DNA end resection during homologous recombination

AbstractDouble stranded DNA Breaks (DSB) that occur in highly transcribed regions of the genome are preferentially repaired by homologous recombination repair (HR). However, the mechanisms that link transcription with HR are unknown. Here we identify a critical role for DHX9, a RNA helicase involved in the processing of pre-mRNA during transcription, in the initiation of HR. Cells that are deficient in DHX9 are impaired in the recruitment of RPA and RAD51 to sites of DNA damage and fail to repair DSB by HR. Consequently, these cells are hypersensitive to treatment with agents such as camptothecin and Olaparib that block transcription and generate DSB that specifically require HR for their repair. We show that DHX9 plays a critical role in HR by promoting the recruitment of BRCA1 to RNA as part of the RNA Polymerase II transcription complex, where it facilitates the resection of DSB. Moreover, defects in DHX9 also lead to impaired ATR-mediated damage signalling and an inability to restart DNA replication at camptothecin-induced DSB. Together, our data reveal a previously unknown role for DHX9 in the DNA Damage Response that provides a critical link between RNA, RNA Pol II and the repair of DNA damage by homologous recombination.

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SAMHD1 Promotes DNA End Resection to Facilitate DNA Repair by Homologous Recombination

Cell Reports ◽

10.1016/j.celrep.2017.08.008 ◽

2017 ◽

Vol 20 (8) ◽

pp. 1921-1935 ◽

Cited By ~ 65

Author(s):

Waaqo Daddacha ◽

Allyson E. Koyen ◽

Amanda J. Bastien ◽

PamelaSara E. Head ◽

Vishal R. Dhere ◽

...

Keyword(s):

Dna Repair ◽

Homologous Recombination ◽

Dna End Resection ◽

End Resection

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Induction of homologous recombination between sequence repeats by the activation induced cytidine deaminase (AID) protein

eLife ◽

10.7554/elife.03110 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 2

Author(s):

Jean-Marie Buerstedde ◽

Noel Lowndes ◽

David G Schatz

Keyword(s):

Homologous Recombination ◽

Somatic Hypermutation ◽

Cytidine Deaminase ◽

Holliday Junctions ◽

Mutagenic Potential ◽

Activation Induced Cytidine Deaminase ◽

Invasion Analysis ◽

Dna End Resection ◽

Cytidine Deamination ◽

End Resection

The activation induced cytidine deaminase (AID) protein is known to initiate somatic hypermutation, gene conversion or switch recombination by cytidine deamination within the immunoglobulin loci. Using chromosomally integrated fluorescence reporter transgenes, we demonstrate a new recombinogenic activity of AID leading to intra- and intergenic deletions via homologous recombination of sequence repeats. Repeat recombination occurs at high frequencies even when the homologous sequences are hundreds of bases away from the positions of AID-mediated cytidine deamination, suggesting DNA end resection before strand invasion. Analysis of recombinants between homeologous repeats yielded evidence for heteroduplex formation and preferential migration of the Holliday junctions to the boundaries of sequence homology. These findings broaden the target and off-target mutagenic potential of AID and establish a novel system to study induced homologous recombination in vertebrate cells.

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Human SIRT6 Promotes DNA End Resection Through CtIP Deacetylation

Science ◽

10.1126/science.1192049 ◽

2010 ◽

Vol 329 (5997) ◽

pp. 1348-1353 ◽

Cited By ~ 261

Author(s):

Abderrahmane Kaidi ◽

Brian T. Weinert ◽

Chunaram Choudhary ◽

Stephen P. Jackson

Keyword(s):

Homologous Recombination ◽

Genome Stability ◽

Protein A ◽

Strand Break ◽

Dsb Repair ◽

Interacting Protein ◽

Dna Double Strand Break ◽

Dna End Resection ◽

Lysine Deacetylases ◽

End Resection

SIRT6 belongs to the sirtuin family of protein lysine deacetylases, which regulate aging and genome stability. We found that human SIRT6 has a role in promoting DNA end resection, a crucial step in DNA double-strand break (DSB) repair by homologous recombination. SIRT6 depletion impaired the accumulation of replication protein A and single-stranded DNA at DNA damage sites, reduced rates of homologous recombination, and sensitized cells to DSB-inducing agents. We identified the DSB resection protein CtIP [C-terminal binding protein (CtBP) interacting protein] as a SIRT6 interaction partner and showed that SIRT6-dependent CtIP deacetylation promotes resection. A nonacetylatable CtIP mutant alleviated the effect of SIRT6 depletion on resection, thus identifying CtIP as a key substrate by which SIRT6 facilitates DSB processing and homologous recombination. These findings further clarify how SIRT6 promotes genome stability.

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CTCF cooperates with CtIP to drive homologous recombination repair of double-strand breaks

Nucleic Acids Research ◽

10.1093/nar/gkz639 ◽

2019 ◽

Vol 47 (17) ◽

pp. 9160-9179 ◽

Cited By ~ 6

Author(s):

Soon Young Hwang ◽

Mi Ae Kang ◽

Chul Joon Baik ◽

Yejin Lee ◽

Ngo Thanh Hang ◽

...

Keyword(s):

Homologous Recombination ◽

Critical Role ◽

Control Point ◽

Double Strand Breaks ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

C Terminus ◽

Dna End Resection ◽

End Resection

Abstract The pleiotropic CCCTC-binding factor (CTCF) plays a role in homologous recombination (HR) repair of DNA double-strand breaks (DSBs). However, the precise mechanistic role of CTCF in HR remains largely unclear. Here, we show that CTCF engages in DNA end resection, which is the initial, crucial step in HR, through its interactions with MRE11 and CtIP. Depletion of CTCF profoundly impairs HR and attenuates CtIP recruitment at DSBs. CTCF physically interacts with MRE11 and CtIP and promotes CtIP recruitment to sites of DNA damage. Subsequently, CTCF facilitates DNA end resection to allow HR, in conjunction with MRE11–CtIP. Notably, the zinc finger domain of CTCF binds to both MRE11 and CtIP and enables proficient CtIP recruitment, DNA end resection and HR. The N-terminus of CTCF is able to bind to only MRE11 and its C-terminus is incapable of binding to MRE11 and CtIP, thereby resulting in compromised CtIP recruitment, DSB resection and HR. Overall, this suggests an important function of CTCF in DNA end resection through the recruitment of CtIP at DSBs. Collectively, our findings identify a critical role of CTCF at the first control point in selecting the HR repair pathway.

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The internal region of CtIP negatively regulates DNA end resection

Nucleic Acids Research ◽

10.1093/nar/gkaa273 ◽

2020 ◽

Vol 48 (10) ◽

pp. 5485-5498 ◽

Cited By ~ 2

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.

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Multiple roles of the splicing complex SF3B in DNA end resection and homologous recombination

DNA Repair ◽

10.1016/j.dnarep.2018.04.003 ◽

2018 ◽

Vol 66-67 ◽

pp. 11-23 ◽

Cited By ~ 12

Author(s):

Rosario Prados-Carvajal ◽

Ana López-Saavedra ◽

Cristina Cepeda-García ◽

Sonia Jimeno ◽

Pablo Huertas

Keyword(s):

Homologous Recombination ◽

Multiple Roles ◽

Dna End Resection ◽

End Resection

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Persistently bound Ku at DNA ends attenuates DNA end resection and homologous recombination

DNA Repair ◽

10.1016/j.dnarep.2011.12.007 ◽

2012 ◽

Vol 11 (3) ◽

pp. 310-316 ◽

Cited By ~ 52

Author(s):

Zhengping Shao ◽

Anthony J. Davis ◽

Kazi R. Fattah ◽

Sairei So ◽

Jingxin Sun ◽

...

Keyword(s):

Homologous Recombination ◽

Dna End Resection ◽

End Resection ◽

Dna Ends

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Reversion and non-reversion mechanisms of resistance to PARP inhibitor or platinum chemotherapy in BRCA1/2-mutant metastatic breast cancer

10.1101/832717 ◽

2019 ◽

Author(s):

Adrienne G. Waks ◽

Ofir Cohen ◽

Bose Kochupurakkal ◽

Dewey Kim ◽

Connor E. Dunn ◽

...

Keyword(s):

Breast Cancer ◽

Metastatic Breast Cancer ◽

Homologous Recombination ◽

Parp Inhibitor ◽

Metastatic Breast ◽

Circulating Tumor Dna ◽

Dna End Resection ◽

Platinum Chemotherapy ◽

End Resection ◽

Tumor Dna

AbstractBackgroundLittle is known about mechanisms of resistance to PARP inhibitors and platinum chemotherapy in patients with metastatic breast cancer and BRCA1/2 mutations. Further investigation of resistance in clinical cohorts may point to strategies to prevent or overcome treatment failure.Patients and MethodsWe obtained tumor biopsies from metastatic breast cancer patients with BRCA1/2 deficiency before and after acquired resistance to PARP inhibitor or platinum chemotherapy. Whole exome sequencing was performed on each tumor, germline DNA, and circulating tumor DNA. Tumors underwent RNA sequencing, and immunohistochemical staining for RAD51 foci on tumor sections was performed for functional assessment of intact homologous recombination.ResultsPre- and post-resistance tumor samples were sequenced from 8 patients (4 with BRCA1 and 4 with BRCA2 mutation; 4 treated with PARP inhibitor and 4 with platinum). Following disease progression on DNA-damaging therapy, four patients (50%) acquired at least one somatic reversion alteration likely to result in functional BRCA1/2 protein detected by tumor or circulating tumor DNA sequencing. Two patients with germline BRCA1 deficiency acquired genomic alterations anticipated to restore homologous recombination through increased DNA end resection: loss of TP53BP1 in one patient and amplification of MRE11A in another. RAD51 foci were acquired post-resistance in all patients with genomic reversion, consistent with reconstitution of homologous recombination. All patients whose tumors demonstrated RAD51 foci post-resistance were intrinsically resistant to subsequent lines of DNA-damaging therapy.ConclusionsGenomic reversion in BRCA1/2 was the most commonly observed mechanism of resistance, occurring in 4 of 8 patients. Novel sequence alterations leading to increased DNA end resection were seen in two patients, and may be targetable for therapeutic benefit. The presence of RAD51 foci by immunohistochemistry was consistent with BRCA1/2 protein functional status from genomic data and predicted response to later DNA-damaging therapy, supporting RAD51 focus formation as a clinically useful biomarker.

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NuA4 and SAGA acetyltransferase complexes cooperate for repair of DNA breaks by homologous recombination

PLoS Genetics ◽

10.1371/journal.pgen.1009459 ◽

2021 ◽

Vol 17 (7) ◽

pp. e1009459

Author(s):

Xue Cheng ◽

Valérie Côté ◽

Jacques Côté

Keyword(s):

Homologous Recombination ◽

Strand Break ◽

Saga Complex ◽

Loop Formation ◽

Dna Breaks ◽

Direct Role ◽

Chromatin Remodelers ◽

D Loop ◽

Dna End Resection ◽

End Resection

Chromatin modifying complexes play important yet not fully defined roles in DNA repair processes. The essential NuA4 histone acetyltransferase (HAT) complex is recruited to double-strand break (DSB) sites and spreads along with DNA end resection. As predicted, NuA4 acetylates surrounding nucleosomes upon DSB induction and defects in its activity correlate with altered DNA end resection and Rad51 recombinase recruitment. Importantly, we show that NuA4 is also recruited to the donor sequence during recombination along with increased H4 acetylation, indicating a direct role during strand invasion/D-loop formation after resection. We found that NuA4 cooperates locally with another HAT, the SAGA complex, during DSB repair as their combined action is essential for DNA end resection to occur. This cooperation of NuA4 and SAGA is required for recruitment of ATP-dependent chromatin remodelers, targeted acetylation of repair factors and homologous recombination. Our work reveals a multifaceted and conserved cooperation mechanism between acetyltransferase complexes to allow repair of DNA breaks by homologous recombination.

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