CMG Pol epsilon dynamics suggests a mechanism for the establishment of leading strand synthesis in the eukaryotic replisome
Significance Faithful and efficient genome duplication is essential for the propagation of life. Aberrant DNA replication can lead to genomic instability and cancer. In eukaryotes, the replication machinery is composed of the DNA unwinding enzyme Cdc45 MCM GINS (CMG) and dedicated DNA synthesis factors. Three different polymerases act sequentially on the leading strand template to establish DNA replication. We describe the architecture and dynamics of the main leading strand polymerase bound to the CMG helicase, and we propose a mechanism for the establishment of efficient leading strand synthesis. Our findings provide important insights into how the eukaryotic replication machinery functions to ensure that genome integrity is maintained during replication. The replisome unwinds and synthesizes DNA for genome duplication. In eukaryotes, the Cdc45 MCM GINS (CMG) helicase and the leading strand polymerase, Pol epsilon, form a stable assembly. The mechanism for coupling DNA unwinding with synthesis is starting to be elucidated, however the architecture and dynamics of the replication fork remain only partially understood, preventing a molecular understanding of chromosome replication. To address this issue, we conducted a systematic single particle EM study on multiple permutations of the reconstituted CMG Pol epsilon assembly. Pol epsilon contains two flexibly tethered lobes. The noncatalytic lobe is anchored to the motor of the helicase, whereas the polymerization domain extends toward the side of the helicase. We observe two alternate configurations of the DNA synthesis domain in the CMG bound Pol epsilon. We propose that this conformational switch might control DNA template engagement and release, modulating replisome progression.
Proceedings of the National Academy of Sciences