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Oncogene (2002) 21, 4191 4199 2002 Nature Publishing Group All rights reserved 0950 9232/02 $25.00www.nature.com/oncORIGINAL PAPERSNbs1 promotes ATM dependent phosphorylation events including those
required for G1/S arrestPierre-Marie Girard1, Enriqueta Riballo1, Adrian C Begg2, Alastair Waugh1 and Penny A Jeggo*,11MRC Cell Mutation Unit, University of Sussex, Brighton, East Sussex, BN1 9RR, UK; 2Department of Experimental Therapy,
The Netherlands Cancer Institute/Antoni van Leeuwenhoekhuis, Plesmanlaan 21, 1066 CX Amsterdam, The NetherlandsCell lines from Nijmegen Breakage Syndrome (NBS)
and ataxia telangiectasia (A-T) patients show defective S
phase checkpoint arrest. In contrast, only A-T but not
NBS cells are significantly defective in radiation-induced
G1/S arrest. Phosphorylation of some ATM substrates
has been shown to occur in NBS cells. It has, therefore,
been concluded that Nbs1 checkpoint function is S phase
specific. Here, we have compared NBS with A-T cell
lines (AT-5762ins137) that express a low level of normal
ATM protein to evaluate the impact of residual Nbs1
function in NBS cells. The radiation-induced cell cycle
response of these NBS and leaky A-T cells is almost
identical; normal G2/M arrest after 2 Gy, intermediate
G1/S arrest depending on the dose and an A-T-like S
phase checkpoint defect. Thus, the checkpoint assays
dier in their sensitivity to low ATM activity. Radiationinduced phosphorylation of the ATM-dependent
substrates Chk2, RPAp34 and p53-Ser15 are similarly
impaired in AT-5762ins137 and NBS cells in a dose
dependent manner. In contrast, NBS cells show normal
ability to activate ATM kinase following irradiation in
vitro and in vivo. We propose that Nbs1 facilitates
ATM-dependent phosphorylation of multiple downstream
substrates, including those required for G1/S arrest.
Oncogene (2002) 21, 4191 4199. doi:10.1038/sj.onc.
1205596Keywords: NBS; ATM; cell cycle checkpoint; ionizing
radiation; kinase activityIntroductionThe presence of DNA double-strand breaks (DSBs) in
mammalian cells elicits a complex array of responses
that result in the onset of apoptosis, cell cycle
checkpoint arrest and/or mechanisms of DNA repair.
Central to these processes is ATM (ataxia-telangiectasia mutated), the protein defective in ataxiatelangiectasia, a hereditary disorder associated with
cancer predisposition and radiosensitivity (Shiloh,
1997). ATM is a serine-threonine PI 3-K related kinase
which becomes activated following exposure to ionizing
radiation (IR) (Banin et al., 1998; Canman et al., 1998;
Savitsky et al., 1995). ATM phosphorylates in vivo a
number of proteins involved in cell cycle checkpoint
control,...