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Irradiation-induced G2/M checkpoint response requires ERK1/2 activation

Oncogene volume 26, pages 4689–4698 (2007)Cite this article

Abstract

Following DNA damage, cells undergo G2/M cell cycle arrest, allowing time for DNA repair. G2/M checkpoint activation involves activation of Wee1 and Chk1 kinases and inhibition of Cdc25A and Cdc25C phosphatases, which results in inhibition of Cdc2 kinase. Results presented in this report indicate that γ-irradiation (IR) exposure of MCF-7 cells resulted in extracellular signal regulated protein kinase 1 and 2 (ERK1/2) activation and induction of G2/M arrest. Furthermore, inhibition of ERK1/2 signaling resulted in ⩾85% attenuation in IR-induced G2/M arrest and concomitant diminution of IR-induced activation of ataxia telangiectasia mutated- and rad3-related (ATR), Chk1 and Wee1 kinases as well as phosphorylation of Cdc25A-Thr506, Cdc25C-Ser216 and Cdc2-Tyr15. Moreover, incubation of cells with caffeine, which inhibits ataxia telangiectasia mutated (ATM)/ATR, or transfection of cells with short interfering RNA targeting ATR abrogated IR-induced Chk1 phosphorylation and G2/M arrest but had no effect on IR-induced ERK1/2 activation. In contrast, inhibition of ERK1/2 signaling resulted in marked attenuation in IR-induced ATR activity with little, if any, effect on IR-induced ATM activation. These results implicate IR-induced ERK1/2 activation as an important regulator of G2/M checkpoint response to IR in MCF-7 cells.

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Abbreviations

ATM:

ataxia telangiectasia mutated

ATR:

ATM- and rad3-related

DMSO:

dimethyl sulfoxide

FACS:

fluorescence-activated cell sorting

ERK:

extracellular signal regulated protein kinase

MAPK:

mitogen-activated protein kinase

MEK1/2:

mitogen-activated protein kinase kinase 1 and 2

pERK:

phosphorylated-extracellular signal regulated protein kinase

PI:

propidium iodide

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Acknowledgements

We thank Dr Helen Piwnica-Worms for GST-Cdc25C construct, Dr Junjie Chen for GST-Chk1 construct, Dr Charles A Kuzynski and Linda A Wilkie for assistance on the flow cytometry analysis, Janice Taylor for assistance with Zeiss LSM510 confocal laser scanning microscope and Dr Janina Baranowska-Kortylewicz for assistance on the operation of Mark I 68A Cesium-137 Irradiator. This work was supported by NSF EPSCoR IDeA Award and Nebraska DHHS-LB506 Grant 2007-45 to YY.

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  1. Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA

    Y Yan, C P Black & K H Cowan

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  1. Y Yan
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  3. K H Cowan
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Correspondence to K H Cowan.

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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc).

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Yan, Y., Black, C. & Cowan, K. Irradiation-induced G2/M checkpoint response requires ERK1/2 activation. Oncogene 26, 4689–4698 (2007). https://doi.org/10.1038/sj.onc.1210268

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