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Molecular and General Genetics MGG

, Volume 255, Issue 3, pp 332–340 | Cite as

Characterisation of the Schizosaccharomyces pombe rad4/cut5 mutant phenotypes: dissection of DNA replication and G2 checkpoint control function

  • R. J. McFarlane
  • A. M. Carr
  • C. Price
ORIGINAL PAPER

Abstract

Mutation of the essential Schizosaccharomyces pombe rad4/cut5 gene causes sensitivity to UV and ionising radiation at the permissive temperature whilst at the restrictive temperature cells fail to undergo DNA replication but still attempt mitosis owing to a defective S-phase checkpoint response. Many mutations in genes encoding DNA replication proteins also abolish checkpoint responses, possibly because the replication machinery is a pre-requisite for the generation of the signal. We demonstrate here that rad4/cut5 cells fail to arrest cell division when treated with the replication inhibitor hydroxyurea at the semi-permissive temperature 32° C, but retain essentially normal replicative capacity. This demonstrates that the replication and checkpoint function of the rad4/cut5 gene product can be separated and that the Rad4 protein differs from other replication proteins in being directly involved in generating the S-phase checkpoint signal. Furthermore, we have investigated the checkpoint response or rad4/cut5-deficient cells to γ-irradiation and UV-mimetic drugs. We find that, at the restrictive temperature, the rad4 /cut5 cells fail to delay mitosis in response to γ-irradiation whilst retaining a normal checkpoint response to the UV-mimetic drug 4-nitroquinoline-1-oxide. The lack of the γ-irradiation checkpoint is reminiscent of the deficiency associated with mutation of the human ATM locus, the causative deficiency of the heritable disorder ataxia telangiectasia. The implications of our results for the organisation of distinct checkpoint-response pathways in both fission yeast and mammalian cells are discussed. Moreover the data are consistent with a model in which the generation of the S-Phase checkpoint signal is DNA polymerase ɛ dependent.

Key wordsSchizosaccharomyces pombe Checkpoint control γ-Irradiation DNA replication 

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Copyright information

© Springer-Verlag Berlin Heidelberg 1997

Authors and Affiliations

  • R. J. McFarlane
    • 1
  • A. M. Carr
    • 2
  • C. Price
    • 1
  1. 1.Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, P. O. Box 594, Western Bank Sheffield S10 2UH, UK e-mail clive.price@sheffield.ac.ukGB
  2. 2.MRC Cell Mutation Unit, University of Sussex, Brighton BN1 9RR, UKGB

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