Termination of DNA replication forks: "Breaking up is hard to do"

Abstract

To ensure duplication of the entire genome, eukaryotic DNA replication initiates from thousands of replication origins. The replication forks move through the chromatin until they encounter forks from neighboring origins. During replication fork termination forks converge, the replisomes disassemble and topoisomerase II resolves the daughter DNA molecules. If not resolved efficiently, terminating forks result in genomic instability through the formation of pathogenic structures. Our recent findings shed light onto the mechanism of replisome disassembly upon replication fork termination. We have shown that termination-specific polyubiquitylation of the replicative helicase component - Mcm7, leads to dissolution of the active helicase in a process dependent on the p97/VCP/Cdc48 segregase. The inhibition of terminating helicase disassembly resulted in a replication termination defect. In this extended view we present hypothetical models of replication fork termination and discuss remaining and emerging questions in the DNA replication termination field.

Keywords: CMG, Cdc45, Mcm2–7, GINS complex; CRL, cullin-RING ligase; D loop, displacement loop; DDR, DNA damage response; DNA replication; DSB, double strand break; DUB, deubiquitylating enzyme; ER, endoplasmic reticulum; ERAD, endoplasmic reticulum associated protein degradation; GINS, Go-Ichi-Ni-San, complex made of Sld5, Psf1, Psf2, Psf3; ICL, intra-strand crosslink; MCM, Minichromosome maintenance; Mcm2–7; OriC, chromosomal replication origin; R loop, RNA:DNA hybrid; RING, really interesting gene; RPC, Replisome Progression Complex; Ter, termination site; Tus-Ter, terminus utilisation substance - termination; Xenopus; p97 segregase; replication termination; replicative helicase; replisome; ubiquitin.

Figure 1.

A speculative model of replisome dissolution at the termination of DNA replication forks based on data published in (5, 55). (A) Two replication forks from neighboring origins approach each other. (B) The Mcm7 subunit of the CMG complex becomes ubiquitylated with lysine-48-linked ubiquitin chains in a process dependent on cullin-type ubiquitin ligase. The ubiquitylated Mcm7 is recognized by protein segregase p97/VCP/Cdc48. (C) p97 activity is required to remodel the active helicase complex resulting in replisome disassembly and removal from chromatin.

Figure 2.

Two possible mechanisms for dormant origin removal from chromatin. Inactive Mcm2–7 complexes may be removed as active forks approach them (left panel, Elongation removal) or pushed in front of the progressing forks and removed at sites of replication fork termination (right panel, Termination removal).

Figure 3.

Speculative models of replication fork termination mechanisms. Two forks approach each other traveling in opposite directions with the CMG complexes unwinding DNA at the tips of the forks. Topo II helps to release the tension created by hemicatenates between approaching forks. (A) Simultaneous removal of converging replisomes. Two helicases collide with each other leading to disassembly of both replisomes and resolution of the terminating DNA with the potential help of additional factors and Topo II. (B) Sequential removal of converging replisomes. The convergence of two helicases leads to disassembly of one of the replisomes while the other finishes unwinding and replicating the remaining DNA. The sister chromatids are detangled by Topo II. (C) Passing replisomes. Two approaching forks pass each other at the termination site as each helicase is traveling only on the leading strand i.e., on the parental strand within the new sister chromatids. The CMG complex may be disassembled when it approaches the last Okazaki fragment of the lagging strand in front of it, start unwinding the last Okazaki fragment or start sliding on the double stranded DNA of the last Okazaki fragment. Again, Topo II detangles the new sister chromatids.