Replication stress and resulting genome instability, a major driver of cancer progression, stem from perturbations of replication fork progression. The first defense against this stress is activation of "dormant" replication origins, which supports replication completion. To determine whether ATR, in itself, contributes to this compensation process, we submitted human cells to a range of low stresses sufficient to activate ATR, not CHK1. Using molecular combing, we developed a dose-response assay that quantifies compensation efficiencies, enabling accurate comparison of cells with different genotypes. Combined with Repli-seq and OK-seq, this assay revealed that ATR activation is key to compensation triggering. We next asked how TopBP1, the main ATR activator, impacts compensation. In stark contradiction to what would be expected from its checkpoint function, we found that TopBP1 represses compensation and acts downstream of ATR. Instead, the function of TopBP1 in replisome assembly, which remains unclear in mammalian cells where the protein is not essential, well-accounts for our results positing that TopBP1 locks dormant origins at the pre-initiation stage, an intermediate in the assembly process, and that ATR activation allows assembly to resume. TopBP1 engagement in the pre-initiation complex would thus serve as a switch linking replisome assembly to the stress response.
Koundrioukoff et al. (Sat,) studied this question.