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Maintenance of replication forks and the S-phase checkpoint by Cdc18p and Orp1p

Nature Cell Biology volume 4, pages 384–388 (2002)Cite this article

Abstract

S-phase and DNA damage checkpoint controls block the onset of mitosis when DNA is damaged or DNA replication is incomplete1,2,3. It has been proposed that damaged or incompletely replicated DNA generates structures that are sensed by the checkpoint control pathway4,5, although little is known about the structures and mechanisms involved. Here, we show that the DNA replication initiation proteins Orp1p6,7 and Cdc18p8,9 are required to induce and maintain the S-phase checkpoint in Schizosaccharomyces pombe. The presence of DNA replication structures correlates with activation of the Cds1p checkpoint protein kinase10 and the S-phase checkpoint pathway. By contrast, induction of the DNA damage pathway is not dependent on Orp1p or Cdc18p. We propose that the presence of unresolved replication forks, together with Orp1p and Cdc18p, are necessary to activate the Cds1p-dependent S-phase checkpoint.

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Figure 1: The DNA replication checkpoint is neither activated nor maintained in the absence of Orp1p and Cdc18p.
Figure 2: Overexpression of Cds1p cannot block entry into mitosis when Orp1p and Cdc18p are inactivated.
Figure 3: The DNA damage checkpoint is maintained in the absence of Orp1p and Cdc18p.
Figure 4: 2D gel electrophoresis.

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Acknowledgements

We are grateful to H. Nishitani for antibodies. We thank all members of the cell cycle lab for their help and F. Uhlmann for critical reading of the manuscript. H.M. was supported by the Japan Society for the Promotion of Science and the Imperial Cancer Research Fund, and S.Y. by the National Sciences and Engineering Research Council of Canada, the British Council and the Association for International Cancer Research.

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Author notes

  1. Hiroshi Murakami and Stephanie K. Yanow: These authors have contributed equally to this work.

Authors and Affiliations

  1. Cell Cycle Laboratory, Cancer Research UK London Research Institute, Lincoln's Inn Fields Laboratories, 44 Lincoln's Inn Fields, London, WC2A 3PX, UK

    Hiroshi Murakami, Stephanie K. Yanow, Dominic Griffiths & Paul Nurse

  2. Department of Biochemistry, Nagoya City University Medical School, 1 Kawasumi, Mizuho-cho, Mizuho-ku, 467-8601, Nagoya, Japan

    Hiroshi Murakami & Makoto Nakanishi

  3. Division of Biology, 216-76, Howard Hughes Medical Institute, California Institute of Technology, Pasadena, 91125, CA, USA

    Stephanie K. Yanow

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  1. Hiroshi Murakami
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Correspondence to Hiroshi Murakami.

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Supplementary information

Figure 1

Cds1p kinase activity. (PDF 18 kb)

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Murakami, H., Yanow, S., Griffiths, D. et al. Maintenance of replication forks and the S-phase checkpoint by Cdc18p and Orp1p. Nat Cell Biol 4, 384–388 (2002). https://doi.org/10.1038/ncb789

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