DNA damage checkpoint maintenance through sustained Chk1 activity.
The G2 DNA damage checkpoint prevents mitotic entry in the presence of DNA damage. This requires the activation of the phosphoinositide-3-kinase-related protein kinases ATR and ATM in human cells and the ATR homologue Rad3 in the fission yeast Schizosaccharomyces pombe. Rad3 activates the effector protein kinase Chk1 by phosphorylation. However, in fission yeast, inactivation of Rad3 following checkpoint activation has no impact on checkpoint duration. This demonstrates that Rad3 is not required for checkpoint maintenance and that the processes of checkpoint initiation and maintenance are distinct. Chk1 is required for checkpoint initiation but its role in checkpoint maintenance has not been investigated. We show here that Chk1 kinase activity is rapidly induced following irradiation and is maintained for the duration of a checkpoint arrest. On entry to mitosis, there is a transient decrease in Chk1 activity and phosphorylation, but Chk1 activity remains higher than that observed in unirradiated cells. We have generated temperature-sensitive alleles of chk1, which phenocopy chk1 deletion at the non-permissive temperature. Using these alleles, we have shown that inactivation of Chk1 during a checkpoint arrest leads to premature checkpoint termination, resulting in catastrophic mitoses that are a hallmark of checkpoint failure. Therefore, unlike Rad3, Chk1 is an important determinant of both checkpoint initiation and maintenance.
Latif C, den Elzen NR, O'Connell MJ