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Initiation of a G2/M checkpoint after ultraviolet radiation requires p38 kinase

Nature volume 411, pages 102–107 (2001)Cite this article

Abstract

Response to genotoxic stress can be considered as a multistage process involving initiation of cell-cycle arrest and maintenance of arrest during DNA repair1. Although maintenance of G2/M checkpoints is known to involve Chk1, Chk2/Rad53 and upstream components, the mechanisms involved in its initiation are less well defined1,2,3,4. Here we report that p38 kinase has a critical role in the initiation of a G2 delay after ultraviolet radiation. Inhibition of p38 blocks the rapid initiation of this checkpoint in both human and murine cells after ultraviolet radiation. In vitro, p38 binds and phosphorylates Cdc25B at serines 309 and 361, and Cdc25C at serine 216; phosphorylation of these residues is required for binding to 14-3-3 proteins. In vivo, inhibition of p38 prevents both phosphorylation of Cdc25B at serine 309 and 14-3-3 binding after ultraviolet radiation, and mutation of this site is sufficient to inhibit the checkpoint initiation. In contrast, in vivo Cdc25C binding to 14-3-3 is not affected by p38 inhibition after ultraviolet radiation. We propose that regulation of Cdc25B phosphorylation by p38 is a critical event for initiating the G2/M checkpoint after ultraviolet radiation.

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Figure 1: A principal role for p38 kinase in G2 checkpoint activation after UV radiation.
Figure 2: The p38α and p38β kinases contribute to UV-induced G2 arrest.
Figure 3: In vitro p38 kinase phosphorylates and binds Cdc25B and Cdc25C, and regulates Cdc25 binding with 14-3-3 proteins.
Figure 4: In vivo phosphorylation of Ser 309 Cdc25B after UV radiation, and its role in regulating 14-3-3 binding and G2/M checkpoint control. a, The level of 14-3-3 in complex with Cdc25 in vivo was determined after sequential immunoprecipitations (see Methods).

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Acknowledgements

We are grateful to J. Pines, I. Hoffmann, H. Piwnica-Worms, J. Han, B. Vogelstein and R. Davis for the plasmids used in the paper; D. Ferris for Cdc2 polyclonal antibody; B. Monia for advice on the antisense oligonucleotide approach; J. Clark for help with DNA sequencing; and O. Kovalsky for help in protein isolation. 

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Authors and Affiliations

  1. Division of Basic Science and,

    Dmitry V. Bulavin & Albert J. Fornace Jr

  2. Laboratory of Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, 20892, Maryland, USA

    Yuichiro Higashimoto, Venkatesha Basrur & Ettore Appella

  3. Isis Pharmaceuticals Inc., Carlsbad, 92008, California, USA

    Ian J. Popoff & William A. Gaarde

  4. Laboratory of Biological Chemistry, National Institute on Aging, NIH, Baltimore, 21224, Maryland, USA

    Olga Potapova

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  1. Dmitry V. Bulavin
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Correspondence to Albert J. Fornace Jr.

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Bulavin, D., Higashimoto, Y., Popoff, I. et al. Initiation of a G2/M checkpoint after ultraviolet radiation requires p38 kinase. Nature 411, 102–107 (2001). https://doi.org/10.1038/35075107

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