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JCS ePress
online publication date 15 Jul 2003
doi: 10.1242/jcs.00677
Research Article
Delineating the position of rad4+/cut5+ within the DNA-structure checkpoint pathways in Schizosaccharomyces pombe
Sheila Harris,
Caroline Kemplen,
Thomas Caspari,
Christopher Chan,
Howard D. Lindsay,
Marius Poitelea,
Antony M. Carr,
and
Clive Price*
* Author for correspondence (e-mail: c.price1{at}lancaster.ac.uk)
The fission yeast BRCT domain protein Rad4/Cut5 is required for genome integrity checkpoint responses and DNA replication. Here we address the position at which Rad4/Cut5 acts within the checkpoint response pathways. Rad4 is shown to act upstream of the effector kinases Chk1 and Cds1, as both Chk1 phosphorylation and Cds1 kinase activity require functional Rad4. Phosphorylation of Rad9, Rad26 and Hus1 in response to either DNA damage or inhibition of DNA replication are independent of Rad4/Cut5 checkpoint function. Further we show that a novel, epitope-tagged allele of rad4+/cut5+ acts as a dominant suppressor of the checkpoint deficiencies of rad3-, rad26- and rad17- mutants. Suppression results in the restoration of mitotic arrest and is dependent upon the remaining checkpoint Rad proteins and the two effector kinases. High-level expression of the rad4+/cut5+ allele in rad17 mutant cells restores the nuclear localization of Rad9, but this does not fully account for the observed suppression. We conclude from these data that Rad4/Cut5 acts with Rad3, Rad26 and Rad17 to effect the checkpoint response, and a model for its function is discussed.
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