A Meier–Gorlin syndrome mutation impairs the loading of the MCM2–7 complex during DNA replication initiation

Chromatin loading of the hexameric replicative helicase MCM2-7 complex requires coordinated interactions with the origin recognition complex (ORC), CDC6, and CDT1. MCM2-7 not bound to DNA forms a single hexamer (SH) with an open DNA entry gate between MCM2 and MCM5. Two MCM2-7 SHs can be loaded sequentially to form the double hexamer (DH) that encircles the DNA duplex. Activated MCM2-7 then unwinds DNA and initiates DNA replication. Our cryoelectron microscopy analyses show that a fraction of human MCM2-7 without DNA exists as DH. Unexpectedly, we find that the MCM3 winged helix domain (WHD) docks on MCM2 in both DNA-free DH and SH, creating a safety latch across the DNA entry gate to block DNA entry into the central channel. The safety latch can be opened by ORC-CDC6 binding. Perturbing this latch by structure-based or disease-related mutations of MCM3 causes replication defects and DNA damage checkpoint activation. Shortening the MCM3 linker between the helicase domain and WHD alleviates the cell cycle defects of the latch-strengthening mutation. Our findings uncover a regulated step in MCM2-7 loading with implications for human diseases.