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Cdc7 kinase mediates Claspin phosphorylation in DNA replication checkpoint

Oncogene volume 27, pages 3475–3482 (2008)Cite this article

Abstract

Cdc7 kinase is evolutionarily conserved and is involved in initiation and progression of DNA replication. However, roles of Cdc7 in checkpoint responses remain largely unknown. In this study, we show that deletion of the Cdc7 genes in mouse embryonic stem (ES) cells abrogates hydroxyurea (HU)- or UV-induced activation of Chk1. HU-induced Chk1 activation is also impaired in human cancer cell lines in which Cdc7 is depleted by siRNA, and Cdc7-depleted cells are more sensitive to HU treatment. In contrast, ATR and Rad17 are relocated to chromatin in these cells following HU treatment, indicating that stalled DNA replication forks are detected normally. Cdc7-depleted cells exhibit defects in chromatin association and phosphorylation of Claspin, suggesting that Cdc7 exerts its effect at least partially through Claspin. Consistent with this prediction, Cdc7 interacts with and phosphorylates Claspin. We propose that Cdc7 is required for activation of the ATR–Chk1 checkpoint pathway through regulation of Claspin.

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Abbreviations

ATM:

ataxia telangiectasia-mutated

ATR:

ATM and Rad3-related

DSB:

double-stranded DNA break

HU:

hydroxyurea

IR:

ionizing radiation

MCM:

minichromosomal maintenance

MMS:

methylmethane sulfate

RPA:

replication protein A

TopBP1:

topoisomerase II binding protein 1

TUNEL:

terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling

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Acknowledgements

We thank Chika Taniyama, Miyuki Kawashima and Ai Ishii for excellent technical assistance. We are grateful to Dr Akiko Kumagai for valuable comments on the manuscript and to the members of our laboratory for helpful discussion. This work was supported in part by grants-in-aid for scientific research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan to HM.

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Authors and Affiliations

  1. Genome Dynamics Project, The Tokyo Metropolitan Institute of Medical Science, Bunkyo-ku, Tokyo, Japan

    J M Kim, N Kakusho, M Yamada, Y Kanoh & H Masai

  2. Cytokine Project, The Tokyo Metropolitan Institute of Medical Science, Bunkyo-ku, Tokyo, Japan

    N Takemoto

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  1. J M Kim
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Correspondence to H Masai.

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

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Kim, J., Kakusho, N., Yamada, M. et al. Cdc7 kinase mediates Claspin phosphorylation in DNA replication checkpoint. Oncogene 27, 3475–3482 (2008). https://doi.org/10.1038/sj.onc.1210994

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