Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 114, Issue 4, Pages 675-680Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1619748114
Keywords
replisome; replication; clamp loader; PCNA; DNA polymerase
Categories
Funding
- National Institutes of Health [T32 CA009673, R01 GM115809]
- Howard Hughes Medical Institute
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The eukaryotic genome is primarily replicated by two DNA polymerases, Pol epsilon and Pol delta, that function on the leading and lagging strands, respectively. Previous studies have established recruitment mechanisms whereby Cdc45-Mcm2-7-GINS (CMG) helicase binds Pol e and tethers it to the leading strand, and PCNA (proliferating cell nuclear antigen) binds tightly to Pol delta and recruits it to the lagging strand. The current report identifies quality control mechanisms that exclude the improper polymerase from a particular strand. We find that the replication factor C (RFC) clamp loader specifically inhibits Pol epsilon on the lagging strand, and CMG protects Pol epsilon against RFC inhibition on the leading strand. Previous studies show that Pol delta is slow and distributive with CMG on the leading strand. However, Saccharomyces cerevisiae Pol delta-PCNA is a rapid and processive enzyme, suggesting that CMG may bind and alter Pol delta activity or position it on the lagging strand. Measurements of polymerase binding to CMG demonstrate Pol epsilon binds CMG with a K-d value of 12 nM, but Pol delta binding CMG is undetectable. Pol delta, like bacterial replicases, undergoes collision release upon completing replication, and we propose Pol delta-PCNA collides with the slower CMG, and in the absence of a stabilizing Pol delta-CMG interaction, the collision release process is triggered, ejecting Pol delta on the leading strand. Hence, by eviction of incorrect polymerases at the fork, the clamp machinery directs quality control on the lagging strand and CMG enforces quality control on the leading strand.
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