Nutrient limitation inactivates mrc1-to-cds1 checkpoint signalling in schizosaccharomyces pombe

Abstract

The S. pombe checkpoint kinase, Cds1, protects the integrity of stalled DNA replication forks after its phosphorylation at threonine-11 by Rad3 (ATR). Modified Cds1 associates through its N-terminal forkhead-associated domain (FHA)-domain with Mrc1 (Claspin) at stalled forks. We report here that nutrient starvation results in post-translational changes to Cds1 and the loss of Mrc1. A drop in glucose after a down-shift from 3% to 0.1–0.3%, or when cells enter the stationary phase, triggers a sharp decline in Mrc1 and the accumulation of insoluble Cds1. Before this transition, Cds1 is transiently activated and phosphorylated by Rad3 when glucose levels fall. Because this coincides with the phosphorylation of histone 2AX at S129 by Rad3, an event that occurs towards the end of every unperturbed S phase, we suggest that a glucose limitation promotes the exit from the S phase. Since nitrogen starvation also depletes Mrc1 while Cds1 is post-translationally modified, we suggest that nutrient limitation is the general signal that promotes exit from S phase before it inactivates the Mrc1–Cds1 signalling component. Why Cds1 accumulates in resting cells while its activator Mrc1 declines is, as yet, unclear but suggests a novel function of Cds1 in non-replicating cells.