The carboxy terminus of NBS1 is required for induction of apoptosis by the MRE11 complex

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Abstract

The MRE11 complex (MRE11, RAD50 and NBS1) and the ataxia-telangiectasia mutated (ATM) kinase function in the same DNA damage response pathway to effect cell cycle checkpoint activation and apoptosis1,2,3. The functional interaction between the MRE11 complex and ATM has been proposed to require a conserved C-terminal domain of NBS1 for recruitment of ATM to sites of DNA damage4,5. Human Nijmegen breakage syndrome (NBS) cells and those derived from multiple mouse models of NBS express a hypomorphic NBS1 allele that exhibits impaired ATM activity despite having an intact C-terminal domain3,6,7,8,9,10,11. This indicates that the NBS1 C terminus is not sufficient for ATM function. We derived Nbs1ΔC/ΔC mice in which the C-terminal ATM interaction domain is deleted. Nbs1ΔC/ΔC cells exhibit intra-S-phase checkpoint defects, but are otherwise indistinguishable from wild-type cells with respect to other checkpoint functions, ionizing radiation sensitivity and chromosome stability. However, multiple tissues of Nbs1ΔC/ΔC mice showed a severe apoptotic defect, comparable to that of ATM- or CHK2-deficient animals. Analysis of p53 transcriptional targets and ATM substrates showed that, in contrast to the phenotype of Chk2-/- mice, NBS1ΔC does not impair the induction of proapoptotic genes. Instead, the defects observed in Nbs1ΔC/ΔC result from impaired phosphorylation of ATM targets including SMC1 and the proapoptotic factor, BID.

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Figure 1: Generation of Nbs1 ΔC/ΔC mice.
Figure 2: Cellular phenotypes of Nbs1ΔC/ΔC.
Figure 3: Apoptotic phenotypes of Nbs1ΔC/ΔC.
Figure 4: Apoptotic signalling in Nbs1ΔC/ΔC.

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Acknowledgements

We thank N. Copeland, N. Jenkins, and C. Adelman for assistance with recombineering and ES cell culture, J. Theunissen for assistance with checkpoint and apoptotic analysis, G. Oltz and E. Rhuley for AC1 ES cells, Y. Shiloh for anti-ATM (MAT3) antibodies, and Petrini laboratory members for helpful suggestions. T.H.S. was supported by an NRSA fellowship and this work was supported by NIH grants awarded to J.H.P. and the Joel and Joan Smilow Initiative.

Author Contributions T.H.S. and J.H.P. conceived the experiments and wrote the paper. T.H.S., M.M., S.S.C., and H.H. performed the experiments.

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Correspondence to John H. J. Petrini.

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Stracker, T., Morales, M., Couto, S. et al. The carboxy terminus of NBS1 is required for induction of apoptosis by the MRE11 complex. Nature 447, 218–221 (2007) doi:10.1038/nature05740

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