Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

Fu, Yu; Yardimci, Hasan; Long, David T.; Guainazzi, Angelo; Bermudez, Vladimir P.; Hurwitz, Jerard; Oijen, Antoine M. van; Schärer, Orlando D.; Walter, Johannes C.

doi:10.1016/j.cell.2011.07.045

Cited by 326 publications

(387 citation statements)

References 51 publications

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“…Unlike the prokaryotic DnaB replicative helicase, which translocates on the lagging strand, 40 the eukaryotic CMG helicase complex translocates on the leading strand. 41 Since the critical Ter C6 residue is on the leading strand of the fork approaching the non-permissive end of Ter, this residue could be masked in the barrel of the MCM helicase, thereby preventing C6 lock formation. However, this hypothesis fails to explain the behavior of Tus/Ter in S. cerevisiae, where the C6 lock mechanism is a significant contributor to replisome arrest, 21 despite the fact that the replicative helicase translocates along the leading strand.…”

Section: Discussionmentioning

confidence: 99%

Spatial separation of replisome arrest sites influences homologous recombination quality at a Tus/Ter-mediated replication fork barrier

Willis

Scully

2016

Cell Cycle

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The Escherichia coli replication fork arrest complex Tus/Ter mediates site-specific replication fork arrest and homologous recombination (HR) on a mammalian chromosome, inducing both conservative "short tract" gene conversion (STGC) and error-prone "long tract" gene conversion (LTGC) products. We showed previously that bidirectional fork arrest is required for the generation of STGC products at Tus/Ter-stalled replication forks and that the HR mediators BRCA1, BRCA2 and Rad51 mediate STGC but suppress LTGC at Tus/Ter-arrested forks. Here, we report the impact of Ter array length on Tus/Ter-induced HR, comparing HR reporters containing arrays of 6, 9, 15 or 21 Ter sites-each targeted to the ROSA26 locus of mouse embryonic stem (ES) cells. Increasing Ter copy number within the array beyond 6 did not affect the magnitude of Tus/Ter-induced HR but biased HR in favor of LTGC. A "lock"-defective Tus mutant, F140A, known to exhibit higher affinity than wild type (wt)Tus for duplex Ter, reproduced these effects. In contrast, increasing Ter copy number within the array reduced HR induced by the I-SceI homing endonuclease, but produced no consistent bias toward LTGC. Thus, the mechanisms governing HR at Tus/Ter-arrested replication forks are distinct from those governing HR at an enzyme-induced chromosomal double strand break (DSB). We propose that increased spatial separation of the 2 arrested forks encountering an extended Tus/Ter barrier impairs the coordination of DNA ends generated by the processing of the stalled forks, thereby favoring aberrant LTGC over conservative STGC.

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Section: Discussionmentioning

confidence: 99%

Spatial separation of replisome arrest sites influences homologous recombination quality at a Tus/Ter-mediated replication fork barrier

Willis

Scully

2016

Cell Cycle

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“…This mechanism requires the convergence of two replication forks at an ICL during DNA replication (Zhang et al , 2015). Both forks initially stall 20–40 nucleotides from the crosslink followed by CMG helicase unloading allowing one fork to approach to within 1 nucleotide of the crosslink (Räschle et al , 2008; Fu et al , 2011; Long et al , 2011). Dual incisions on either side of the ICL then unhook the lesion from one of the strands.…”

Section: Introductionmentioning

confidence: 99%

Recruitment and positioning determine the specific role of the XPF ‐ ERCC 1 endonuclease in interstrand crosslink repair

et al. 2017

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XPF‐ERCC1 is a structure‐specific endonuclease pivotal for several DNA repair pathways and, when mutated, can cause multiple diseases. Although the disease‐specific mutations are thought to affect different DNA repair pathways, the molecular basis for this is unknown. Here we examine the function of XPF‐ERCC1 in DNA interstrand crosslink (ICL) repair. We used Xenopus egg extracts to measure both ICL and nucleotide excision repair, and we identified mutations that are specifically defective in ICL repair. One of these separation‐of‐function mutations resides in the helicase‐like domain of XPF and disrupts binding to SLX4 and recruitment to the ICL. A small deletion in the same domain supports recruitment of XPF to the ICL, but inhibited the unhooking incisions most likely by disrupting a second, transient interaction with SLX4. Finally, mutation of residues in the nuclease domain did not affect localization of XPF‐ERCC1 to the ICL but did prevent incisions on the ICL substrate. Our data support a model in which the ICL repair‐specific function of XPF‐ERCC1 is dependent on recruitment, positioning and substrate recognition.

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“…The model proposes that the initial incisions of ICLs, a process termed “ICL unhooking”, require the convergence of two replication forks upon the ICL. Both nascent leading strands initially stall ~20–40 nt from the ICL (“−20” position) due to steric hindrance imposed by the CMG helicase complexes (Fu et al , 2011). BRCA1‐dependent eviction of the CMG helicase at the replication forks enables the subsequent extension of one nascent leading strand to immediately adjacent to the crosslinked nucleotides (“approach” to “−1” position; Long et al , 2014).…”

Section: Introductionmentioning

confidence: 99%

RPA activates the XPF ‐ ERCC 1 endonuclease to initiate processing of DNA interstrand crosslinks

et al. 2017

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During replication‐coupled DNA interstrand crosslink (ICL) repair, the XPF‐ERCC1 endonuclease is required for the incisions that release, or “unhook”, ICLs, but the mechanism of ICL unhooking remains largely unknown. Incisions are triggered when the nascent leading strand of a replication fork strikes the ICL. Here, we report that while purified XPF‐ERCC1 incises simple ICL‐containing model replication fork structures, the presence of a nascent leading strand, modelling the effects of replication arrest, inhibits this activity. Strikingly, the addition of the single‐stranded DNA (ssDNA)‐binding replication protein A (RPA) selectively restores XPF‐ERCC1 endonuclease activity on this structure. The 5′–3′ exonuclease SNM1A can load from the XPF‐ERCC1‐RPA‐induced incisions and digest past the crosslink to quantitatively complete the unhooking reaction. We postulate that these collaborative activities of XPF‐ERCC1, RPA and SNM1A might explain how ICL unhooking is achieved in vivo.

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Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

Cited by 326 publications

References 51 publications

Spatial separation of replisome arrest sites influences homologous recombination quality at a Tus/Ter-mediated replication fork barrier

Spatial separation of replisome arrest sites influences homologous recombination quality at a Tus/Ter-mediated replication fork barrier

Recruitment and positioning determine the specific role of the XPF ‐ ERCC 1 endonuclease in interstrand crosslink repair

RPA activates the XPF ‐ ERCC 1 endonuclease to initiate processing of DNA interstrand crosslinks

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