Article

Novel pro- and anti-recombination activities of the Bloom's syndrome helicase

Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, USA.
Genes & Development (Impact Factor: 10.8). 01/2008; 21(23):3085-94. DOI: 10.1101/gad.1609007
Source: PubMed

ABSTRACT

Bloom's syndrome (BS) is an autosomal recessive disorder characterized by a strong cancer predisposition. The defining feature of BS is extreme genome instability. The gene mutated in Bloom's syndrome, BLM, encodes a DNA helicase (BLM) of the RecQ family. BLM plays a role in homologous recombination; however, its exact function remains controversial. Mutations in the BLM cause hyperrecombination between sister chromatids and homologous chromosomes, indicating an anti-recombination role. Conversely, other data show that BLM is required for recombination. It was previously shown that in vitro BLM helicase promotes disruption of recombination intermediates, regression of stalled replication forks, and dissolution of double Holliday junctions. Here, we demonstrate two novel activities of BLM: disruption of the Rad51-ssDNA (single-stranded DNA) filament, an active species that promotes homologous recombination, and stimulation of DNA repair synthesis. Using in vitro reconstitution reactions, we analyzed how different biochemical activities of BLM contribute to its functions in homologous recombination.

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    • "The differences in metabolism of RAD51 filaments may also be reflected in the absence of a clear ortholog of Srs2 (i.e., of a factor dismantling RAD51 filaments from ssDNA) (Krejci et al. 2003; Veaute et al. 2003) in higher eukaryotes. Despite the fact that in vitro several human RecQ orthologs (e.g., BLM, RECQL5) have been shown to dismantle RAD51 filaments (Hu et al. 2007; Bugreev et al. 2007), in vivo evidence suggests it is more plausible that PARI, FHB1, and RTEL1 are the bona fide Srs2 functional orthologs (Chiolo et al. 2007; Barber et al. 2008; Moldovan et al. 2012; Vannier et al. 2013). It is noteworthy that, mechanistically, RTEL1 resembles yeast Mph1 but not Srs2, as it dismantles D-loop intermediate and not RAD51 filaments (Prakash et al. 2009; Barber et al. 2008; Vannier et al. 2012). "
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    ABSTRACT: Homologous recombination (HR) maintains genome stability by repairing DNA double-strand breaks and gaps and restarting replication forks. It is an error-free pathway that uses a homologous sequence in the genome to copy the damaged genetic information. In the present chapter, we will discuss in detail the mechanism by which HR operates to maintain genome stability as revealed by studies predominantly performed in Saccharomyces cerevisiae. We will then discuss the similarities and dissimilarities between yeast and humans while emphasizing the importance of HR in suppressing carcinogenesis and as a potential therapeutic target.
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    • "It is unclear why depletion of two independent factors (namely BLM or FBH1) is able to compensate for a lack of BOD1L, although BLM and FBH1 have partially redundant functions in DT40 cells (Kohzaki et al., 2007). While FBH1 and BLM both have pro-and anti-recombinogenic activities (Bugreev et al., 2007; Fugger et al., 2009), BLM can displace RAD51 from ssDNA and can also potentiate HR through its ability to stimulate DNA2-dependent end-resection by binding to RPA (Chen et al., 2013; Xue et al., 2013; Sturzenegger et al., 2014). It is possible that loss of BLM activity in BOD1L/BLM knockdown cells has two effects: (1) increases RAD51 filament stability and; (2) compromises DNA2- dependent resection of damaged forks, the latter of which causes the genome instability apparent in BOD1L-deficient cells. "
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    ABSTRACT: Recognition and repair of damaged replication forks are essential to maintain genome stability and are coordinated by the combined action of the Fanconi anemia and homologous recombination pathways. These pathways are vital to protect stalled replication forks from uncontrolled nucleolytic activity, which otherwise causes irreparable genomic damage. Here, we identify BOD1L as a component of this fork protection pathway, which safeguards genome stability after replication stress. Loss of BOD1L confers exquisite cellular sensitivity to replication stress and uncontrolled resection of damaged replication forks, due to a failure to stabilize RAD51 at these forks. Blocking DNA2-dependent resection, or downregulation of the helicases BLM and FBH1, suppresses both catastrophic fork processing and the accumulation of chromosomal damage in BOD1L-deficient cells. Thus, our work implicates BOD1L as a critical regulator of genome integrity that restrains nucleolytic degradation of damaged replication forks. Copyright © 2015 Elsevier Inc. All rights reserved.
    No preview · Article · Jul 2015 · Molecular cell
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    • "Recombination intermediates can be alternatively resolved by the RecQ helicase Sgs1 that acts in a complex with the DNA topoisomerase Top3 and the stimulatory protein Rmi1 [30] [31] [32]. In addition, Sgs1/BLM plays multiple roles during replication, promoting intra-S phase checkpoint activation and preventing deleterious recombination events [33] [34] [35] [36]. Interestingly, lack of Uls1 moderately suppresses the sensitivity of sgs1 to HU and MMS [23]. "
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    ABSTRACT: The Saccharomyces cerevisiae Uls1 belongs to the Swi2/Snf2 family of DNA-dependent ATPases and a new protein family of SUMO-targeted ubiquitin ligases. Here we show that Uls1 is implicated in DNA repair independently of the replication stress response pathways mediated by the endonucleases Mus81 and Yen1 and the helicases Mph1 and Srs2. Uls1 works together with Sgs1 and we demonstrate that the attenuation of replication stress-related defects in sgs1Δ by deletion of ULS1 depends on a functional of Rad51 recombinase and post-replication repair pathway mediated by Rad18 and Rad5, but not on the translesion polymerase, Rev3. The higher resistance of sgs1Δ uls1Δ mutants to genotoxic stress compared to single sgs1Δ cells is not the result of decreased formation or accelerated resolution of recombination-dependent DNA structures. Instead, deletion of ULS1 restores stability of the rDNA region in sgs1Δ cells. Our data suggest that Uls1 may contribute to genomic stability during DNA synthesis and channel the repair of replication lesions into the Sgs1-dependent pathway, with DNA translocase and SUMO binding activities of Uls1 as well as a RING domain being essential for its functions in replication stress response.
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