title: CMG-Pol epsilon dynamics suggests a mechanism for the establishment of leading-strand synthesis in the eukaryotic replisome.
abstract: 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.
authors.affiliation: Macromolecular Machines Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: Jin Chuan
authors.identifier: 0000-0001-5716-5170
authors.initials: JC
authors.lastname: Zhou
authors.affiliation: Chromosome Replication Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Janska
authors.affiliation: Macromolecular Machines Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: Panchali
authors.initials: P
authors.lastname: Goswami
authors.affiliation: Macromolecular Machines Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: Ludovic
authors.initials: L
authors.lastname: Renault
authors.affiliation: Macromolecular Machines Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: Ferdos
authors.initials: F
authors.lastname: Abid Ali
authors.affiliation: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, OX3 7BN, United Kingdom.
authors.forename: Abhay
authors.initials: A
authors.lastname: Kotecha
authors.affiliation: Chromosome Replication Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom.
authors.forename: John F X
authors.identifier: 0000-0001-5184-7680
authors.initials: JFX
authors.lastname: Diffley
authors.affiliation: Macromolecular Machines Laboratory, The Francis Crick Institute, London, NW1 1AT, United Kingdom; alessandro.costa@crick.ac.uk.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Costa
mesh_terms: D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004257:DNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D005057:Eukaryotic Cells / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D004257:DNA Polymerase II; D004265:DNA Helicases
keywords: CMG helicase; DNA polymerase; DNA replication; single-particle electron microscopy