DNA-PK Is Involved in Repairing a Transient Surge of DNA Breaks Induced by Deceleration of DNA Replication

Cells that suffer substantial inhibition of DNA replication halt their cell cycle via a checkpoint response mediated by the PI3 kinases ATM and ATR. It is unclear how cells cope with milder replication insults, which are under the threshold for ATM and ATR activation. A third PI3 kinase, DNA-dependent protein kinase (DNA–PK), is also activated following replication inhibition, but the role DNA–PK might play in response to perturbed replication is unclear, since this kinase does not activate the signaling cascades involved in the S-phase checkpoint. Here we report that mild, transient drug-induced perturbation of DNA replication rapidly induced DNA breaks that promptly disappeared in cells that contained a functional DNA–PK whereas such breaks persisted in cells that were deficient in DNA–PK activity. After the initial transient burst of DNA breaks, cells with a functional DNA–PK did not halt replication and continued to synthesize DNA at a slow pace in the presence of replication inhibitors. In contrast, DNA–PK deficient cells subject to low levels of replication inhibition halted cell cycle progression via an ATR-mediated S-phase checkpoint. The ATM kinase was dispensable for the induction of the initial DNA breaks. These observations suggest that DNA-PK is involved in setting a high threshold for the ATR-Chk1-mediated S-phase checkpoint by promptly repairing DNA breaks that appear immediately following inhibition of DNA replication.