The Eukaryotic Replisome Goes Under the Microscope

Abstract

The machinery at the eukaryotic replication fork has seen many new structural advances using electron microscopy and crystallography. Recent structures of eukaryotic replisome components include the Mcm2-7 complex, the CMG helicase, DNA polymerases, a Ctf4 trimer hub and the first look at a core replisome of 20 different proteins containing the helicase, primase, leading polymerase and a lagging strand polymerase. The eukaryotic core replisome shows an unanticipated architecture, with one polymerase sitting above the helicase and the other below. Additionally, structures of Mcm2 bound to an H3/H4 tetramer suggest a direct role of the replisome in handling nucleosomes, which are important to DNA organization and gene regulation. This review provides a summary of some of the many recent advances in the structure of the eukaryotic replisome.

Structure of eukaryotic CMG helicase at a replication fork and implications to replisome architecture and origin initiation

“…S15C). We noted earlier that the catalytic NTD of Pol2 may be mobile and averaged out in the CMG-Pol e 3D reconstruction, thus explaining the noted loss of about 30% of the expected volume of Pol e in CMG-Pol e, as suggested previously (17,26,27).…”

“…Pol e is positioned on the C-tier, below the helicase to immediately extend the leading-strand 3′ terminus upon exit from CMG. The Pol e catalytic domain is in the NTD of Pol2 which has been proposed to be attached to the CTD of Pol e by a flexible tether (see text for details) (17,26,27). This may allow the catalytic Pol e NTD to transiently release the leading strand while remaining attached to CMG, exposing the leading 3′ terminus to RFC (or other proteins) for loading additional PCNA needed for Caf1-mediated nucleosome assembly and mismatch repair of the leading strand.…”

Regulation and Modulation of Human DNA Polymerase δ Activity and Function

“…ChIP analysis with antibody against the p125 subunit showed that PDIP46 was present together with two components of the mammalian replisome [56]. These are Mcm2, a component of the CMG (Cdc45-MCM-GINS) helicase [151], and Ctf4, which associates with CMG [152,153]. Thus, the ChIP data supports the idea that PDIP46 is associated with chromatin at or near the replisome.…”

From structure to mechanism—understanding initiation of DNA replication

“…DNA polymerases are known to associate with the CMG in part via Ctf4 to form a coupled DNA-unwinding and DNA synthesis assembly ( Fig. 1G; Gambus et al 2006;Simon et al 2014;O'Donnell and Li 2016). Initial priming of DNA synthesis is carried out by DNA polymerase α, while leading and lagging strand DNA synthesis occurs mainly via polymerase ε (Pursell et al 2007;Burgers et al 2016) and polymerase δ (Nick McElhinny et al 2008), respectively.…”

“…Initiation of DNA replication is a multistep reaction that is carefully choreographed to promote replication fork assembly and regulated firing of replication origins (Costa et al 2013;Riera et al 2014;Tognetti et al 2015;Bell and Labib 2016;O'Donnell and Li 2016;Pellegrini and Costa 2016;Riera and Speck 2016;Bleichert et al 2017). Moreover, this process is highly regulated in order to coordinate DNA synthesis with the cell cycle and the energy status of the cell.…”