ATM phosphorylates p95/nbs1 in an S-phase checkpoint pathway

The rare diseases ataxia-telangiectasia (AT), caused by mutations in the AT M gene, and Nijmegen breakage syndrome (NBS), with mutations in the p95/nbs 1 gene, share a variety of phenotypic abnormalities such as chromosomal ins tability, radiation sensitivity and defects in cell-cycle checkpoints in re sponse to ionizing radiation(1-4). The ATM gene encodes a protein kinase th at is activated by ionizing radiation or radiomimetic drugs(5,6), whereas p 95/nbs1 is part of a protein complex that is involved in responses to DNA d ouble-strand breaks(3,7). Here, because of the similarities between AT and NBS, we evaluated the functional interactions between ATM and p95/nbs1. Act ivation of the ATM kinase by ionizing radiation and induction of ATM-depend ent responses in NBS cells indicated that p95/nbs1 may not be required for signalling to ATM after ionizing radiation. However, p95/nbs1 was phosphory lated on serine 343 in an ATM-dependent manner in vitro and in vivo after i onizing radiation. A p95/nbs1 construct mutated at the ATM phosphorylation site abrogated an S-phase checkpoint induced by ionizing radiation in norma l cells and failed to compensate for this functional deficiency in NBS cell s. These observations link ATM and p95/nbs1 in a common signalling pathway and provide an explanation for phenotypic similarities in these two disease s.