Suppressors of Cdc25p overexpression identify two pathways that influence the G2/M checkpoint in fission yeast

Checkpoints maintain the order of cell-cycle events. At G2/M, a checkpoint blocks mitosis in response to damaged or unreplicated DNA. There are signif icant differences in the checkpoint responses to damaged DNA. and unreplica ted DNA, although many of the same genes are involved in both responses. To identify new genes that function specifically in the DNA replication check point pathway, we searched for high-copy suppressors of overproducer of Cdc 25p (OPcdc25(+)), which lacks a DNA replication checkpoint. Two classes of suppressors were isolated. One class includes a new gene encoding a putativ e DEAD box helicase, suppressor of uncontrolled mitosis (sum3(+)). This gen e negatively regulates the cell-cycle response to stress when overexpressed and restores the checkpoint response by a mechanism that is independent of Cdc2p tyrosine phosphorylation. The second class includes chk1(+) and the two Schizosaccharomyces pombe 14-3-3 genes, rad24(+) and rad25(+), which ap pear to suppress the checkpoint defect by inhibiting Cdc25p. We show that r ad24 Delta mutants are defective in the checkpoint response to the DNA repl ication inhibitor hydroxyurea at 37 degrees and that cds1 Delta rad24 Delta mutants, like cds1 Delta chk1 Delta mutants, are entirely checkpoint defic ient at 29 degrees. These results suggest that chk1(+) and rad24(+) may fun ction redundantly with cds1(+) in the checkpoint response to unreplicated D NA.