Autophosphorylation and ATM activation: Additional sites add to the complexity

Sergei V. Kozlov, Mark E. Graham, Burkhard Jakob, Frank Tobias, Amanda W. Kijas, Marcel Tanuji, Philip Chen, Phillip J. Robinson, Gisela Taucher-Scholz, Keiji Suzuki, Sairai So, David Chen, Martin F. Lavin

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Abstract

The recognition and signaling of DNA double strand breaks involves the participation of multiple proteins, including the protein kinase ATM (mutated in ataxia-telangiectasia). ATM kinase is activated in the vicinity of the break and is recruited to the break site by the Mre11-Rad50-Nbs1 complex, where it is fully activated. In human cells, the activation process involves autophosphorylation on three sites (Ser367, Ser1893, and Ser1981) and acetylation on Lys3016. We now describe the identification of a new ATM phosphorylation site, Thr(P)1885 and an additional autophosphorylation site, Ser(P)2996, that is highly DNA damage-inducible. We also confirm that human and murine ATM share five identical phosphorylation sites. We targeted the ATM phosphorylation sites, Ser 367 and Ser2996, for further study by generating phosphospecific antibodies against these sites and demonstrated that phosphorylation of both was rapidly induced by radiation. These phosphorylations were abolished by a specific inhibitor of ATM and were dependent on ATM and the Mre11-Rad50-Nbs1 complex. As found for Ser(P)1981, ATM phosphorylated at Ser367 and Ser2996 localized to sites of DNA damage induced by radiation, but ATM recruitment was not dependent on phosphorylation at these sites. Phosphorylation at Ser367 and Ser2996 was functionally important because mutant forms of ATM were defective in correcting the S phase checkpoint defect and restoring radioresistance in ataxia-telangiectasia cells. These data provide further support for the importance of autophosphorylation in the activation and function of ATM in vivo.

Original languageEnglish (US)
Pages (from-to)9107-9119
Number of pages13
JournalJournal of Biological Chemistry
Volume286
Issue number11
DOIs
StatePublished - Mar 18 2011

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Kozlov, S. V., Graham, M. E., Jakob, B., Tobias, F., Kijas, A. W., Tanuji, M., Chen, P., Robinson, P. J., Taucher-Scholz, G., Suzuki, K., So, S., Chen, D., & Lavin, M. F. (2011). Autophosphorylation and ATM activation: Additional sites add to the complexity. Journal of Biological Chemistry, 286(11), 9107-9119. https://doi.org/10.1074/jbc.M110.204065