ATM-dependent phosphorylation of nibrin in response to radiation exposure

Mutations in the gene ATM are responsible for the genetic disorder ataxia-t elangiectasia (A-T), which is characterized by cerebellar dysfunction, radi osensitivity, chromosomal instability and cancer predisposition. Both the A -T phenotype and the similarity of the ATM protein to other DNA-damage sens ors suggests a role for ATM in biochemical pathways involved in the recogni tion, signalling and repair of DNA double-strand breaks (DSBs). Them are st rong parallels between the pattern of radiosensitivity, chromosomal instabi lity and cancer predisposition in A-T patients and that in patients with Ni jmegen breakage syndrome (NBS). The protein defective in NBS, nibrin (encod ed by NBS1), forms a complex with MRE11 and RAD50 (refs 1,2). This complex localizes to DSBs within 30 minutes after cellular exposure to ionizing rad iation (1R) and is observed in brightly staining nuclear foci after a longe r period of time(3). The overlap between clinical and cellular phenotypes i n A-T and NBS suggests that ATM and nibrin may function in the same biochem ical pathway. Here we demonstrate that nibrin is phosphorylated within one hour of treatment of cells with IR. This response is abrogated in A-T cells that either do not express ATM protein or express near full-length mutant protein. We also show that ATM physically interacts with and phosphorylates nibrin on serine 343 both in vivo and in vitro. Phosphorylation of this si te appears to be functionally important because mutated nibrin (S343A) does not completely complement radiosensitivity in NBS cells. ATM phosphorylati on of nibrin does not affect nibrin-MRE11-RAD50 association as revealed by radiation-induced foci formation. Our data provide a biochemical explanatio n for the similarity in phenotype between A-T and NBS.