Journal
MUTATION RESEARCH-FUNDAMENTAL AND MOLECULAR MECHANISMS OF MUTAGENESIS
Volume 708, Issue 1-2, Pages 28-36Publisher
ELSEVIER
DOI: 10.1016/j.mrfmmm.2011.01.005
Keywords
ATM; DNA damage signaling; p53
Funding
- Lung Cancer Research Foundation
- Frieda G. and Saul F. Shapira BRCA Cancer Research Program
- Lung Cancer SPORE [P50 CA090440]
- Breast Cancer Research Foundation
- [CA148644]
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Ataxia-telangiectasia mutated (ATM) encodes a nuclear serine/threonine protein kinase whose activity is increased in cells exposed to low doses of ionizing radiation (IR). Here we examine ATM kinase activation in cells exposed to either P-32- or P-33-orthophosp hate under conditions typically employed in metabolic labelling experiments. We calculate that the absorbed dose of IR delivered to a 5 cm x 5 cm monolayer of cells incubated in 2 ml media containing 1 mCi of the high-energy (1.70 MeV) beta-particle emitter P-32-orthophosphate for 30 min is similar to 1 Gy IR. The absorbed dose of IR following an otherwise identical exposure to the low-energy (0.24 MeV) beta-particle emitter P-33-orthophosp hate is similar to 0.18 Gy IR. We show that low-energy beta-particles emitted by P-33 induce a greater number of ionizing radiation-induced foci (IRIF) and greater ATM kinase signaling than energetic beta-particles emitted by P-32. Hence, we demonstrate that it is inappropriate to use P-33-orthophosp hate as a negative control for P-32-orthophosphate in experiments investigating DNA damage responses to DNA double-strand breaks (DSBs). Significantly, we show that ATM accumulates in the chromatin fraction when ATM kinase activity is inhibited during exposure to either radionuclide. Finally, we also show that chromosome aberrations accumulate in cells when ATM kinase activity is inhibited during exposure to similar to 0.36 Gy beta-particles emitted by P-33. We therefore propose that direct cellular exposure to P-33-orthophosphate is an excellent means to induce and label the IR-induced. ATM kinase-dependent phosphoproteome. (C) 2011 Elsevier B.V. All rights reserved.
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