Purified human BRCA2 stimulates RAD51-mediated recombination

Jensen, Ryan B.; Carreira, Aura; Kowalczykowski, Stephen C.

doi:10.1038/nature09399

Cited by 592 publications

(740 citation statements)

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“…However, Srs2 contains a BRC repeat variant (BRCv, a.a.836–860). Srs2‐BRCv structurally resembles the BRC repeat of the human tumour suppressor BRCA2 (Islam et al , 2012) which mediates BRCA2–RAD51 interaction, thereby promoting recruitment of RAD51 to DSBs and regulation of RAD51 recombinase activity (Jensen et al , 2010; Liu et al , 2010; Thorslund et al , 2010). Srs2‐BRCv also promotes Srs2–Rad51 interaction in vitro (Islam et al , 2012).…”

Section: Discussionmentioning

confidence: 99%

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

et al. 2017

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Cells use homology‐dependent DNA repair to mend chromosome breaks and restore broken replication forks, thereby ensuring genome stability and cell survival. DNA break repair via homology‐based mechanisms involves nuclease‐dependent DNA end resection, which generates long tracts of single‐stranded DNA required for checkpoint activation and loading of homologous recombination proteins Rad52/51/55/57. While recruitment of the homologous recombination machinery is well characterized, it is not known how its presence at repair loci is coordinated with downstream re‐synthesis of resected DNA. We show that Rad51 inhibits recruitment of proliferating cell nuclear antigen (PCNA), the platform for assembly of the DNA replication machinery, and that unloading of Rad51 by Srs2 helicase is required for efficient PCNA loading and restoration of resected DNA. As a result, srs2Δ mutants are deficient in DNA repair correlating with extensive DNA processing, but this defect in srs2Δ mutants can be suppressed by inactivation of the resection nuclease Exo1. We propose a model in which during re‐synthesis of resected DNA, the replication machinery must catch up with the preceding processing nucleases, in order to close the single‐stranded gap and terminate further resection.

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Section: Discussionmentioning

confidence: 99%

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

et al. 2017

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“…A well-defined regulatory function of RPA in HR is manifest at the level of RAD51 filament formation: RPA binding to ssDNA prevents assembly of RAD51 nucleoprotein filaments, but this inhibition is alleviated by BRCA2 (Kowalczykowski 2015). In this regard, BRCA2 is serving as a mediator for RAD51 filament assembly (Jensen et al 2010). Here we reveal another aspect of HR regulation by RPA, but, in this case, regulation is at the level DNA resection.…”

Section: Discussionmentioning

confidence: 99%

PHF11 promotes DSB resection, ATR signaling, and HR

et al. 2017

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Resection of double-strand breaks (DSBs) plays a critical role in their detection and appropriate repair. The 3 ′ ssDNA protrusion formed through resection activates the ATR-dependent DNA damage response (DDR) and is required for DSB repair by homologous recombination (HR). Here we report that PHF11 (plant homeodomain finger 11) encodes a previously unknown DDR factor involved in 5 ′ end resection, ATR signaling, and HR. PHF11 was identified based on its association with deprotected telomeres and localized to sites of DNA damage in S phase. Depletion of PHF11 diminished the ATR signaling response to telomere dysfunction and genome-wide DNA damage, reduced end resection at sites of DNA damage, resulted in compromised HR and misrejoining of S-phase DSBs, and increased the sensitivity to DNA-damaging agents. PHF11 interacted with the ssDNA-binding protein RPA and was found in a complex with several nucleases, including the 5 ′ dsDNA exonuclease EXO1. Biochemical experiments demonstrated that PHF11 stimulates EXO1 by overcoming its inhibition by RPA, suggesting that PHF11 acts (in part) by promoting 5 ′ end resection at RPA-bound sites of DNA damage. These findings reveal a role for PHF11 in DSB resection, DNA damage signaling, and DSB repair.[Keywords: PHF11; EXO1; RPA; ATR; DSB; resection; homologous recombination] Supplemental material is available for this article.Received October 9, 2016; revised version accepted December 22, 2016.Nuclear double-strand breaks (DSBs) activate the ATM and ATR kinase-dependent DNA damage response (DDR) pathways (for review, see Ciccia and Elledge 2010). Whereas the ATM kinase responds to the presence of dsDNA ends, activation of the ATR kinase requires the presence of ssDNA that is bound by RPA. In addition, activation of the ATR kinase requires the loading of the 9-1-1 complex on the double-stranded-single-stranded junction formed after 5 ′ end resection. Resection of the 5 ′ end of DSBs is therefore a critical step in the activation of ATR signaling. DSB resection is also required for the initiation of several DNA repair pathways, including homologous recombination (HR) and single-strand annealing (SSA) (for review, see Symington and Gautier 2011). HR can restore the original DNA sequence using the sister chromatid as a template for repair. After 5 ′ end resection, HR is initiated by the BRCA2-mediated loading of the Rad51 recombinase onto the ssDNA. In the absence of HR, S-phase DSBs can become a substrate for more error-prone DSB repair, including SSA and classical or alternative nonhomologous end-joining (NHEJ). DSB resection is initiated through the agency of BRCA1, the MRE11/RAD50/NBS1 (MRN) complex, and CtIP (for review, see Cejka 2015). The outcome is a ssDNA end with a short (∼20-nucleotide [nt]) 3 ′ overhang that is a substrate for further nucleolytic attack by EXO1, a dsDNA exonuclease that degrades only the 5 ′ strand to leave a 3 ′ overhang that can be thousands of nucleotides in length. In addition, an overlapping pathway involving the DNA2 nuclease acting in conjun...

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“…The BRCA2 protein has been shown to interact with and stimulate RAD51 loading onto RPA-coated ssDNA [29][30][31][32] . Therefore, we tested the proposal that RAD51 ubiquitylation might interfere with its ability to interact with BRCA2.…”

Section: Rad51mentioning

confidence: 99%

FBH1 influences DNA replication fork stability and homologous recombination through ubiquitylation of RAD51

et al. 2015

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Unscheduled homologous recombination (HR) can lead to genomic instability, which greatly increases the threat of neoplastic transformation in humans. The F-box DNA helicase 1 (FBH1) is a 3 0 -5 0 DNA helicase with a putative function as a negative regulator of HR. It is the only known DNA helicase to contain an F-box, suggesting that one of its functions is to act as a ubiquitin ligase as part of an SCF (SKP1, CUL1 and F-box) complex. Here we report that the central player in HR, RAD51, is ubiquitylated by the SCF FBH1 complex. Expression of an ubiquitylation-resistant form of RAD51 in human cells leads to hyperrecombination, as well as several phenotypes indicative of an altered response to DNA replication stress. These effects are likely to be mediated by the enhanced nuclear matrix association of the ubiquitylationresistant RAD51. These data are consistent with FBH1 acting as a negative regulator of RAD51 function in human cells.

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Purified human BRCA2 stimulates RAD51-mediated recombination

Cited by 592 publications

References 50 publications

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

Unloading of homologous recombination factors is required for restoring double‐stranded DNA at damage repair loci

PHF11 promotes DSB resection, ATR signaling, and HR

FBH1 influences DNA replication fork stability and homologous recombination through ubiquitylation of RAD51

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