Human Fbh1 helicase contributes to genome maintenance via pro- and anti-recombinase activities

Institute of Cancer Biology and Center for Genotoxic Stress Research, Danish Cancer Society, DK-2100 Copenhagen, Denmark.
The Journal of Cell Biology (Impact Factor: 9.83). 10/2009; 186(5):655-63. DOI: 10.1083/jcb.200812138
Source: PubMed


Homologous recombination (HR) is essential for faithful repair of DNA lesions yet must be kept in check, as unrestrained HR may compromise genome integrity and lead to premature aging or cancer. To limit unscheduled HR, cells possess DNA helicases capable of preventing excessive recombination. In this study, we show that the human Fbh1 (hFbh1) helicase accumulates at sites of DNA damage or replication stress in a manner dependent fully on its helicase activity and partially on its conserved F box. hFbh1 interacted with single-stranded DNA (ssDNA), the formation of which was required for hFbh1 recruitment to DNA lesions. Conversely, depletion of endogenous Fbh1 or ectopic expression of helicase-deficient hFbh1 attenuated ssDNA production after replication block. Although elevated levels of hFbh1 impaired Rad51 recruitment to ssDNA and suppressed HR, its small interfering RNA-mediated depletion increased the levels of chromatin-associated Rad51 and caused unscheduled sister chromatid exchange. Thus, by possessing both pro- and anti-recombinogenic potential, hFbh1 may cooperate with other DNA helicases in tightly controlling cellular HR activity.

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Available from: Jacob Falck Hansen, Apr 16, 2014
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    • "Particularly, the human Fbox DNA helicase protein 1 (FBH1) has been shown to be a key factor in fork reversal when nucleotide supply is scarce. Indeed, FBH1 accumulates at stalling forks following a short treatment of HU [112] and it is responsible for fork reversal either in vitro or in vivo [113]. Additional proteins are involved in fork reversal including SMARCAL1 (SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A-like protein1), which appears to suppress genomic instability arising from RS. Loss of this DNAdependent ATPase leads to Schimke immunoosseous dysplasia (SIOD), a disease characterized by immunodeficiency, skeletal and kidney abnormalities [114]. "
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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.
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    • "Cells were grown on coverslips and treated as indicated in the figure legends. The coverslips were fixed and stained as described previously (Fugger et al., 2009). Briefly, the coverslips were fixed in Lillies with or without prior preextraction in CSK buffer (0.5% Triton X-100 in 20 mM HEPES [pH 7.4], 50 mM NaCl, 3 mM MgCl, and 300 mM sucrose), permeabilized in 0.2% Triton X-100/PBS, and incubated with the indicated antibodies diluted in DMEM supplemented with 10% serum. "
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