Autophosphorylation and ATM activation: additional sites add to the complexity

Area of Science:

• Molecular Biology
• Cellular Biology
• Biochemistry

Background:

• DNA double-strand breaks (DSBs) trigger complex signaling pathways for repair.
• ATM (ataxia-telangiectasia mutated) kinase is a key regulator in the DSB response.
• ATM activation involves autophosphorylation and recruitment to break sites by the Mre11-Rad50-Nbs1 complex.

Purpose of the Study:

• To identify and characterize novel ATM phosphorylation sites.
• To investigate the role of specific ATM phosphorylation sites in DNA damage signaling and cellular response.
• To confirm conserved ATM phosphorylation sites between human and murine cells.

Main Methods:

• Mass spectrometry to identify novel ATM phosphorylation sites.
• Generation of phosphospecific antibodies against Ser(367) and Ser(2996).
• Irradiation of cells to induce DNA damage and assess phosphorylation.
• Use of ATM inhibitors and analysis of ATM-deficient cells (ataxia-telangiectasia).

Main Results:

• Identification of Thr(1885) and Ser(2996) as new ATM phosphorylation sites.
• Phosphorylation of Ser(367) and Ser(2996) is rapidly induced by radiation and dependent on ATM and the MRN complex.
• Phosphorylation at Ser(367) and Ser(2996) is crucial for ATM's role in the S phase checkpoint and radioresistance.
• Phosphorylated ATM localizes to DNA damage sites, but recruitment is independent of these specific phosphorylations.

Conclusions:

• ATM autophosphorylation at Ser(367) and Ser(2996) plays a critical role in its activation and function.
• These novel phosphorylation sites are essential for maintaining genomic integrity and cellular survival following DNA damage.
• The findings underscore the importance of post-translational modifications in regulating ATM kinase activity.