Inhibiting a checkpoint (bottom) allows both early (green) and late (red) replication origins to fire.

When DNA replication goes awry, it is critical for a cell to detect and correct the error as soon as possible. Thus, when replication from an early-firing origin is blocked, an intra-S-phase checkpoint prevents the initiation of replication from late-firing origins. The molecular machinery of this checkpoint has remained poorly understood. Now, Feijoo et al. (page 913) describe the activities of the kinases Chk1 and Chk2 in mammalian cells during replication arrest, and demonstrate that Chk1 is required for this intra-S-phase checkpoint.

Arresting DNA replication in HeLa cells with hydroxyurea causes both Chk1 and Chk2 to be phosphorylated and activated, whereas neither protein is activated during normal S phase progression, suggesting that the proteins are phosphorylated in response to stalled or slowed replication. When the replication block is released, Chk1 is rapidly inactivated, and DNA synthesis resumes, but Chk2 remains activated. In arrested cells, blocking Chk1 activation causes destabilization of stalled replication forks and allows late replication origins to fire, indicating that Chk1 is required for this intra- S-phase checkpoint.

The results suggest that stalled replication forks in S phase cause the phosphorylation and activation of both Chk1 and Chk2, and that Chk1 is primarily responsible for blocking the activation of late replication origins. Since DNA synthesis can proceed in the presence of activated Chk2, it is possible that Chk2 controls a later mitotic checkpoint. ▪