Stochasticity Triggers Activation of the S-phase Checkpoint Pathway in Budding Yeast

In a complex and ever-changing environment, various signal transduction pathways mediate outside signals and stress to a living cell and its intracellular responses. Eukaryotic cells utilize the DNA synthesis phase (S-phase) checkpoint to respond to DNA damage and replication stress, and the activation of the S-phase checkpoint defers the routine progression in the S phase. Through the analysis of microfluidic single-cell measurements, we find that the behavior of yeast cells exhibits bimodal distribution in the activation of the S-phase checkpoint, and the nonactivated portion of cells obeys the exponential decay law over time, the rate of which is dictated by HU dosage. Mathematical modeling and further experimental evidence from different mutant strains support the idea that the activation of the yeast S-phase checkpoint is a stochastic barrier-crossing process in a double-well system, where the barrier height is determined by both DNA replication stress and autophosphorylation of the key effector kinase Rad53. Our approach, as a novel methodology, is generally applicable to quantitative analysis of the signal transduction pathways at the single-cell level.