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Structure of the intact ATM/Tel1 kinase.

ABSTRACT: The ataxia-telangiectasia mutated (ATM) protein is an apical kinase that orchestrates the multifaceted DNA-damage response. Normally, ATM kinase is in an inactive, homodimer form and is transformed into monomers upon activation. Besides a conserved kinase domain at the C terminus, ATM contains three other structural modules, referred to as FAT, FATC and N-terminal helical solenoid. Here we report the first cryo-EM structure of ATM kinase, which is an intact homodimeric ATM/Tel1 from Schizosaccharomyces pombe. We show that two monomers directly contact head-to-head through the FAT and kinase domains. The tandem N-terminal helical solenoid tightly packs against the FAT and kinase domains. The structure suggests that ATM/Tel1 dimer interface and the consecutive HEAT repeats inhibit the binding of kinase substrates and regulators by steric hindrance. Our study provides a structural framework for understanding the mechanisms of ATM/Tel1 regulation as well as the development of new therapeutic agents.

SUBMITTER: Wang X

PROVIDER: S-EPMC4894967 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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Structure of the intact ATM/Tel1 kinase.

Wang Xuejuan X Chu Huanyu H Lv Mengjuan M Zhang Zhihui Z Qiu Shuwan S Liu Haiyan H Shen Xuetong X Wang Weiwu W Cai Gang G

Nature communications 20160527

The ataxia-telangiectasia mutated (ATM) protein is an apical kinase that orchestrates the multifaceted DNA-damage response. Normally, ATM kinase is in an inactive, homodimer form and is transformed into monomers upon activation. Besides a conserved kinase domain at the C terminus, ATM contains three other structural modules, referred to as FAT, FATC and N-terminal helical solenoid. Here we report the first cryo-EM structure of ATM kinase, which is an intact homodimeric ATM/Tel1 from Schizosaccha ...[more]

PMID: 27229179

Similar Datasets

Structural basis of allosteric regulation of Tel1/ATM kinase.

Project description:ATM/Tel1 is an apical kinase that orchestrates the multifaceted DNA damage response. Mutations of ATM/Tel1 are associated with ataxia telangiectasia syndrome. Here, we report cryo-EM structures of symmetric dimer (4.1?Å) and asymmetric dimer (4.3?Å) of Saccharomyces cerevisiae Tel1. In the symmetric state, the side chains in Tel1 C-terminus (residues 1129-2787) are discernible and an atomic model is built. The substrate binding groove is completely embedded in the symmetric dimer by the intramolecular PRD and intermolecular LID domains. Point mutations in these domains sensitize the S. cerevisiae cells to DNA damage agents and hinder Tel1 activation due to reduced binding affinity for its activator Xrs2/Nbs1. In the asymmetric state, one monomer becomes more compact in two ways: the kinase N-lobe moves down and the Spiral of ?-solenoid moves upwards, which resemble the conformational changes observed in active mTOR. The accessibility of the activation loop correlates with the synergistic conformational disorders in the TRD1-TRD2 linker, FATC and PRD domains, where critical post-translational modifications and activating mutations are coincidently condensed. This study reveals a tunable allosteric network in ATM/Tel1, which is important for substrate recognition, recruitment and efficient phosphorylation.

| S-EPMC6796912 | biostudies-literature

Tel2 mediates activation and localization of ATM/Tel1 kinase to a double-strand break.

Project description:The kinases ATM and ATR (Tel1 and Mec1 in the yeast Saccharomyces cerevisiae) control the response to DNA damage. We report that S. cerevisiae Tel2 acts at an early step of the TEL1/ATM pathway of DNA damage signaling. We show that Tel1 and Tel2 interact, and that even when Tel1 protein levels are high, this interaction is specifically required for Tel1 localization to a DNA break and its activation of downstream targets. Computational analysis revealed structural homology between Tel2 and Ddc2 (ATRIP in vertebrates), a partner of Mec1, suggesting a common structural principle used by partners of phoshoinositide 3-kinase-like kinases.

| S-EPMC2279195 | biostudies-literature

Structure of the human dimeric ATM kinase.

Project description:DNA-double strand breaks activate the serine/threonine protein kinase ataxia-telangiectasia mutated (ATM) to initiate DNA damage signal transduction. This activation process involves autophosphorylation and dissociation of inert ATM dimers into monomers that are catalytically active. Using single-particle electron microscopy (EM), we determined the structure of dimeric ATM in its resting state. The EM map could accommodate the crystal structure of the N-terminal truncated mammalian target of rapamycin (mTOR), a closely related enzyme of the phosphatidylinositol 3-kinase-related protein kinase (PIKK) family, allowing for the localization of the N- and the C-terminal regions of ATM. In the dimeric structure, the actives sites are buried, restricting the access of the substrates to these sites. The unanticipated domain organization of ATM provides a basis for understanding its mechanism of inhibition.

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Activation of Tel1<sup>ATM</sup> kinase requires Rad50 ATPase and long nucleosome-free DNA but no DNA ends.

Project description:In Saccharomyces cerevisiae, Tel1 protein kinase, the ortholog of human ataxia telangiectasia-mutated (ATM), is activated in response to DNA double-strand breaks. Biochemical studies with human ATM and genetic studies in yeast suggest that recruitment and activation of Tel1ATM depends on the heterotrimeric MRXMRN complex, composed of Mre11, Rad50, and Xrs2 (human Nbs1). However, the mechanism of activation of Tel1 by MRX remains unclear, as does the role of effector DNA. Here we demonstrate that dsDNA and MRX activate Tel1 synergistically. Although minimal activation was observed with 80-mer duplex DNA, the optimal effector for Tel1 activation is long, nucleosome-free DNA. However, there is no requirement for DNA double-stranded termini. The ATPase activity of Rad50 is critical for activation. In addition to DNA and Rad50, either Mre11 or Xrs2, but not both, is also required. Each of the three MRX subunits shows a physical association with Tel1. Our study provides a model of how the individual subunits of MRX and DNA regulate Tel1 kinase activity.

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Tel1(ATM)-mediated interference suppresses clustered meiotic double-strand-break formation.

Project description:Meiotic recombination is a critical step in gametogenesis for many organisms, enabling the creation of genetically diverse haploid gametes. In each meiotic cell, recombination is initiated by numerous DNA double-strand breaks (DSBs) created by Spo11, the evolutionarily conserved topoisomerase-like protein, but how these DSBs are distributed relatively uniformly across the four chromatids that make up each chromosome pair is poorly understood. Here we employ Saccharomyces cerevisiae to demonstrate distance-dependent DSB interference in cis (in which the occurrence of a DSB suppresses adjacent DSB formation)--a process that is mediated by the conserved DNA damage response kinase, Tel1(ATM). The inhibitory function of Tel1 acts on a relatively local scale, while over large distances DSBs have a tendency to form independently of one another even in the presence of Tel1. Notably, over very short distances, loss of Tel1 activity causes DSBs to cluster within discrete zones of concerted DSB activity. Our observations support a hierarchical view of recombination initiation where Tel1(ATM) prevents clusters of DSBs, and further suppresses DSBs within the surrounding chromosomal region. Such collective negative regulation will help to ensure that recombination events are dispersed evenly and arranged optimally for genetic exchange and efficient chromosome segregation.

| S-EPMC7116500 | biostudies-literature

Tel1/ATM prevents degradation of replication forks that reverse after topoisomerase poisoning.

Project description:In both yeast and mammals, the topoisomerase poison camptothecin (CPT) induces fork reversal, which has been proposed to stabilize replication forks, thus providing time for the repair of CPT-induced lesions and supporting replication restart. We show that Tel1, the Saccharomyces cerevisiae orthologue of human ATM kinase, stabilizes CPT-induced reversed forks by counteracting their nucleolytic degradation by the MRX complex. Tel1-lacking cells are hypersensitive to CPT specifically and show less reversed forks in the presence of CPT The lack of Mre11 nuclease activity restores wild-type levels of reversed forks in CPT-treated tel1? cells without affecting fork reversal in wild-type cells. Moreover, Mrc1 inactivation prevents fork reversal in wild-type, tel1?, and mre11 nuclease-deficient cells and relieves the hypersensitivity of tel1? cells to CPT Altogether, our data indicate that Tel1 counteracts Mre11 nucleolytic activity at replication forks that undergo Mrc1-mediated reversal in the presence of CPT.

| S-EPMC6030699 | biostudies-literature

A kinase-independent role for the Rad3(ATR)-Rad26(ATRIP) complex in recruitment of Tel1(ATM) to telomeres in fission yeast.

Project description:ATM and ATR are two redundant checkpoint kinases essential for the stable maintenance of telomeres in eukaryotes. Previous studies have established that MRN (Mre11-Rad50-Nbs1) and ATRIP (ATR Interacting Protein) interact with ATM and ATR, respectively, and recruit their partner kinases to sites of DNA damage. Here, we investigated how Tel1(ATM) and Rad3(ATR) recruitment to telomeres is regulated in fission yeast. Quantitative chromatin immunoprecipitation (ChIP) assays unexpectedly revealed that the MRN complex could also contribute to the recruitment of Tel1(ATM) to telomeres independently of the previously established Nbs1 C-terminal Tel1(ATM) interaction domain. Recruitment of Tel1(ATM) to telomeres in nbs1-c60Delta cells, which lack the C-terminal 60 amino acid Tel1(ATM) interaction domain of Nbs1, was dependent on Rad3(ATR)-Rad26(ATRIP), but the kinase domain of Rad3(ATR) was dispensable. Thus, our results establish that the Rad3(ATR)-Rad26(ATRIP) complex contributes to the recruitment of Tel1(ATM) independently of Rad3(ATR) kinase activity, by a mechanism redundant with the Tel1(ATM) interaction domain of Nbs1. Furthermore, we found that the N-terminus of Nbs1 contributes to the recruitment of Rad3(ATR)-Rad26(ATRIP) to telomeres. In response to replication stress, mammalian ATR-ATRIP also contributes to ATM activation by a mechanism that is dependent on the MRN complex but independent of the C-terminal ATM interaction domain of Nbs1. Since telomere protection and DNA damage response mechanisms are very well conserved between fission yeast and mammalian cells, mammalian ATR-ATRIP may also contribute to the recruitment of ATM to telomeres and to sites of DNA damage independently of ATR kinase activity.

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Fission yeast Tel1(ATM) and Rad3(ATR) promote telomere protection and telomerase recruitment.

Project description:The checkpoint kinases ATM and ATR are redundantly required for maintenance of stable telomeres in diverse organisms, including budding and fission yeasts, Arabidopsis, Drosophila, and mammals. However, the molecular basis for telomere instability in cells lacking ATM and ATR has not yet been elucidated fully in organisms that utilize both the telomere protection complex shelterin and telomerase to maintain telomeres, such as fission yeast and humans. Here, we demonstrate by quantitative chromatin immunoprecipitation (ChIP) assays that simultaneous loss of Tel1(ATM) and Rad3(ATR) kinases leads to a defect in recruitment of telomerase to telomeres, reduced binding of the shelterin complex subunits Ccq1 and Tpz1, and increased binding of RPA and homologous recombination repair factors to telomeres. Moreover, we show that interaction between Tpz1-Ccq1 and telomerase, thought to be important for telomerase recruitment to telomeres, is disrupted in tel1Delta rad3Delta cells. Thus, Tel1(ATM) and Rad3(ATR) are redundantly required for both protection of telomeres against recombination and promotion of telomerase recruitment. Based on our current findings, we propose the existence of a regulatory loop between Tel1(ATM)/Rad3(ATR) kinases and Tpz1-Ccq1 to ensure proper protection and maintenance of telomeres in fission yeast.

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Reduced Crossover Interference and Increased ZMM-Independent Recombination in the Absence of Tel1/ATM.

Project description:Meiotic recombination involves the repair of double-strand break (DSB) precursors as crossovers (COs) or noncrossovers (NCOs). The proper number and distribution of COs is critical for successful chromosome segregation and formation of viable gametes. In budding yeast the majority of COs occurs through a pathway dependent on the ZMM proteins (Zip2-Zip3-Zip4-Spo16, Msh4-Msh5, Mer3), which form foci at CO-committed sites. Here we show that the DNA-damage-response kinase Tel1/ATM limits ZMM-independent recombination. By whole-genome mapping of recombination products, we find that lack of Tel1 results in higher recombination and reduced CO interference. Yet the number of Zip3 foci in tel1? cells is similar to wild type, and these foci show normal interference. Analysis of recombination in a tel1? zip3? double mutant indicates that COs are less dependent on Zip3 in the absence of Tel1. Together these results reveal that in the absence of Tel1, a significant proportion of COs occurs through a non-ZMM-dependent pathway, contributing to a CO landscape with poor interference. We also see a significant change in the distribution of all detectable recombination products in the absence of Tel1, Sgs1, Zip3, or Msh4, providing evidence for altered DSB distribution. These results support the previous finding that DSB interference depends on Tel1, and further suggest an additional level of DSB interference created through local repression of DSBs around CO-designated sites.

| S-EPMC4549261 | biostudies-literature

Kinase-independent functions of TEL1 in telomere maintenance.

Project description:TEL1 is important in Saccharomyces cerevisiae telomere maintenance, and its kinase activity is required. Tel1p associates with telomeres in vivo, is enriched at short telomeres, and enhances the binding of telomerase components to short telomeres. However, it is unclear how the kinase activity and telomere association contribute to Tel1p's overall function in telomere length maintenance. To investigate this question, we generated a set of single point mutants and a double point mutant (tel1(KD)) of Tel1p that were kinase deficient and two Xrs2p mutants that failed to bind Tel1p. Using these separation-of-function alleles in a de novo telomere elongation assay, we found, surprisingly, that the tel1(KD) allele and xrs2 C-terminal mutants were both partially functional. Combining the tel1(KD) and xrs2 C-terminal mutants had an additive effect and resembled the TEL1 null (tel1Delta) phenotype. These data indicate that Tel1p has two separate functions in telomere maintenance and that the Xrs2p-dependent recruitment of Tel1p to telomeres plays an important role even in the absence of its kinase activity.

| S-EPMC2738284 | biostudies-literature

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