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Exo1 plays a major role in DNA end resection in humans and influences double-strand break repair and damage signaling decisions

Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, USA.
DNA repair (Impact Factor: 3.36). 02/2012; 11(4):441-8. DOI: 10.1016/j.dnarep.2012.01.006
Source: PubMed

ABSTRACT The resection of DNA double-strand breaks (DSBs) to generate ssDNA tails is a pivotal event in the cellular response to these breaks. In the two-step model of resection, primarily elucidated in yeast, initial resection by Mre11-CtIP is followed by extensive resection by two distinct pathways involving Exo1 or BLM/WRN-Dna2. However, resection pathways and their exact contributions in humans in vivo are not as clearly worked out as in yeast. Here, we examined the contribution of Exo1 to DNA end resection in humans in vivo in response to ionizing radiation (IR) and its relationship with other resection pathways (Mre11-CtIP or BLM/WRN). We find that Exo1 plays a predominant role in resection in human cells along with an alternate pathway dependent on WRN. While Mre11 and CtIP stimulate resection in human cells, they are not absolutely required for this process and Exo1 can function in resection even in the absence of Mre11-CtIP. Interestingly, the recruitment of Exo1 to DNA breaks appears to be inhibited by the NHEJ protein Ku80, and the higher level of resection that occurs upon siRNA-mediated depletion of Ku80 is dependent on Exo1. In addition, Exo1 may be regulated by 53BP1 and Brca1, and the restoration of resection in BRCA1-deficient cells upon depletion of 53BP1 is dependent on Exo1. Finally, we find that Exo1-mediated resection facilitates a transition from ATM- to ATR-mediated cell cycle checkpoint signaling. Our results identify Exo1 as a key mediator of DNA end resection and DSB repair and damage signaling decisions in human cells.

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    • "Although Fun30 facilitates both extensive resection mechanisms, the phenotype of fun30D is similar to exo1D and overexpression of Exo1 suppresses the DNA damage sensitivity of the fun30D mutant (Chen et al. 2012; Costelloe et al. 2012). Additionally, SMARCAD1, the human ortholog of Fun30, is required for RPA localization to laser-induced DNA damage, similar to the role of EXO1 (Costelloe et al. 2012; Tomimatsu et al. 2012). Although the recruitment of RSC, INO80, and Fun30 would be expected to precede resection, localization of these factors to DSBs is reduced in exo1D sgs1D cells; furthermore, recruitment of Sgs1, Dna2, and Exo1 is reduced in the fun30D mutant, indicating a complex interdependency. "
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    ABSTRACT: RecA/Rad51 catalyzed pairing of homologous DNA strands, initiated by polymerization of the recombinase on single-stranded DNA (ssDNA), is a universal feature of homologous recombination (HR). Generation of ssDNA from a double-strand break (DSB) requires nucleolytic degradation of the 5'-terminated strands to generate 3'-ssDNA tails, a process referred to as 5'-3' end resection. The RecBCD helicase-nuclease complex is the main end-processing machine in Gram-negative bacteria. Mre11-Rad50 and Mre11-Rad50-Xrs2/Nbs1 can play a direct role in end resection in archaea and eukaryota, respectively, by removing end-blocking lesions and act indirectly by recruiting the helicases and nucleases responsible for extensive resection. In eukaryotic cells, the initiation of end resection has emerged as a critical regulatory step to differentiate between homology-dependent and end-joining repair of DSBs.
    Cold Spring Harbor perspectives in biology 08/2014; 6(8). DOI:10.1101/cshperspect.a016436 · 8.23 Impact Factor
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    • "EXO1 has been proven to be directly involved in DNA repair mechanism[18], [27], [28]. Thus, we next explored whether EXO1 accounts for FOXM1-mediated cisplatin resistance. "
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    ABSTRACT: Cisplatin is commonly used in ovarian cancer chemotherapy, however, chemoresistance to cisplatin remains a great clinical challenge. Oncogenic transcriptional factor FOXM1 has been reported to be overexpressed in ovarian cancer. In this study, we aimed to investigate the potential role of FOXM1 in ovarian cancers with chemoresistance to cisplatin. Our results indicate that FOXM1 is upregulated in chemoresistant ovarian cancer samples, and defends ovarian cancer cells against cytotoxicity of cisplatin. FOXM1 facilitates DNA repair through regulating direct transcriptional target EXO1 to protect ovarian cancer cells from cisplatin-mediated apoptosis. Attenuating FOXM1 and EXO1 expression by small interfering RNA, augments the chemotherapy efficacy against ovarian cancer. Our findings indicate that targeting FOXM1 and its target gene EXO1 could improve cisplatin effect in ovarian cancer, confirming their role in modulating cisplatin sensitivity.
    PLoS ONE 05/2014; 9(5):e96989. DOI:10.1371/journal.pone.0096989 · 3.23 Impact Factor
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    • "Among them, the association of Ku70–Ku80 to DNA ends favors end protection and promotes accurate C-NHEJ through the recruitment of the core components of the repair pathway (10,76). Several studies have shown that the absence of Ku80 not only impairs accurate NHEJ but also enhances end resection catalyzed by Exo1 and consequently HR-directed repair (53,71,77). Similarly, we find that PARP3 depleted-cells display deficient NHEJ, but enhanced DNA end resection that is marked by increased DNA-damage-induced accumulation of RPA and RAD54 foci together with accelerated RPA phosphorylation and increased activation of Mre11. "
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    ABSTRACT: The repair of toxic double-strand breaks (DSB) is critical for the maintenance of genome integrity. The major mechanisms that cope with DSB are: homologous recombination (HR) and classical or alternative nonhomologous end joining (C-NHEJ versus A-EJ). Because these pathways compete for the repair of DSB, the choice of the appropriate repair pathway is pivotal. Among the mechanisms that influence this choice, deoxyribonucleic acid (DNA) end resection plays a critical role by driving cells to HR, while accurate C-NHEJ is suppressed. Furthermore, end resection promotes error-prone A-EJ. Increasing evidence define Poly(ADP-ribose) polymerase 3 (PARP3, also known as ARTD3) as an important player in cellular response to DSB. In this work, we reveal a specific feature of PARP3 that together with Ku80 limits DNA end resection and thereby helps in making the choice between HR and NHEJ pathways. PARP3 interacts with and PARylates Ku70/Ku80. The depletion of PARP3 impairs the recruitment of YFP-Ku80 to laser-induced DNA damage sites and induces an imbalance between BRCA1 and 53BP1. Both events result in compromised accurate C-NHEJ and a concomitant increase in DNA end resection. Nevertheless, HR is significantly reduced upon PARP3 silencing while the enhanced end resection causes mutagenic deletions during A-EJ. As a result, the absence of PARP3 confers hypersensitivity to anti-tumoral drugs generating DSB.
    Nucleic Acids Research 03/2014; 42(9). DOI:10.1093/nar/gku174 · 9.11 Impact Factor
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Emma Bolderson