Molecular basis of human ATM kinase inhibition

Stakyte, K.; Rotheneder, M.; Lammens, Katja; Bartho, J.D.; Grädler, Ulrich; Fuchß, Thomas; Pehl, Ulrich; Alt, Aaron; Logt, Erik van de; Hopfner, Karl‐Peter

doi:10.1038/s41594-021-00654-x

Cited by 36 publications

(35 citation statements)

References 74 publications

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“…The refined model contains over 90% of the ATM residues, with the remaining residues in poorly-ordered or disordered loops. While this manuscript was being prepared, a cryo-EM structure of ATM bound to the inhibitor KU-55933 was reported at an overall resolution of 2.8 Å ( Stakyte et al, 2021 ). Our structure of ATM bound to AMP-PNP is highly similar with a root mean square deviation (RMSD) of 0.9 Å based on 2,453 aligned Cα atoms.…”

Section: Resultsmentioning

confidence: 99%

Structure of the human ATM kinase and mechanism of Nbs1 binding

Warren

Pavletich

2022

eLife

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DNA double-strand breaks (DSBs) can lead to mutations, chromosomal rearrangements, genome instability, and cancer. Central to the sensing of DSBs is the ATM (Ataxia-telangiectasia mutated) kinase, which belongs to the phosphatidylinositol 3-kinase-related protein kinase (PIKK) family. In response to DSBs, ATM is activated by the MRN (Mre11-Rad50-Nbs1) protein complex through a poorly-understood process that also requires double-stranded DNA. Previous studies indicate that the FxF/Y motif of Nbs1 directly binds to ATM, and is required to retain active ATM at sites of DNA damage. Here we report the 2.5 Å resolution cryo-EM structures of human ATM and its complex with the Nbs1 FxF/Y motif. In keeping with previous structures of ATM and its yeast homolog Tel1, the dimeric human ATM kinase adopts a symmetric, butterfly-shaped structure. The conformation of the ATM kinase domain is most similar to the inactive states of other PIKKs, suggesting that activation may involve an analogous realigning the N and C lobes along with relieving the blockage of the substrate-binding site. We also show that the Nbs1 FxF/Y motif binds to a conserved hydrophobic cleft within the Spiral domain of ATM, suggesting an allosteric mechanism of activation. We evaluate the importance of these structural findings with mutagenesis and biochemical assays.

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Section: Resultsmentioning

confidence: 99%

Structure of the human ATM kinase and mechanism of Nbs1 binding

Warren

Pavletich

2022

eLife

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“…Derived discoveries will in turn lead to novel, testable hypotheses concerning clinical aspects of p53 with links to the development of drug-resistance or cancer progression ( Somarelli, Gardner, et al 2020 ; Salomao et al 2021 ). Accumulating evidence present how comparative genomic studies aiming to shed light on complex human traits with several translational aspects ( Sulak et al 2016 ; Tollis et al 2019 ; Somarelli, Boddy, et al 2020 ; Somarelli, Gardner, et al 2020 ; Farre et al 2021 ; Stakyte et al 2021 ) indicate the structural basis of the interfaces, which can be used for the development of highly specific antibodies to target neoantigens and cancer mutations that are difficult to target in conventional ways ( Hsiue et al 2021 ).…”

Section: Discussionmentioning

confidence: 99%

The Elephant Evolved p53 Isoforms that Escape MDM2-Mediated Repression and Cancer

Padariya

Jooste

Hupp

et al. 2022

Molecular Biology and Evolution

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The p53 tumour suppressor is a transcription factor with roles in cell development, apoptosis, oncogenesis, ageing and homeostasis in response to stresses and infections. p53 is tightly regulated by the MDM2 E3 ubiquitin ligase. The p53-MDM2 pathway has co-evolved, with MDM2 remaining largely conserved while the TP53 gene morphed into various isoforms. Studies on pre-vertebrate ancestral homologues revealed the transition from an environmentally-induced mechanism activating p53 to a tightly regulated system involving cell signaling. The evolution of this mechanism depends on structural changes in the interacting protein motifs. Elephants such as Loxodonta africana constitute ideal models to investigate this co-evolution as they are large and long-living as well as having 20 copies of TP53 isoformic sequences expressing a variety of BOX-I MDM2-binding motifs. Collectively, these isoforms would enhance sensitivity to cellular stresses, such as DNA damage, presumably accounting for strong cancer defenses and other adaptations favouring healthy ageing. Here we investigate the molecular evolution of the p53-MDM2 system by combining in silico modelling and in vitro assays to explore structural and functional aspects of p53 isoforms retaining the MDM2 interaction while forming distinct pools of cell-signalling. The methodology used demonstrates, for the first time, that in silico docking simulations can be used to explore functional aspects of elephant p53 isoforms. Our observations elucidate structural and mechanistic aspects of p53 regulation, facilitate understanding of complex cell signalling and suggest testable hypotheses of p53 evolution referencing Peto’s Paradox.

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“…The maps show clear density for the majority of the side chains, allowing for the sequence assignment of the Pincer and Spiral domains and the accurate mapping of conserved residues and cancer-associated missense mutations (Supplemental Figures 3 to 7). While this manuscript was being prepared, a cryo-EM structure of ATM bound to the inhibitor KU-55933 was reported at an overall resolution of 2.8 Å 40 . Our structure of ATM bound to AMP-PNP is highly similar with a root mean square deviation (RMSD) of 0.9 Å based on 2453 aligned Cα atoms.…”

Section: Resultsmentioning

confidence: 99%

Structure of the human ATM kinase and mechanism of Nbs1 binding

Warren

Pavletich

2021

Preprint

View full text Add to dashboard Cite

DNA double-strand breaks (DSBs) can lead to mutations, chromosomal rearrangements, genome instability, and ultimately cancer. Central to the sensing of DSBs are ATM (Ataxia telangiectasia mutated) kinase, which belongs to the phosphatidylinositol 3-kinase-related protein kinase (PIKK) family, and the MRN (Mre11-Rad50-Nbs1) protein complex that activates ATM. How the MRN complex recruits and activates ATM kinase is poorly understood. Previous studies indicate that the FxF/Y motif of Nbs1 directly binds to ATM kinase, and is required to retain active ATM at sites of DNA damage. Here, we report the 2.5 Å resolution cryo-EM structures of human ATM and its complex with the Nbs1 FxF/Y motif. In keeping with previous structures of ATM and its yeast homolog Tel1, the dimeric human ATM kinase adopts a symmetric, butterfly-shaped autoinhibited structure. The conformation of the ATM kinase domain is most similar to the inactive states of other PIKKs, suggesting that activation may involve an analogous realigning the N and C lobes along with relieving the blockage of the substrate-binding site. We show that the Nbs1 FxF/Y motif binds to a conserved hydrophobic cleft within the Spiral domain of ATM, suggesting an allosteric mechanism of activation. We evaluate the importance of these interactions with mutagenesis and biochemical assays.

show abstract

Molecular basis of human ATM kinase inhibition

Cited by 36 publications

References 74 publications

Structure of the human ATM kinase and mechanism of Nbs1 binding

Structure of the human ATM kinase and mechanism of Nbs1 binding

The Elephant Evolved p53 Isoforms that Escape MDM2-Mediated Repression and Cancer

Structure of the human ATM kinase and mechanism of Nbs1 binding

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