Structure of Mre11-Nbs1 complex yields insights into ataxia-telangiectasia-like disease mutations and DNA damage signaling

Gene Center, Ludwig Maximilians University Munich, Munich, Germany.
Nature Structural & Molecular Biology (Impact Factor: 13.31). 06/2012; 19(7):693-700. DOI: 10.1038/nsmb.2323
Source: PubMed


The Mre11-Rad50-Nbs1 (MRN) complex tethers, processes and signals DNA double-strand breaks, promoting genomic stability. To understand the functional architecture of MRN, we determined the crystal structures of the Schizosaccharomyces pombe Mre11 dimeric catalytic domain alone and in complex with a fragment of Nbs1. Two Nbs1 subunits stretch around the outside of the nuclease domains of Mre11, with one subunit additionally bridging and locking the Mre11 dimer via a highly conserved asymmetrical binding motif. Our results show that Mre11 forms a flexible dimer and suggest that Nbs1 not only is a checkpoint adaptor but also functionally influences Mre11-Rad50. Clinical mutations in Mre11 are located along the Nbs1-interaction sites and weaken the Mre11-Nbs1 interaction. However, they differentially affect DNA repair and telomere maintenance in Saccharomyces cerevisiae, potentially providing insight into their different human disease pathologies.

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    • "Indeed, the flexibility of the Mre11 dimer appears to be higher in eukaryotes, as highlighted by stabilization of Schizosaccharomyces pombe Mre11 dimers by Nbs1 interaction (Schiller et al., 2012). Indeed, in structures of Mre11 with Nbs1, two Nbs1 molecules interact with the Mre11 dimer (Schiller et al., 2012). Only 14 residues of Nbs1 are involved in this interaction (Fig. 5). "
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    ABSTRACT: The Mre11-Rad50-Nbs1 (MRN) complex is a dynamic macromolecular machine that acts in the first steps of DNA double strand break repair, and each of its components has intrinsic dynamics and flexibility properties that are directly linked with their functions. As a result, deciphering the functional structural biology of the MRN complex is driving novel and integrated technologies to define the dynamic structural biology of protein machinery interacting with DNA. Rad50 promotes dramatic long-range allostery through its coiled-coil and zinc-hook domains. Its ATPase activity drives dynamic transitions between monomeric and dimeric forms that can be modulated with mutants modifying the ATPase rate to control end joining versus resection activities. The biological functions of Mre11's dual endo- and exonuclease activities in repair pathway choice were enigmatic until recently, when they were unveiled by the development of specific nuclease inhibitors. Mre11 dimer flexibility, which may be regulated in cells to control MRN function, suggests new inhibitor design strategies for cancer intervention. Nbs1 has FHA and BRCT domains to bind multiple interaction partners that further regulate MRN. One of them, CtIP, modulates the Mre11 excision activity for homologous recombination repair. Overall, these combined properties suggest novel therapeutic strategies. Furthermore, they collectively help to explain how MRN regulates DNA repair pathway choice with implications for improving the design and analysis of cancer clinical trials that employ DNA damaging agents or target the DNA damage response.
    Progress in Biophysics and Molecular Biology 01/2015; 117(2-3). DOI:10.1016/j.pbiomolbio.2014.12.004 · 2.27 Impact Factor
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    • "Two a helices of Mre11 located carboxy terminal to the nuclease core domain are responsible for interaction with the Rad50 coiled-coil base (Lammens et al. 2011; Lim et al. 2011; Williams et al. 2011). Schizosaccharomyces pombe Mre11 interacts with Nbs1 via a eukaryotic-specific insertion between phosphoesterases motifs II and III, referred to as the latching loop, and through additional residues in the amino-terminal region (Schiller et al. 2012). Mutations within the latching loop "
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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 · 8.68 Impact Factor
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    • "ATM is recruited to DNA double strand breaks (DSB) by the Mre11/Rad50/NBN (MRN) complex where it is activated to phosphorylate a multitude of proteins involved in the response to DNA damage [7]. Disorders arising due to mutations in members of the MRN complex are also characterised by defects in the response to DNA DSB [8]. Hypomorphic mutations in Mre11 give rise to A-T like disorder (ATLD), which overlaps in its clinical phenotype with A-T and also features radiosensitivity and cell cycle defects [9]. "
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    ABSTRACT: Disruption of the Setx gene, defective in ataxia oculomotor apraxia type 2 (AOA2) leads to the accumulation of DNA/RNA hybrids (R-loops), failure of meiotic recombination and infertility in mice. We report here the presence of R-loops in the testes from other autosomal recessive ataxia mouse models, which correlate with fertility in these disorders. R-loops were coincident in cells showing high basal levels of DNA double strand breaks and in those cells undergoing apoptosis. Depletion of Setx led to high basal levels of R-loops and these were enhanced further by DNA damage both in vitro and in vivo in tissues with proliferating cells. There was no evidence for accumulation of R-loops in the brains of mice where Setx, Atm, Tdp1 or Aptx genes were disrupted. These data provide further evidence for genome destabilization as a consequence of disrupted transcription in the presence of DNA double strand breaks arising during DNA replication or recombination. They also suggest that R-loop accumulation does not contribute to the neurodegenerative phenotype in these autosomal recessive ataxias.
    PLoS ONE 03/2014; 9(3):e90219. DOI:10.1371/journal.pone.0090219 · 3.23 Impact Factor
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