Characterization of the mechanisms of ATM activation by the MRN complex and DNA
The complex containing the Mre11, Rad50, and Nbs1 proteins (MRN) is essential for the cellular response to DNA double-strand breaks, integrating DNA repair with the activation of checkpoint signaling through the protein kinase ATM (ataxia Telangiectasia mutated). The ATM kinase signals the presence of DNA double-strand breaks in mammalian cells by phosphorylating proteins which initiate cell cycle arrest or apoptosis. We demonstrate that MRN stimulates the kinase activity of ATM in vitro toward its substrates p53, Chk2, and histone H2AX. We also show that the MRN complex acts as the double-strand break sensor for ATM and recruits ATM to broken DNA molecules. Inactive ATM dimers can be activated in vitro with DNA in the presence of MRN, leading to phosphorylation of downstream targets p53 and Chk2. ATM dimers are dissociated into monomers by MRN in a process that does not require ATM autophosphorylation. Unwinding of DNA ends by MRN is required for ATM stimulation, consistent with the central role of single-stranded DNA as an evolutionary conserved signal for DNA damage.