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A more recent version of this article appeared on February 1, 2002
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Submitted on March 9, 2001
Revised on October 24, 2001
Accepted on November 1, 2001
1 Department of Genetics, Harvard Medical School, 200 Longwood Avenue, Boston, MA 02115
* Corresponding author. E-mail address: wolkow{at}rascal.med.harvard.
Fission yeast Rad3 is a member of a family of PI-3 kinase related kinases required for the maintenance of genomic stability in all eukaryotic cells. In fission yeast, Rad3 regulates the cell cycle arrest and recovery activities associated with the G2/M checkpoint. We have developed an assay that directly measures Rad3 kinase activity in cells expressing physiological levels of the protein. Using the assay, we demonstrate directly that Rad3 kinase activity is stimulated by checkpoint signals. Of the five other G2/M checkpoint proteins (Hus1, Rad1, Rad9, Rad17 and Rad26), only Rad26 was required for Rad3 kinase activity. As Rad26 has previously been shown to interact constitutively with Rad3, our results demonstrate that Rad26 is a regulatory subunit, and Rad3 is the catalytic subunit, of the Rad3/Rad26 kinase complex. Analysis of Rad3/Rad26 kinase activation in rad26.T12, a mutant that is proficient for cell cycle arrest, but defective in recovery, suggests that these two responses to checkpoint signals require quantitatively different levels of kinase activity from the Rad3/Rad26 complex.
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