Oncogene (2007) 26, 7749–7758 & 2007 Nature Publishing Group All rights reserved 0950-9232/07 $30.00 www.nature.com/onc REVIEW ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks MF Lavin1,2 1Radiation Biology and Oncology Laboratory, Queensland Institute of Medical Research, Brisbane, Queensland, Australia and 2Faculty of Health Sciences, University of Queensland, Brisbane, Queensland, Australia The recognition and repair of DNA double-strand breaks rearrangement of immunoglobulin and T-cell receptor (DSBs) is a complex process that draws upon a multitude genes (Livak, 2004; Dudley et al., 2005). of proteins. This is not surprising since this is a lethal Induced DNA DSBs are potentially lethal to the cell and lesion if left unrepaired and also contributes to genome therefore must be rapidly recognized and repaired to avoid instability and the consequential riskof cancer and other genetic damage (Agarwal et al., 2006). Not unexpectedly pathologies. Some of the key proteins that recognize these there are cellular mechanisms in place to recognize and breaks in DNA are mutated in distinct genetic disorders signal the presence of DNA DSB to the cell-cycle that predispose to agent sensitivity, genome instability, checkpoints to delay the passage of cells through the cycle cancer predisposition and/or neurodegeneration. These and facilitate DNA repair (Zhou and Elledge, 2000). include members of the Mre11 complex (Mre11/Rad50/ Signalling also occurs to the DNA repair machinery. Two Nbs1) and ataxia-telangiectasia (A-T) mutated (ATM), majormechanisms exist forthe repairofthe DNA DSB, mutated in the human genetic disorder A-T. The mre11 homologous recombination repair and non-homologous (MRN) complex appears to be the major sensor of the end joining (NHEJ) (Valerie and Povirk, 2003). Ulti- breaks and subsequently recruits ATM where it is mately, it is paramount that the break be repaired by one activated to phosphorylate in turn members of that of these mechanisms, but this is preceded by detection, complex and a variety of other proteins involved in cell- recognition and signalling of the break to other cellular cycle control and DNA repair. The MRN complex is also processes such as cell-cycle control and transcriptional upstream of ATM and ATR (A-T-mutated and rad3- events (Zhou and Elledge, 2000). Most progress on the related) protein in responding to agents that blockDNA recognition and signalling of DNA DSB has been acquired replication. To date, more than 30 ATM-dependent from a series of rare human genetic disorders sharing substrates have been identified in multiple pathways that sensitivity to agents that cause breaks in DNA (Lukas maintain genome stability and reduce the riskof disease. et al., 2006). Prominent among these are ataxia-telangiec- We focus here on the relationship between ATM and the tasia (A-T) with A-T-mutated (ATM) being the mutated MRN complex in recognizing and responding to DNA protein; Nijmegen breakage syndrome (NBS), Nbs1 DSBs. defective; A-T like syndrome (A-TLD), Mre11 defective; Oncogene (2007) 26, 7749–7758; doi:10.1038/sj.onc.1210880 and A-T and Rad3 related (ATR) (Seckels), ATR defective. (Table 1). A single case with a defect in Rad50 Keywords: ATM; Mre11 complex; DNA double strand protein has also been described (Dork et al., in prepara- breaks; signal transduction; functional consequences tion). It should be pointed out that while the gene products in all of these syndromes recognize and signal DNA DSB, they do not show dramatic reductions in capacity to repair these breaks (Foray et al., 1997; Girard et al., 2000; Riballo et al., 2004). On the otherhand, defects in DNA-PKcs, Introduction Ku70/80 and ligase IV, all of which participate in NHEJ, are associated with more marked defects in the capacity to Exposure of cells to ionizing radiation and radiomimetic repair DNA DSB (Featherstone and Jackson, 1999; Meek chemicals gives rise to a variety of DNA damage, including et al., 2004; Orii et al., 2006). In this review, the emphasis double-strand breaks (DSB) (Ward, 1985). DNA damage will be on the mre11 (MRN) complex and ATM. As we during S phase can lead to the collapse of DNA replication shall see the MRN complex is the primary sensor of DNA forks also generating DNA DSB (Paulsen and Cimprich, DSB. It recruits ATM to the break where it is activated 2007). DSBs also arise under normal conditions during and subsequently phosphorylates members of the complex and a variety of other proteins that signal to different cellular processes. This ensures that DNA repair complexes Correspondence: Dr MF Lavin, Radiation Biology and Oncology Laboratory, Queensland Institute of Medical Research, Brisbane, efficiently remove and repair the break. Failure to do so Queensland 4029, Australia. results in genome instability, which may give rise to cancer, E-mail: [email protected] neurodegeneration and other pathologies. Recognition and signalling of DNA DSB MF Lavin 7750 Table 1 DNA damage recognition/repair syndromes defective in DNA double-strand break repair Syndrome Defective Mutant Cancer Neurological Developmental/ Agent/sensitivity gene protein susceptibility changes growth delay Ataxia-telangiectasia (A-T) ATM ATM Yes Neurodegeneration No Ionizing radiation A-T-like disorder (ATLD) Mre11 Mre11 No Neurodegeneration No Ionizing radiation Nijmegen breakage syndrome (NS) Nbs1 Nbs1 Yes Microcephaly Yes Ionizing radiation Rad50-deficient patient Rad50 Rad50 ? Microcephaly ? Ionizing radiation Ataxia oculomotorapraxiatype 1 Aptx Aprataxin No Neurodegeneration No H2O2 (AOA1) Ataxia oculomotorapraxiatype 2 Setx Senataxin No Neurodegeneration No H2O2 Spinocerebellar ataxia with axonal Tdp1 TDP1 No Neurodegeneration No H2O2 neuropathy (SCAN1) A-T and Rad30 related disorder ATR ATR No Neurodegeneration No HU, UV (Seckels) DNA ligase Lig IV DNA No Microcephaly Yes Ionizing radiation Ligase IV Abbreviations: ATM, ataxia-telangiectasia mutated; ATR, ataxia-telangiectasia mutated and Rad3 related; NBS, Nijmegen breakage syndrome. C C The MRN complex acts as a sensor of DNA DSB X X X Zn X C C The MRN complex is a highly conserved protein complex involved in the following DNA repair mechanisms: both DNA DNA B homologous recombination repair and NHEJ, DNA B A M M A A A B M N N B replication, telomere maintenance and in signalling to the M cell-cycle checkpoints (D’Amours and Jackson, 2002; van C C den Bosch et al., 2003). The complex is rapidly localized X Zn X X X to nuclear foci in response to radiation exposure (Maser C C et al., 1997), which were shown to be sites of DNA Figure 1 Schematic representation of the binding of the Mre11 damage using irradiation masks and ultrasoft X-rays complex to DNA free ends. These ends are tethered by interaction (Nelms et al., 1998). It was subsequently shown that of the hinge regions of Rad50 molecules, assisted by co-ordination ATM, which phosphorylates Nbs1 for activation of the with Zn þ 2 ions. S-phase checkpoint, was not required for association of the MRN complex with sites of DNA damage (Mirzoeva and Petrini, 2001). However, the complex also binds Artemis, also play a role in DSB processing at least in a tightly to chromatin in the absence of DNA damage subgroup of breaks with damaged termini (Riballo et al., during S phase (Mirzoeva and Petrini, 2003). The Mre11/ 2004). Thus, the initial event in recognizing and Rad50 complex binds to DNA as a heterotetramer, responding to DNA DSB is the binding of the Mre11 tethering broken ends of a DSB (de Jager et al., 2001). complex, which tethers the broken ends together as a The binding appears to be achieved through the two means of preparing for repair. DNA-binding motifs of Mre11 (van den Bosch et al., 2003). This is arranged as a globular domain with Rad50 WalkerA and B motifs (ATPase domains) and the bridging of DNA molecules is achieved through CXXC ATM is recruited to and activated by the MRN complex sequences in the middle of Rad50 (Figure 1). These sequences are displayed at the ends of coiled-coil regions Evidence using rare genetic disorders and mouse models and appearto dimerize by the coordination of a Zn 2 þ ion While the recruitment of the MRN complex to damaged (Hopfner et al., 2002). Upon binding to DNA, the DNA is rapid, so too is the activation of ATM dynamic architecture of the MRN complex is altered to (Bakkenist and Kastan, 2003; Kozlov et al., 2003), give rise to parallel orientation of the coiled-coils of making it difficult to discern the sequence of events Rad50, preventing intracomplex interaction and favour- involved. If the MRN complex were the sensor of DNA ing intercomplex association (Moreno-Herrero et al., breaks then it might be expected that it would be 2005). Association with Rad50 stimulates both the upstream of ATM activation. Evidence for this was exonuclease and endonuclease activities of Mre11 (Paull provided from studies with NBS and A-TLD cells, and Gellert, 1998; Trujillo and Sung, 2001) and Nbs1 hypomorphic for members of the complex; in cells in stimulates its endonuclease activity (Paull and Gellert, which the MRN complex was depleted during viral 1999). The complete complex can also partly unwind or infection; in vitro investigations using recombinant dissociate a short DNA duplex with a 30 overhang and proteins and in Xenopus extracts reconstituted for this activity is stimulated by ATP (Paull and Gellert, DNA damage signalling. In these experiments, ATM 1999). It seems likely that these and otheractivities may activation was determined by autophosphorylation on be responsible for the processing of DNA DSB prior to S1981 or indirectly through its capacity to phosphor- repair. However, it is evident that other nucleases, such as ylate downstream substrates. ATM activation was Oncogene Recognition and signalling of DNA DSB MF Lavin 7751 retarded in NBS cells in response to neocarzinostatin the phenotype of NBS cells and that from Nbs1tr735 treatment (Uziel et al., 2003).