. 2001;477:131–153. [PubMed] [Google Scholar]
4. Karanjawala ZE, Lieber MR. Deoxyribonucleic acid damage and crumbling. Mech Ageing Dev. 2004;125:405–416. [PubMed] [Google Scholar]
5. Lieber MR, Karanjawala ZE. Ageing, repetitive genomes and DNA damage. Nat Rev Mol Cell Biol. 2004;v:69–75. [PubMed] [Google Scholar]
vi. Thompson LH, Schild D. Recombinational Deoxyribonucleic acid repair and human disease. Mutat Res. 2002;509:49–78. [PubMed] [Google Scholar]
7. Sonoda Due east, Hochegger H, Saberi A, Taniguchi Y, Takeda S. Differential usage of not-homologous finish-joining and homologous recombination in double strand break repair. Dna Repair (Amst) 2006;v:1021–1029. [PubMed] [Google Scholar]
8. Sargent RG, Branneman MA, Wilson JH. Repair of site-specific double-strand breaks in mammalian chromosome by homologous and illegitimate recombination. Mol Cell Biol. 1997;17:267–277. [PMC gratis commodity] [PubMed] [Google Scholar]
9. Takata Yard, Sasaki MS, Sonoda E, Morrison C, Hashimoto M, Utsumi H, et al. Homologous recombination and non-homologous end-joining pathways of Deoxyribonucleic acid double-strand break repair accept overlapping roles in the maintenance of chromosomal integrity in vertebrate cells. Embo J. 1998;17:5497–5508. [PMC free article] [PubMed] [Google Scholar]
10. Rothkamm K, Kruger I, Thompson LH, Lobrich M. Pathways of Dna double-strand break repair during the mammalian prison cell cycle. Mol Cell Biol. 2003;23:5706–5715. [PMC free commodity] [PubMed] [Google Scholar]
11. Hinz JM, Yamada NA, Salazar EP, Tebbs RS, Thompson LH. Influence of double-strand-intermission repair pathways on radiosensitivity throughout the cell bicycle in CHO cells. Deoxyribonucleic acid Repair (Amst) 2005;4:782–792. [PubMed] [Google Scholar]
12. Morales CP, Holt I, Ouellette Yard, Kaur KJ, Yan Y, Wilson KS, et al. Absenteeism of cancer-associated changes in man fibroblasts immortalized with telomerase. Nat Genet. 1999;21:115–118. [PubMed] [Google Scholar]
thirteen. Mao Z, Seluanov A, Jiang Y, Gorbunova 5. TRF2 is required for repair of nontelomeric Dna double-strand breaks by homologous recombination. Proc Natl Acad Sci U South A. 2007;104:13068–13073. [PMC free article] [PubMed] [Google Scholar]
14. Mao Z, Bozzella M, Seluanov A, Gorbunova V. Comparing of nonhomologous end joining and homologous recombination in man cells. DNA Repair (Amst) 2008 In Press. [PMC complimentary commodity] [PubMed] [Google Scholar]
15. Seluanov A, Mittelman D, Pereira-Smith OM, Wilson JH, Gorbunova V. DNA end joining becomes less efficient and more than error-decumbent during cellular senescence. Proc Natl Acad Sci U S A. 2004;101:7624–7629. [PMC complimentary commodity] [PubMed] [Google Scholar]
16. Jasin Yard. Genetic manipulation of genomes with rare-cut endonucleases. Trends Genet. 1996;12:224–228. [PubMed] [Google Scholar]
17. Johnson RD, Jasin M. Sister chromatid gene conversion is a prominent double strand suspension repair in mammalian cells. EMBO J. 2000;19:3398–3407. [PMC gratuitous article] [PubMed] [Google Scholar]
eighteen. Ikegami Southward, Taguchi T, Ohashi Yard, Oguro Grand, Nagano H, Mano Y. Aphidicolin prevents mitotic jail cell division by interfering with the activity of Deoxyribonucleic acid polymerase-blastoff. Nature. 1978;275:458–460. [PubMed] [Google Scholar]
19. Luduena RF, Banerjee A, Khan IA. Tubulin structure and biochemistry. Curr Opin Cell Biol. 1992;four:53–57. [PubMed] [Google Scholar]
20. Rouet P, Smih F, Jasin K. Introduction of double-strand breaks into the genome of mouse cells by expression of a rare-cut endonuclease. Mol Cell Biol. 1994;14:8096–8106. [PMC gratis article] [PubMed] [Google Scholar]
21. Ferreira MG, Cooper JP. Two modes of Dna double-strand break repair are reciprocally regulated through the fission yeast cell cycle. Genes Dev. 2004;18:2249–2254. [PMC complimentary article] [PubMed] [Google Scholar]
22. Stamato TD, Dipatri A, Giaccia A. Jail cell-cycle-dependent repair of potentially lethal damage in the XR-i gamma-ray-sensitive Chinese hamster ovary cell. Radiat Res. 1988;115:325–333. [PubMed] [Google Scholar]
23. Jeggo PA. Studies on mammalian mutants defective in rejoining double-strand breaks in DNA. Mutat Res. 1990;239:1–16. [PubMed] [Google Scholar]
24. Mills KD, Ferguson DO, Essers J, Eckersdorff M, Kanaar R, Alt FW. Rad54 and DNA Ligase 4 cooperate to maintain mammalian chromatid stability. Genes Dev. 2004;18:1283–1292. [PMC free article] [PubMed] [Google Scholar]
25. Couedel C, Mills KD, Barchi Thou, Shen L, Olshen A, Johnson RD, et al. Collaboration of homologous recombination and nonhomologous end-joining factors for the survival and integrity of mice and cells. Genes Dev. 2004;18:1293–1304. [PMC free commodity] [PubMed] [Google Scholar]
26. Kruger I, Rothkamm K, Lobrich 1000. Enhanced fidelity for rejoining radiation-induced DNA double-strand breaks in the G2 phase of Chinese hamster ovary cells. Nucleic Acids Res. 2004;32:2677–2684. [PMC complimentary article] [PubMed] [Google Scholar]
27. Cheong Northward, Wang Ten, Wang Y, Iliakis Thou. Loss of S-phase-dependent radioresistance in irs-1 cells exposed to X-rays. Mutat Res. 1994;314:77–85. [PubMed] [Google Scholar]
28. Puchta H. The repair of double-strand breaks in plants: mechanisms and consequences for genome evolution. J Exp Bot. 2005;56:one–14. [PubMed] [Google Scholar]
29. Shalev K, Levy AA. The maize transposable element Ac induces recombination between the donor site and an homologous ectopic sequence. Genetics. 1997;146:1143–1151. [PMC complimentary commodity] [PubMed] [Google Scholar]
xxx. Saintigny Y, Delacote F, Vares 1000, Petitot F, Lambert Due south, Averbeck D, et al. Characterization of homologous recombination induced by replication inhibition in mammalian cells. Embo J. 2001;20:3861–3870. [PMC free article] [PubMed] [Google Scholar]
31. Arnaudeau C, Lundin C, Helleday T. Dna double-strand breaks associated with replication forks are predominantly repaired past homologous recombination involving an exchange mechanism in mammalian cells. J Mol Biol. 2001;307:1235–1245. [PubMed] [Google Scholar]
Which Double-strand Repair Mechanism Is Most Active During The G1 Phase?,
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