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A Rad3–Rad26 complex responds to DNA damage independently of other checkpoint proteins

Nature Cell Biology volume 1, pages 393–398 (1999)Cite this article

Abstract

The conserved PIK-related kinase Rad3 is required for all DNA-integrity-checkpoint responses in fission yeast. Here we report a stable association between Rad3 and Rad26 in soluble protein extracts. Rad26 shows Rad3-dependent phosphorylation after DNA damage. Unlike phosphorylation of Hus1, Crb2/Rhp9, Cds1 and Chk1, phosphorylation of Rad26 does not require other known checkpoint proteins. Rad26 phosphorylation is the first biochemical marker of Rad3 function, indicating that Rad3-related checkpoint kinases may have a direct role in DNA-damage recognition.

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Figure 1: Rad26 is phosphorylated in response to DNA damage.
Figure 2: Rad26 and Rad3 define a protein complex.
Figure 3: HA–rad26 phenocopies rad26-T12 and is complexed with Rad3.
Figure 4: Genetic dependency of Rad26 phosphorylation.
Figure 5: S-phase-induced DNA damage is distinct from G2 DNA damage.

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References

  1. Carr, A. M. Control of cell cycle arrest by the Mec1sc/Rad3sp DNA structure checkpoint pathway. Curr. Opin. Genet.Dev. 7, 93–98 (1997).

    Article  CAS  Google Scholar 

  2. Elledge, S. J. Cell cycle checkpoints: preventing an identity crisis. Science 274, 1664–1672 ( 1996).

    Article  CAS  Google Scholar 

  3. Matsuoka, S., Huang, M. & Elledge, S. J. Linkage of ATM to cell cycle regulation by the Chk2 protein kinase. Science 282, 1893– 1897 (1998).

    Article  CAS  Google Scholar 

  4. Hoekstra, M. F. Responses to DNA damage and regulation of cell cycle checkpoints by the ATM protein kinase family. Curr. Opin. Genet. Dev 7, 170–175 (1997).

    Article  CAS  Google Scholar 

  5. Savitsky, K. et al. A single ataxia telangiectasia gene with a product similar to PI 3-kinase. Science 268, 1749– 1753 (1995).

    Article  CAS  Google Scholar 

  6. Wright, J. A. et al. Protein kinase mutants of human ATR increase sensitivity to UV and ionizing radiation and abrogate cell cycle checkpoint control. Proc. Natl Acad. Sci. USA 95, 7445– 7450 (1998).

    Article  CAS  Google Scholar 

  7. Cliby, W. A. et al. Overexpression of a kinase-inactive ATR protein causes sensitivity to DNA-damaging agents and defects in cell cycle checkpoints. EMBO J. 17, 159–169 ( 1998).

    Article  CAS  Google Scholar 

  8. Tibbetts, R. S. et al. A role for ATR in the DNA damage-induced phosphorylation of p53. Genes Dev. 13, 153– 157 (1999).

    Article  Google Scholar 

  9. Desany, B. A., Alcasabas, A. A., Bachant, J. B. & Elledge, S. J. Recovery from DNA replicational stress is the essential function of the S-phase checkpoint pathway. Genes Dev. 12, 2956– 2970 (1998).

    Article  CAS  Google Scholar 

  10. Zhao, X., Muller, E. G. & Rothstein, R. A suppressor of two essential checkpoint genes identifies a novel protein that negatively affects dNTP pools. Mol. Cell 2, 329–340 (1998).

    Article  CAS  Google Scholar 

  11. Weinert, T. A., Kiser, G. L. & Hartwell, L. H. Mitotic checkpoint genes in budding yeast and the dependence of mitosis on DNA replication and repair. Genes Dev. 8, 652–665 ( 1994).

    Article  CAS  Google Scholar 

  12. Greenwell, P. W. et al. TEL1, a gene involved in controlling telomere length in S. cerevisiae, is homologous to the human ataxia telangiectasia gene. Cell 82, 823–829 (1995).

    Article  CAS  Google Scholar 

  13. Morrow, D. M., Tagle, D. A., Shiloh, Y., Collins, F. S. & Hieter, P. TEL1, an S. cerevisiae homologue of the human gene mutated in ataxia telangiectasia, is functionally related to the yeast checkpoint gene MEC1. Cell 82, 831– 840 (1995).

    Article  CAS  Google Scholar 

  14. Bentley, N. J. et al. The Schizosaccharomyces pombe rad3 checkpoint gene . EMBO J. 15, 6641–6651 (1996).

    Article  CAS  Google Scholar 

  15. Naito, T., Matsuura, A. & Ishikawa, F. Circular chromosome formation in a fission yeast mutant defective in two ATM homologues. Nature Genet. 2, 203–206 (1998).

    Article  Google Scholar 

  16. Griffiths, D. J. F. & Carr, A. M. in DNA Repair in Prokaryates and Lower Eukaryotes Vol. 1 (eds Nickoloff, J. A. & Hoekstra, M. F.) 449–475 (Humana, Totowa, 1998).

    Google Scholar 

  17. Critchlow, S. E. & Jackson, S. P. DNA end-joining: from yeast to man. Trends Biochem. Sci. 23, 394–398 (1998).

    Article  CAS  Google Scholar 

  18. Lydall, D. & Weinert, T. Yeast checkpoint genes in DNA damage processing: implications for repair and arrest. Science 270, 1488–1491 (1995).

    Article  CAS  Google Scholar 

  19. de la Torre-Ruiz, M. A., Green, C. M. & Lowndes, N. F. RAD9 and RAD24 define two additive, interacting branches of the DNA damage checkpoint pathway in budding yeast normally required for Rad53 modification and activation. EMBO J. 17, 2687–2698 (1998).

    Article  CAS  Google Scholar 

  20. Murray, J. M., Lindsay, H. D., Munday, C. A. & Carr, A. M. Role of Schizosaccharomyces pombe RecQ homolog, recombination, and checkpoint genes in UV damage tolerance. Mol. Cell. Biol. 17, 6868–6875 (1997).

    Article  CAS  Google Scholar 

  21. Lindsay, H. D. et al. S-phase specific activation of Cds1 kinase defines a subpathway of the checkpoint response in Schizosaccharomyces pombe. Genes Dev. 12, 382–395 ( 1998).

    Article  CAS  Google Scholar 

  22. Martinho, R. G. et al. Analysis of Rad3 and Chk1 protein kinases defines different checkpoint responses. EMBO J. 17, 7239– 7249 (1998).

    Article  CAS  Google Scholar 

  23. Al-Khodairy, F. et al. Identification and characterisation of new elements involved in checkpoints and feedback controls in fission yeast. Mol. Biol. Cell 5, 147–160 ( 1994).

    Article  CAS  Google Scholar 

  24. Kostrub, C., Knudsen, K., Subramani, S. & Enoch, T. Hus1p, a conserved fission yeast checkpoint protein, interacts with Rad1p and is phosphorylated in response to DNA damage. EMBO J. 17, 2055–2066 (1998).

    Article  CAS  Google Scholar 

  25. Walworth, N. & Bernards, R. rad-dependent responses of the chk1-encoded protein kinase at the DNA damage checkpoint. Science 271, 353–356 ( 1996).

    Article  CAS  Google Scholar 

  26. Saka, Y., Esashi, F., Matsusaka, T., Mochida, S. & Yanagida, M. Damage and replication checkpoint control in fission yeast is ensured by interactions of Crb2, a protein with BRCT motif, with Cut5 and Chk1. Genes Dev. 11, 3387–3400 (1997).

    Article  CAS  Google Scholar 

  27. Lydall, D. & Weinert, T. G2/M checkpoint genes of Saccharomyces cerevisiae: further evidence for roles in DNA replication and/or repair . Mol. Gen. Genet. 256, 638– 651 (1997).

    Article  CAS  Google Scholar 

  28. Garvik, B., Carson, M. & Hartwell, L. Single-stranded DNA arising at telomeres in cdc13 mutants may constitute a specific signal for the RAD9 checkpoint . Mol. Cell. Biol. 15, 6128– 6138 (1995).

    Article  CAS  Google Scholar 

  29. Francesconi, S., Grenon, M., Bouvier, D. & Baldacci, G. p56chk1 protein kinase is required for the DNA replication checkpoint at 37 °C in fission yeast. EMBO J. 16, 1332–1341 (1997).

    Article  CAS  Google Scholar 

  30. Gutz, H., Heslot, H., Leupold, U. & Loprieno, N. in Handbook of Genetics Vol. 1 (ed. King, R. C.) 395–446 (Plenum, New York, 1974).

    Google Scholar 

  31. Craven, R. A. et al. Vectors for the expression of tagged proteins in Schizosaccharomyces pombe. Gene 221, 59– 68 (1998).

    Article  CAS  Google Scholar 

  32. Murray, J. M. et al. Cloning and characterisation of the S. pombe rad15 gene, a homologue to the S. cerevisiae RAD3 and human ERCC2 genes. Nucleic Acids Res. 20, 2673–2678 (1992).

    Article  CAS  Google Scholar 

  33. Bahler, J. et al. Heterologous modules for efficient and versatile PCR-based gene targeting in Schizosaccharomyces pombe. Yeast 14, 943–951 (1998).

    Article  CAS  Google Scholar 

  34. Santocanale, C. & Diffley, J. F. A Mec1- and Rad53-dependent checkpoint controls late-firing origins of DNA replication . Nature 395, 615–618 (1998).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank T. Caspari for discussions and the ICOS Corporation for financial assistance. This work was supported in part by Euratom contract F14PCT950010.

Correspondence and requests for materials should be addressed to A.M.C.

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  1. MRC Cell Mutation Unit, Sussex University, Falmer, BN1 9RR, UK

    Rhian J. Edwards, Nicola J. Bentley & Antony M. Carr

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  3. Antony M. Carr
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Correspondence to Antony M. Carr.

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Edwards, R., Bentley, N. & Carr, A. A Rad3–Rad26 complex responds to DNA damage independently of other checkpoint proteins. Nat Cell Biol 1, 393–398 (1999). https://doi.org/10.1038/15623

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