scholarly journals The checkpoint clamp protein Rad9 facilitates DNA-end resection and prevents alternative non-homologous end joining

Cell Cycle ◽  
2014 ◽  
Vol 13 (21) ◽  
pp. 3460-3464 ◽  
Author(s):  
Feng-Ling Tsai ◽  
Mihoko Kai
Keyword(s):  
End Joining ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
End Resection

scholarly journals Nucleosome disassembly during human non-homologous end joining followed by concerted HIRA- and CAF-1-dependent reassembly

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Xuan Li ◽  
Jessica K Tyler
Keyword(s):  
Strand Break ◽  
End Joining ◽  
Efficient Removal ◽  
Dsb Repair ◽  
Dna Double Strand Break ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
Nucleosome Disassembly ◽  
End Resection ◽  
Assembly Pathways

The cell achieves DNA double-strand break (DSB) repair in the context of chromatin structure. However, the mechanisms used to expose DSBs to the repair machinery and to restore the chromatin organization after repair remain elusive. Here we show that induction of a DSB in human cells causes local nucleosome disassembly, apparently independently from DNA end resection. This efficient removal of histone H3 from the genome during non-homologous end joining was promoted by both ATM and the ATP-dependent nucleosome remodeler INO80. Chromatin reassembly during DSB repair was dependent on the HIRA histone chaperone that is specific to the replication-independent histone variant H3.3 and on CAF-1 that is specific to the replication-dependent canonical histones H3.1/H3.2. Our data suggest that the epigenetic information is re-established after DSB repair by the concerted and interdependent action of replication-independent and replication-dependent chromatin assembly pathways.

scholarly journals DNA end-resection in highly accessible chromatin produces a toxic break

2019 ◽  
Author(s):  
Jeroen van den Berg ◽  
Stacey E.P. Joosten ◽  
YongSoo Kim ◽  
Anna G. Manjón ◽  
Lenno Krenning ◽  
...  
Keyword(s):  
Strand Break ◽  
Open Chromatin ◽  
End Joining ◽  
Close Proximity ◽  
Damage Response ◽  
Hypersensitive Sites ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
End Resection ◽  
Accessible Chromatin

AbstractOf all damage occurring to DNA, the double strand break (DSB) is the most toxic lesion. Luckily, cells have developed multiple repair pathways to cope with these lesions. These different pathways compete for the same break, and the location of the break can influence this competition. However, the exact contribution of break location in repair pathway preference is not fully understood. We observe that most breaks prefer classical non-homologous end-joining, whereas some depend on DNA end-resection for their repair. Surprisingly, we find that for a subset of these sites, the activation of resection-dependent repair induces a detrimental DNA damage response. These sites exhibit extensive DNA end-resection due to improper recruitment of 53BP1 and the Shieldin complex due to low levels of H4K20me2. Most of these sites reside in close proximity to DNAseI hypersensitive sites. Compacting or removing these regions reduces extensive DNA end-resection and restores normal repair. Taken together, we found that DSB in open chromatin is highly toxic, due to the improper activity of 53BP1 and Shieldin, resulting in extensive DNA end-resection.

scholarly journals The non-homologous end joining factor Ku orchestrates replication fork resection and fine-tunes Rad51-mediated fork restart

2017 ◽  
Author(s):  
Ana Teixeira-Silva ◽  
Anissia Ait Saada ◽  
Ismail Iraqui ◽  
Marina Charlotte Nocente ◽  
Karine Fréon ◽  
...  
Keyword(s):  
Replication Fork ◽  
Double Strand Breaks ◽  
Fine Tuning ◽  
List Type ◽  
End Joining ◽  
Strand Breaks ◽  
Replication Restart ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
End Resection

AbstractReplication requires Homologous Recombination (HR) to stabilize and restart terminally-arrested forks. HR-mediated fork processing requires single stranded DNA (ssDNA) gaps and not necessarily Double Strand Breaks. We used genetic and molecular assays to investigate fork-resection and restart at dysfunctional, unbroken forks in Schizosaccharomyces pombe. We found that fork-resection is a two-step process coordinated by the non-homologous end joining factor Ku. An initial resection mediated by MRN/Ctp1 removes Ku from terminally-arrested forks, generating ~ 110 bp sized gaps obligatory for subsequent Exo1-mediated long-range resection and replication restart. The lack of Ku results in slower fork restart, excessive resection, and impaired RPA recruitment. We propose that terminally-arrested forks undergo fork reversal, providing a single DNA end for Ku binding which primes RPA-coated ssDNA. We uncover an unprecedented role for Ku in orchestrating resection of unbroken forks and in fine-tuning HR-mediated replication restart.Ku orchestrates a two-steps DNA end-resection of terminally-arrested and unbroken forksMRN/Ctp1 removes Ku from terminally-arrested forks to initiate fork-resectiona ~110 bp sized ssDNA gap is sufficient and necessary to promote fork restart.The lack of Ku decreases ssDNA RPA-coating, and slows down replication fork restart.

scholarly journals Competing roles of DNA end resection and non-homologous end joining functions in the repair of replication-born double-strand breaks by sister-chromatid recombination

2012 ◽  
Vol 41 (3) ◽  
pp. 1669-1683 ◽  
Author(s):  
Sandra Muñoz-Galván ◽  
Ana López-Saavedra ◽  
Stephen P. Jackson ◽  
Pablo Huertas ◽  
Felipe Cortés-Ledesma ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
End Resection ◽  
Sister Chromatid Recombination

scholarly journals Antagonistic relationship of NuA4 with the Non-Homologous End-Joining machinery at DNA damage sites

2021 ◽  
Author(s):  
Salar Ahmad ◽  
Valerie Côté ◽  
Xue Cheng ◽  
Gaëlle Bourriquen ◽  
Vasileia Sapountzi ◽  
...  
Keyword(s):  
Budding Yeast ◽  
Double Strand Breaks ◽  
Repair Pathway ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
Relationship Of ◽  
End Resection

AbstractThe NuA4 histone acetyltransferase complex, apart from its known role in gene regulation, has also been directly implicated in the repair of DNA double-strand breaks (DSBs), favoring homologous recombination (HR) in S/G2 during the cell cycle. Here, we investigate the antagonistic relationship of NuA4 with non-homologous end joining (NHEJ) factors. We show that budding yeast Rad9, the 53BP1 ortholog, can inhibit NuA4 acetyltransferase activity when bound to chromatin in vitro. While we previously reported that NuA4 is recruited at DSBs during the S/G2 phase, we can also detect its recruitment in G1 when genes for NHEJ factors Rad9, Yku80 and Nej1 are mutated. This is accompanied with the binding of single-strand DNA binding protein RPA and Rad52, indicating DNA end resection in G1 as well as recruitment of the HR machinery. This NuA4 recruitment to DSBs in G1 depends on both Xrs2 and Lcd1/Ddc2. Introducing an acetyltransferase defective allele in these NHEJ mutant backgrounds decreases their hyper-resection phenotype in G1. Interestingly, we identified two novel non-histone acetylation targets of NuA4, Nej1 and Yku80. Acetyl-mimicking mutant of Nej1 inhibits repair of DNA breaks by NHEJ, decreases its interaction with other core NHEJ factors such as Yku80 and Lif1 and favors end resection. Altogether, these results establish a strong reciprocal antagonistic regulatory function of NuA4 and NHEJ factors in repair pathway choice and suggests a role of NuA4 in alternative repair mechanism that involves DNA-end resection in G1.Author SummaryDNA double-strand breaks (DSBs) are one of the most harmful form of DNA damage. Cells employ two major repair pathways to resolve DSBs: Homologous Recombination (HR) and Non-Homologous End Joining (NHEJ). Here we wanted to dissect further the role played by the NuA4 (Nucleosome acetyltransferase of histone H4) complex in the repair of DSBs. Budding yeast NuA4 complex, like its mammalian homolog TIP60 complex, has been shown to favor repair by HR. Here, we show that indeed budding yeast NuA4 and components of the NHEJ repair pathway share an antagonistic relationship. Deletion of NHEJ components enables increased recruitment of NuA4 in the vicinity of DSBs, where NuA4 favors the end resection process which is an underlying mechanism for HR repair. We also describe two independent modes responsible for the recruitment of NuA4 to DSB sites. Additionally, we also present two NHEJ core components as new targets of NuA4 acetyltransferase activity and suggest that these acetylation events can disassemble the NHEJ repair complex from DSBs, favoring repair by HR. Our study demonstrates the importance of NuA4 in the modulation of DSB repair pathway choice.

scholarly journals Antagonistic relationship of NuA4 with the non-homologous end-joining machinery at DNA damage sites

PLoS Genetics ◽  
2021 ◽  
Vol 17 (9) ◽  
pp. e1009816
Author(s):  
Salar Ahmad ◽  
Valérie Côté ◽  
Xue Cheng ◽  
Gaëlle Bourriquen ◽  
Vasileia Sapountzi ◽  
...  
Keyword(s):  
Single Strand ◽  
Regulatory Function ◽  
Repair Pathway ◽  
Dna Breaks ◽  
End Joining ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
Relationship Of ◽  
End Resection

The NuA4 histone acetyltransferase complex, apart from its known role in gene regulation, has also been directly implicated in the repair of DNA double-strand breaks (DSBs), favoring homologous recombination (HR) in S/G2 during the cell cycle. Here, we investigate the antagonistic relationship of NuA4 with non-homologous end joining (NHEJ) factors. We show that budding yeast Rad9, the 53BP1 ortholog, can inhibit NuA4 acetyltransferase activity when bound to chromatin in vitro. While we previously reported that NuA4 is recruited at DSBs during the S/G2 phase, we can also detect its recruitment in G1 when genes for Rad9 and NHEJ factors Yku80 and Nej1 are mutated. This is accompanied with the binding of single-strand DNA binding protein RPA and Rad52, indicating DNA end resection in G1 as well as recruitment of the HR machinery. This NuA4 recruitment to DSBs in G1 depends on Mre11-Rad50-Xrs2 (MRX) and Lcd1/Ddc2 and is linked to the hyper-resection phenotype of NHEJ mutants. It also implicates NuA4 in the resection-based single-strand annealing (SSA) repair pathway along Rad52. Interestingly, we identified two novel non-histone acetylation targets of NuA4, Nej1 and Yku80. Acetyl-mimicking mutant of Nej1 inhibits repair of DNA breaks by NHEJ, decreases its interaction with other core NHEJ factors such as Yku80 and Lif1 and favors end resection. Altogether, these results establish a strong reciprocal antagonistic regulatory function of NuA4 and NHEJ factors in repair pathway choice and suggests a role of NuA4 in alternative repair mechanisms in situations where some DNA-end resection can occur in G1.

scholarly journals The internal region of CtIP negatively regulates DNA end resection

2020 ◽  
Vol 48 (10) ◽  
pp. 5485-5498 ◽  
Author(s):  
Sean Michael Howard ◽  
Ilaria Ceppi ◽  
Roopesh Anand ◽  
Roger Geiger ◽  
Petr Cejka
Keyword(s):  
Homologous Recombination ◽  
Regulatory Function ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Internal Region ◽  
Dna End Resection ◽  
End Resection

Abstract DNA double-strand breaks are repaired by end-joining or homologous recombination. A key-committing step of recombination is DNA end resection. In resection, phosphorylated CtIP first promotes the endonuclease of MRE11–RAD50–NBS1 (MRN). Subsequently, CtIP also stimulates the WRN/BLM–DNA2 pathway, coordinating thus both short and long-range resection. The structure of CtIP differs from its orthologues in yeast, as it contains a large internal unstructured region. Here, we conducted a domain analysis of CtIP to define the function of the internal region in DNA end resection. We found that residues 350–600 were entirely dispensable for resection in vitro. A mutant lacking these residues was unexpectedly more efficient than full-length CtIP in DNA end resection and homologous recombination in vivo, and consequently conferred resistance to lesions induced by the topoisomerase poison camptothecin, which require high MRN–CtIP-dependent resection activity for repair. This suggested that the internal CtIP region, further mapped to residues 550–600, may mediate a negative regulatory function to prevent over resection in vivo. The CtIP internal deletion mutant exhibited sensitivity to other DNA-damaging drugs, showing that upregulated resection may be instead toxic under different conditions. These experiments together identify a region within the central CtIP domain that negatively regulates DNA end resection.

scholarly journals A role for the p53 tumour suppressor in regulating the balance between homologous recombination and non-homologous end joining

Open Biology ◽  
2016 ◽  
Vol 6 (9) ◽  
pp. 160225 ◽  
Author(s):  
Sylvie Moureau ◽  
Janna Luessing ◽  
Emma Christina Harte ◽  
Muriel Voisin ◽  
Noel Francis Lowndes
Keyword(s):  
Dna Damage ◽  
Homologous Recombination ◽  
Tumour Suppressor ◽  
Cycle Phase ◽  
Repair Pathway ◽  
End Joining ◽  
Dsb Repair ◽  
Pathway Choice ◽  
Non Homologous End Joining ◽  
End Resection

Loss of p53, a transcription factor activated by cellular stress, is a frequent event in cancer. The role of p53 in tumour suppression is largely attributed to cell fate decisions. Here, we provide evidence supporting a novel role for p53 in the regulation of DNA double-strand break (DSB) repair pathway choice. 53BP1, another tumour suppressor, was initially identified as p53 Binding Protein 1, and has been shown to inhibit DNA end resection, thereby stimulating non-homologous end joining (NHEJ). Yet another tumour suppressor, BRCA1, reciprocally promotes end resection and homologous recombination (HR). Here, we show that in both human and mouse cells, the absence of p53 results in impaired 53BP1 focal recruitment to sites of DNA damage induced by ionizing radiation. This effect is largely independent of cell cycle phase and the extent of DNA damage. In p53-deficient cells, diminished localization of 53BP1 is accompanied by a reciprocal increase in BRCA1 recruitment to DSBs. Consistent with these findings, we demonstrate that DSB repair via NHEJ is abrogated, while repair via homology-directed repair (HDR) is stimulated. Overall, we propose that in addition to its role as an ‘effector’ protein in the DNA damage response, p53 plays a role in the regulation of DSB repair pathway choice.

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