CDC7 and APC/C^Cdh1 gate distinct routes to initiate DNA replication

Precise temporal control of DNA replication initiation is essential for faithful cell division, yet there are conflicting prevailing models for how cells trigger origin firing. Here, we demonstrate that human origins are not fired at fixed thresholds of cell cycle regulators E2F, APC/C^Cdh1, CDK2/1, and CDC7. Instead, origin firing is triggered at a tunable CDK2/1 threshold that is gated by either CDC7 or APC/C^Cdh1 activity. CDC7 phosphorylates MCM helicase, enabling Cyclin E-CDK2 to trigger origin firing at low CDK2/1 activity, independent of APC/C^Cdh1 inactivation. In contrast, APC/C^Cdh1 inactivation enables Cyclin A-CDK1 to phosphorylate MCM at distinct CDK2/1-specific sites and trigger origin firing, independent of CDC7 activity. Strikingly, this requires much higher CDK2/1 activity, which CDK2 cannot normally reach alone. Thus, having two distinct routes necessitates blocking both routes to prevent S phase: either by inhibiting CDC7 or CDK2 while also preventing APC/C^Cdh1 inactivation or inhibiting CDK1.