Problems that arise during DNA replication are thought to be the root cause of much of the genome instability, including duplications, deletions and translocations of chromosomal segments, that is associated with cancer and other disease states (Aguilera and García-Muse, 2013; Arlt et al., 2012; Macheret and Halazonetis, 2015). One common problem that arises is accidental collision of the replication fork with proteins that are tightly bound to the DNA (Lambert and Carr, 2013). Such collisions can precipitate a process known as replication fork collapse in which the protein components of the fork (collectively known as the replisome) are thought to partially or completely dissociate from the DNA (Cortez, 2015). In some cases this process may also be associated with DNA breakage, detaching one arm of the replication fork and exposing a single duplex DNA end. Replication fork collapse, either with or without DNA breakage, causes a localized cessation of DNA synthesis that can be remedied through the intervention of homologous recombination (HR) proteins, which restart replication.

In eukaryotes, recombination-dependent replication (RDR) has been studied mostly in the budding yeast Saccharomyces cerevisiae, using experimental systems in which RDR is initiated from a site-specific DNA double-strand break (DSB) (Anand et al., 2013; Kramara et al., 2018; Sakofsky and Malkova, 2017). HR uses intact homologous DNA sequences as templates to repair DSBs and, if such sequences are available for only one side of the break, repair occurs via a RDR process termed break induced replication (BIR). In most cases, BIR has been studied outside of the context of S-phase, with a DSB, rather than a collapsed or broken replication fork, acting as its inducer. However, studies, using a site-specific single-strand DNA break to induce replication fork breakage, have shown that essentially the same BIR process can repair broken forks during S-phase (Mayle et al., 2015).

Similar to other HR-mediated repair processes, BIR involves the 5’ to 3’ nucleolytic resection of a duplex DNA end to provide a single-stranded DNA (ssDNA) tail for the Rad51 recombination protein to load on to (Anand et al., 2013). In yeast, this loading reaction depends on Rad52, which acts as a mediator between ssDNA coated with the ssDNA-binding protein RPA and Rad51 (Krogh and Symington, 2004). Once loaded, Rad51 performs a DNA homology search, followed by strand invasion of the intact DNA sequence that it finds, forming a displacement (D) loop. The D-loop acts as a platform for replication protein reassembly, which seemingly involves most of the main replisome components (Lydeard et al., 2010). However, BIR exhibits a number of features that distinguishes it from canonical replication initiated from a replication origin, including: a requirement for the non-essential DNA Polymerase ∂ subunit Pol32 (Lydeard et al., 2007); a dependence on the Pif1 DNA helicase to promote DNA synthesis (Saini et al., 2013; Wilson et al., 2013); progression via a migrating D-loop or bubble resulting in conservative rather than semi-conservative replication (Saini et al., 2013); a massively increased rate of mutagenesis (Deem et al., 2011); an association with clustered mutations (Sakofsky et al., 2014); and an increased frequency of template switching (TS) (Smith et al., 2007). Given that a BIR-like process has been documented in human cells (Costantino et al., 2014; Roumelioti et al., 2016), it is highly likely that its error-prone nature and propensity for TS causes many of the mutations and genome rearrangements that are associated with cancer and other diseases. It is therefore imperative that the factors that influence BIR’s tendency to cause genomic change are identified, and their conservation of function in related RDR processes assessed, especially in the context of ongoing DNA replication during S-phase.

We, and others, have documented a related process to BIR in the fission yeast Schizosaccharomyces pombe, which is induced by the RTS1 protein-DNA polar replication fork barrier (RFB), as opposed to a DNA break (Lambert et al., 2010; Miyabe et al., 2015; Mizuno et al., 2013; Nguyen et al., 2015). This form of RDR has been termed homologous recombination-restarted replication (HoRReR) (Miyabe et al., 2015) but, in this paper, we will refer to it simply as RDR. Replication forks collapse, and recruit HR proteins, within approximately 10 min of collision with RTS1 (Mohebi et al., 2015; Nguyen et al., 2015). Similar to BIR, Rad51 and Rad52 promote replication restart at RTS1 and, in doing so, can inadvertently cause genomic rearrangements through the recombination of repetitive DNA elements (Ahn et al., 2005; Lambert et al., 2005; Lambert et al., 2010; Mizuno et al., 2009; Morrow et al., 2017; Nguyen et al., 2015). Also like BIR, RDR induced by RTS1 is highly mutagenic and prone to TS (Iraqui et al., 2012; Mizuno et al., 2013; Nguyen et al., 2015). In this paper, we advance from our earlier study by identifying some of the key factors that influence TS during RDR, including key antirecombinogenic DNA helicases and collisions between restarted replication and tRNA transcription. We also highlight apparent similarities and differences between TS associated with BIR in S. cerevisiae and RTS1-induced RDR in S. pombe.

The factors affecting TS associated with recombination-mediated DNA replication have hitherto been studied mainly in the context of BIR in S. cerevisiae and outside of S-phase. In our study we have measured TS associated with RDR from a collapsed, yet unbroken, replication fork in S. pombe. With this system, we have identified similarities with BIR in S. cerevisiae, as well as some potential differences. To place our findings into a conceptual framework, we will first discuss a hypothetical model for TS, which is elaborated from a previous model (Figure 7) (Nguyen et al., 2015). We envisage that the collapsed replication fork regresses to form a classic ‘chicken foot’ structure with an exposed ssDNA tail formed by the nucleolytic activity of the Mre11-Rad50-Nbs1-Ctp1 complex and Exo1 (Figure 7, step 1) (Teixeira-Silva et al., 2017). Rad51 loads onto the ssDNA tail to form a nucleoprotein filament that first locates a homologous duplex and then invades it to form a D-loop (Figure 7, step 2). Pol∂ extends the 3’ end of the invading strand (Miyabe et al., 2015) and, as it does, the D-loop moves in tandem (Figure 7, step 3) (Saini et al., 2013). Although not shown in Figure 7, lagging strand DNA synthesis may be established on the nascent leading strand giving rise to conservative replication, which could then be directed into semi-conservative replication by branch migration of the regressed replication fork behind the D-loop (Nguyen et al., 2015). Such a model would be compatible with the finding that RDR, triggered by fork collapse at RTS1, is semi-conservative (Miyabe et al., 2015), and contrasts with BIR, which is conservative (Donnianni and Symington, 2013; Saini et al., 2013). As DNA replication progresses, the associated migrating D-loop is likely to be susceptible to dissociation, either through the action of DNA helicases and/or by branch migration of the regressed replication fork ‘catching up’ with it (Nguyen et al., 2015). Dissociation of the D-loop in a region of repetitive DNA would lead to TS if the ejected DNA strand re-invades the ‘wrong’ repeat (Figure 7, steps 4 and 5; note that the DNA repeats, in yellow, are directly orientated as they are in our genetic assay). RDR is terminated upon encounter with the oncoming replication fork (Figure 7, steps 6 and 7) and, during this process, the invading DNA strand may either be ligated to the nascent lagging strand of the oncoming fork (Figure 7, step 7a) or be unwound (Figure 7, step 7b). If unwound, a second TS may be triggered (not shown) or the regressed replication fork may simply re-set by branch migration and merge with the oncoming fork (Figure 7, steps 8b and 9b). Note that these events can give rise to a gene conversion in our genetic assay (Figure 1B). Ligation of the invading DNA strand to the nascent lagging strand of the oncoming fork would prevent further TS and consolidate the formation of a deletion, with branched DNA structures being resolved by nucleases (not shown) (Lorenz et al., 2009) or by the completion of DNA replication (Figure 7, steps 8a and 9a). In the latter case, the region being deleted would form a heterologous loop of ssDNA (Figure 7, step 9a), which could be removed by a large DNA loop repair pathway (Clikeman et al., 2001; Sommer et al., 2008).

Figure 7

Download asset Open asset

TS associated with BIR in S. cerevisiae is thought to reflect an inherent instability of the migrating D-loop, which has evolved to encourage DSB repair by synthesis-dependent strand annealing (Smith et al., 2007). The observation that TS in S. cerevisae is constrained to the first 10 kb of template DNA that is copied, suggests that the D-loop eventually matures into a more stable structure (Mayle et al., 2015; Smith et al., 2007). This maturation process may involve cleavage of the D-loop by structure-specific nucleases such as Mus81 (Mayle et al., 2015; Osman and Whitby, 2007). Our finding that TS downstream of RTS1-AO is generally biased in favour of deletions, whereas at the RFB gene conversions predominate, suggests that the frequency of double TS events declines soon after replication has restarted. Interestingly, Carr and colleagues reported that the propensity for restarted replication to perform a U-turn at small inverted repeats positioned downstream of RTS1 declined 4-fold when the distance between RFB and repeats was increased from 0 to 2 kb, and then appeared to plateu (Mizuno et al., 2013). These observations suggest that the putative migrating D-loop, which is formed following replication fork collapse at RTS1-AO, undergoes partial stabilization shortly after it is formed. However, unlike with BIR in S. cerevisiae, a degree of D-loop instability must be retained because we can detect TS over distances of at least 75 kb downstream of the site of fork collapse. Our finding, that tRNA^GLU08HO increases TS downstream of RTS1-AO, suggests that head-on collisions with transcription complexes might exacerbate D-loop instability and be a key trigger in promoting TS.

The apparent disparity in D-loop maturation, between BIR in S. cerevisiae and RDR in S. pombe, may reflect differences in D-loop processing. In S. cerevisiae, Mus81 constrains TS during BIR presumably by resolving the D-loop into a more stable fork structure (Mayle et al., 2015). However, in S. pombe, we found no evidence that Mus81 is required to constrain TS during RDR. In fact the only constraint, over the region in which TS can occur in S. pombe, seems to be the point at which the restarted replication converges with the oncoming replication fork - a property which is shared with BIR in S. cerevisiae (Mayle et al., 2015; Nguyen et al., 2015).

TS associated with restarted replication is thought to be responsible for generating many of the genome rearrangements that are associated with cancer and other disease states (Carvalho and Lupski, 2016; Hastings et al., 2009). However, some complex genome rearrangements can span distances of hundreds of kilobases and, therefore, could not be formed from a single restarted replication event if TS was limited to a 10 kb region downstream of the site of fork collapse (Hastings et al., 2009). Our finding, that TS events occur over a much greater distance in S. pombe than they do for BIR in S. cerevisiae, shows that TS, associated with the repair of a single collapsed replication fork, may be capable of generating some of the long-range and complex genome rearrangements seen in humans.

Similar to BIR in S. cerevisiae, the Pif1 family DNA helicase Pfh1 promotes RDR, which explains why there is a general reduction in the levels of TS downstream of RTS1-AO in pfh1-m21 and -mt* mutants. In S. cerevisiae, Pif1 aids DNA synthesis during BIR by driving D-loop migration (Wilson et al., 2013), and Pfh1 might do likewise during RDR in S. pombe. Moreover, our observation, that resected regressed replication forks accumulate at RTS1-AO in a pfh1-mt* mutant, suggests that Pfh1 is a candidate for promoting the putative branch migration and/or re-setting of the regressed replication fork behind the migrating D-loop (see Discussion above). Pfh1 also has the ability to drive DNA replication by dislodging protein-DNA barriers via its so-called sweepase activity (Sabouri et al., 2012; Steinacher et al., 2012). Efficient RDR past RTS1 presumably requires removal of the barrier protein Rtf1, which may depend on Pfh1’s sweepase activity.

Despite Pfh1 being needed for efficient RDR, we were able to detect an increase in TS in a pfh1-m21 mutant if we provided more time for replication restart to occur, which shows that Pfh1, in addition to promoting RDR, also plays a role in suppressing TS. This finding contrasts with BIR in S. cerevisisae where Pif1 has no affect on TS (Stafa et al., 2014). Unlike S. pombe and humans, which have only one Pif1 family helicase, S. cerevisiae has a second Pif1 helicase called Rrm3, which, whilst seemingly not required for BIR, is important for the repair of replication-associated DSBs as well as replication past stable protein-DNA complexes (Ivessa et al., 2003; Muñoz-Galván et al., 2017; Wilson et al., 2013). It is possible that Pif1 and Rrm3 act redundantly in the suppression of TS in S. cerevisiae. Alternatively, the role of Pif1 helicases in suppressing TS may only be evident when RDR acts during S-phase alongside ongoing canonical DNA replication. Specifically, they may play an important role in directing the termination of DNA replication where a canonical fork encounters a D-loop. We speculate that Pfh1 drives D-loop dissociation during termination through its action as an accessory helicase at the oncoming DNA replication fork, where it likely translocates on the lagging template strand in a 5’ – 3’ direction (i.e. towards the D-loop and on the same DNA strand that the invading strand is annealed to) (Sabouri, 2017) (Figure 7, step 7b). Without this activity there would be a greater tendancy for the 3’ end of the invading strand at the D-loop to ligate to the lagging strand of the oncoming fork, which would specifically favour deletion formation in our genetic assay (Figure 7, steps 7a – 9a). Indeed, in the instances where we can detect an increase in Ade⁺ recombinants when pfh1 is mutated, it is specifically deletions that increase, with the frequency of gene conversions either decreasing or remaining unchanged (Figure 2, Figure 2—figure supplement 1, Figure 4, Figure 4—figure supplement 1).

In addition to Pfh1, we have identified a further three conserved DNA helicases that suppress TS downstream of RTS1-AO, namely Rqh1, Srs2 and Fbh1. These helicases, and their homologues in other organisms, are well known as major players in directing the outcome of homologous recombination reactions (Doe and Whitby, 2004; Fugger et al., 2009; Kohzaki et al., 2007; Larsen and Hickson, 2013; Lorenz et al., 2009; Morishita et al., 2005; Niu and Klein, 2017; Osman et al., 2005; Simandlova et al., 2013; Tsutsui et al., 2014). However, their role in suppressing TS downstream of a collapsed replication fork has not previously been demonstrated, though it has been suggested that the homologue of Rqh1 in S. cerevisiae (Sgs1) might limit translocations between highly diverged genes by suppressing TS during BIR (Schmidt et al., 2006). Based on the known activities of their homologues, Rqh1, Srs2 and Fbh1 have the potential to influence TS in various ways. For example, Sgs1 and its human homologue BLM, as well as S. cerevisiae Srs2, are capable of dissociating D-loops in vitro (Bachrati et al., 2006; Bugreev et al., 2007; Dupaigne et al., 2008; Fasching et al., 2015; Liu et al., 2017; van Brabant et al., 2000). The dissociation of a nascent D-loop can abort a TS event before a genetic change is made (e.g. a deletion or gene conversion in our genetic assay) and, therefore, the loss of this activity might account for the increase in TS we observe in both rqh1∆ and srs2∆ mutants (Figure 5). Indeed, Lambert and colleagues showed that Rqh1 was responsible for the disappearance of recombination intermediates at RTS1 in cells lacking CAF1, and proposed that this was due to its ability to drive D-loop dissociation (Pietrobon et al., 2014).

Another way in which Rqh1 may affect TS is by catalyzing the branch migration of the regressed replication fork behind the migrating D-loop. The regressed fork forms a four-way/Holliday junction, which is a structure that several RecQ–type helicases have been shown to unwind/branch migrate in vitro (Cejka and Kowalczykowski, 2010; Karow et al., 2000; LeRoy et al., 2005; Machwe et al., 2011). As discussed above, branch migration of the four-way junction into the ‘back’ of the D-loop would cause the D-loop to unwind. This might happen more frequently during an attempted TS event if the four-way junction continues to migrate whilst the D-loop is being re-established.

In S. cerevisiae, Rad51 promotes strand re-invasion, leading to a TS event during BIR (Anand et al., 2014). Presumably, Rad51 loads onto the ejected DNA strand following dissociation of the migrating D-loop to form a nucleoprotein filament. Both Srs2 and Fbh1 regulate nucleoprotein filament formation by dissociating Rad51 from DNA (Krejci et al., 2003; Simandlova et al., 2013; Tsutsui et al., 2014; Veaute et al., 2003). The loss of this anti-recombinogenic activity might account for the increase in TS observed in srs2∆ and fbh1∆ mutants.

Whilst dissociation of a nascent D-loop might prevent TS recombination, the unwinding of a fully established migrating D-loop would promote it. Indeed, this is exactly the activity that Mph1 is thought to catalyze during BIR in S. cerevisiae (Stafa et al., 2014). Surprisingly, we found that Fml1, despite promoting ectopic recombination during the restart of replication, appeared to have no effect on TS once replication had restarted. It is possible that a reduction in TS caused by lower levels of migrating D-loop dissociation is offset by an increased number of D-loops reaching our 12.4 kb genetic reporter. We are currently developing methods to measure restarted replication tract lengths, which will allow us to better calibrate our TS data.

In conclusion, we have identified a number of factors that influence TS associated with replication restart from a collapsed replication fork, revealing both similarities and potential differences with TS during BIR in S. cerevisiae. The fact that S. pombe and S. cerevisiae are only distant relatives, being about as divergent from each other as each is to animals (Heckman et al., 2001; Sipiczki, 2000), means that the similarities we have reported (e.g. Pif1 helicases promoting replication restart) are likely to be conserved in humans. Even the apparent disparities may simply reflect differences in the recombination assays used rather than fundamental mechanistic differences between RDR in S. pombe and S. cerevisiae. This is exemplified in S. cerevisiae, where the importance of Mus81 in limiting TS during BIR appears to depend on the recombination assay (Mayle et al., 2015; Stafa et al., 2014). In future studies, it will be important to establish whether the apparent disparities between RDR in S. pombe and BIR in S. cerevisiae reflect real species-specific differences, fundamental differences in the restart of replication from a DSB/broken replication fork versus a collapsed, yet unbroken, fork or simply differences in the recombination assays that have been used.

All data generated or analysed during this study are included in the manuscript and supporting files.

1. 1. Aguilera A
   2. García-Muse T

   (2013) Causes of genome instability

   Annual Review of Genetics 47:1–32.

   https://doi.org/10.1146/annurev-genet-111212-133232

   • PubMed
   • Google Scholar
2. 1. Ahn JS
   2. Osman F
   3. Whitby MC

   (2005) Replication fork blockage by RTS1 at an ectopic site promotes recombination in fission yeast

   The EMBO Journal 24:2011–2023.

   https://doi.org/10.1038/sj.emboj.7600670

   • PubMed
   • Google Scholar
3. 1. Ait Saada A
   2. Teixeira-Silva A
   3. Iraqui I
   4. Costes A
   5. Hardy J
   6. Paoletti G
   7. Fréon K
   8. Lambert SAE

   (2017) Unprotected replication forks are converted into mitotic sister chromatid bridges

   Molecular Cell 66:398–410.

   https://doi.org/10.1016/j.molcel.2017.04.002

   • PubMed
   • Google Scholar
4. 1. Anand RP
   2. Lovett ST
   3. Haber JE

   (2013) Break-induced DNA replication

   Cold Spring Harbor Perspectives in Biology 5:a010397.

   https://doi.org/10.1101/cshperspect.a010397

   • PubMed
   • Google Scholar
5. 1. Anand RP
   2. Tsaponina O
   3. Greenwell PW
   4. Lee CS
   5. Du W
   6. Petes TD
   7. Haber JE

   (2014) Chromosome rearrangements via template switching between diverged repeated sequences

   Genes & Development 28:2394–2406.

   https://doi.org/10.1101/gad.250258.114

   • PubMed
   • Google Scholar
6. 1. Arlt MF
   2. Wilson TE
   3. Glover TW

   (2012) Replication stress and mechanisms of CNV formation

   Current Opinion in Genetics & Development 22:204–210.

   https://doi.org/10.1016/j.gde.2012.01.009

   • PubMed
   • Google Scholar
7. 1. Bachrati CZ
   2. Borts RH
   3. Hickson ID

   (2006) Mobile D-loops are a preferred substrate for the bloom's syndrome helicase

   Nucleic Acids Research 34:2269–2279.

   https://doi.org/10.1093/nar/gkl258

   • PubMed
   • Google Scholar
8. 1. Bugreev DV
   2. Yu X
   3. Egelman EH
   4. Mazin AV

   (2007) Novel pro- and anti-recombination activities of the bloom's syndrome helicase

   Genes & Development 21:3085–3094.

   https://doi.org/10.1101/gad.1609007

   • PubMed
   • Google Scholar
9. 1. Carvalho CM
   2. Lupski JR

   (2016) Mechanisms underlying structural variant formation in genomic disorders

   Nature Reviews Genetics 17:224–238.

   https://doi.org/10.1038/nrg.2015.25

   • PubMed
   • Google Scholar
10. 1. Cejka P
    2. Kowalczykowski SC

    (2010) The full-length saccharomyces cerevisiae Sgs1 protein is a vigorous DNA helicase that preferentially unwinds holliday junctions

    Journal of Biological Chemistry 285:8290–8301.

    https://doi.org/10.1074/jbc.M109.083196

    • PubMed
    • Google Scholar
11. 1. Chiolo I
    2. Saponaro M
    3. Baryshnikova A
    4. Kim JH
    5. Seo YS
    6. Liberi G

    (2007) The human F-Box DNA helicase FBH1 faces saccharomyces cerevisiae Srs2 and postreplication repair pathway roles

    Molecular and Cellular Biology 27:7439–7450.

    https://doi.org/10.1128/MCB.00963-07

    • PubMed
    • Google Scholar
12. 1. Clikeman JA
    2. Wheeler SL
    3. Nickoloff JA

    (2001)

    Efficient incorporation of large (>2 kb) heterologies into heteroduplex DNA: pms1/Msh2-dependent and -independent large loop mismatch repair in saccharomyces cerevisiae

    Genetics 157:1481–1491.

    • PubMed
    • Google Scholar
13. 1. Cortez D

    (2015) Preventing replication fork collapse to maintain genome integrity

    DNA Repair 32:149–157.

    https://doi.org/10.1016/j.dnarep.2015.04.026

    • PubMed
    • Google Scholar
14. 1. Costantino L
    2. Sotiriou SK
    3. Rantala JK
    4. Magin S
    5. Mladenov E
    6. Helleday T
    7. Haber JE
    8. Iliakis G
    9. Kallioniemi OP
    10. Halazonetis TD

    (2014) Break-induced replication repair of damaged forks induces genomic duplications in human cells

    Science 343:88–91.

    https://doi.org/10.1126/science.1243211

    • PubMed
    • Google Scholar
15. 1. Deem A
    2. Keszthelyi A
    3. Blackgrove T
    4. Vayl A
    5. Coffey B
    6. Mathur R
    7. Chabes A
    8. Malkova A

    (2011) Break-induced replication is highly inaccurate

    PLOS Biology 9:e1000594.

    https://doi.org/10.1371/journal.pbio.1000594

    • PubMed
    • Google Scholar
16. 1. Deshpande AM
    2. Newlon CS

    (1996) DNA replication fork pause sites dependent on transcription

    Science 272:1030–1033.

    https://doi.org/10.1126/science.272.5264.1030

    • PubMed
    • Google Scholar
17. 1. Doe CL
    2. Whitby MC

    (2004) The involvement of Srs2 in post-replication repair and homologous recombination in fission yeast

    Nucleic Acids Research 32:1480–1491.

    https://doi.org/10.1093/nar/gkh317

    • PubMed
    • Google Scholar
18. 1. Donnianni RA
    2. Symington LS

    (2013) Break-induced replication occurs by conservative DNA synthesis

    PNAS 110:13475–13480.

    https://doi.org/10.1073/pnas.1309800110

    • Google Scholar
19. 1. Dupaigne P
    2. Le Breton C
    3. Fabre F
    4. Gangloff S
    5. Le Cam E
    6. Veaute X

    (2008) The Srs2 helicase activity is stimulated by Rad51 filaments on dsDNA: implications for crossover incidence during mitotic recombination

    Molecular Cell 29:243–254.

    https://doi.org/10.1016/j.molcel.2007.11.033

    • PubMed
    • Google Scholar
20. 1. Fasching CL
    2. Cejka P
    3. Kowalczykowski SC
    4. Heyer WD

    (2015) Top3-Rmi1 dissolve Rad51-mediated D loops by a topoisomerase-based mechanism

    Molecular Cell 57:595–606.

    https://doi.org/10.1016/j.molcel.2015.01.022

    • PubMed
    • Google Scholar
21. 1. Fugger K
    2. Mistrik M
    3. Danielsen JR
    4. Dinant C
    5. Falck J
    6. Bartek J
    7. Lukas J
    8. Mailand N

    (2009) Human Fbh1 helicase contributes to genome maintenance via pro- and anti-recombinase activities

    The Journal of Cell Biology 186:655–663.

    https://doi.org/10.1083/jcb.200812138

    • PubMed
    • Google Scholar
22. 1. Hastings PJ
    2. Ira G
    3. Lupski JR

    (2009) A microhomology-mediated break-induced replication model for the origin of human copy number variation

    PLOS Genetics 5:e1000327.

    https://doi.org/10.1371/journal.pgen.1000327

    • PubMed
    • Google Scholar
23. 1. Heckman DS
    2. Geiser DM
    3. Eidell BR
    4. Stauffer RL
    5. Kardos NL
    6. Hedges SB

    (2001) Molecular evidence for the early colonization of land by fungi and plants

    Science 293:1129–1133.

    https://doi.org/10.1126/science.1061457

    • PubMed
    • Google Scholar
24. 1. Inagawa T
    2. Yamada-Inagawa T
    3. Eydmann T
    4. Mian IS
    5. Wang TS
    6. Dalgaard JZ

    (2009) Schizosaccharomyces pombe Rtf2 mediates site-specific replication termination by inhibiting replication restart

    PNAS 106:7927–7932.

    https://doi.org/10.1073/pnas.0812323106

    • PubMed
    • Google Scholar
25. 1. Iraqui I
    2. Chekkal Y
    3. Jmari N
    4. Pietrobon V
    5. Fréon K
    6. Costes A
    7. Lambert SA

    (2012) Recovery of arrested replication forks by homologous recombination is error-prone

    PLOS Genetics 8:e1002976.

    https://doi.org/10.1371/journal.pgen.1002976

    • PubMed
    • Google Scholar
26. 1. Ivessa AS
    2. Lenzmeier BA
    3. Bessler JB
    4. Goudsouzian LK
    5. Schnakenberg SL
    6. Zakian VA

    (2003) The saccharomyces cerevisiae helicase Rrm3p facilitates replication past nonhistone protein-DNA complexes

    Molecular Cell 12:1525–1536.

    https://doi.org/10.1016/S1097-2765(03)00456-8

    • PubMed
    • Google Scholar
27. 1. Karow JK
    2. Constantinou A
    3. Li JL
    4. West SC
    5. Hickson ID

    (2000) The bloom's syndrome gene product promotes branch migration of holliday junctions

    PNAS 97:6504–6508.

    https://doi.org/10.1073/pnas.100448097

    • PubMed
    • Google Scholar
28. 1. Kohzaki M
    2. Hatanaka A
    3. Sonoda E
    4. Yamazoe M
    5. Kikuchi K
    6. Vu Trung N
    7. Szüts D
    8. Sale JE
    9. Shinagawa H
    10. Watanabe M
    11. Takeda S

    (2007) Cooperative roles of vertebrate Fbh1 and blm DNA helicases in avoidance of crossovers during recombination initiated by replication fork collapse

    Molecular and Cellular Biology 27:2812–2820.

    https://doi.org/10.1128/MCB.02043-06

    • PubMed
    • Google Scholar
29. 1. Kramara J
    2. Osia B
    3. Malkova A

    (2018) Break-Induced replication: the where, the why, and the how

    Trends in Genetics 34:518–531.

    https://doi.org/10.1016/j.tig.2018.04.002

    • PubMed
    • Google Scholar
30. 1. Krejci L
    2. Van Komen S
    3. Li Y
    4. Villemain J
    5. Reddy MS
    6. Klein H
    7. Ellenberger T
    8. Sung P

    (2003) DNA helicase Srs2 disrupts the Rad51 presynaptic filament

    Nature 423:305–309.

    https://doi.org/10.1038/nature01577

    • PubMed
    • Google Scholar
31. 1. Krogh BO
    2. Symington LS

    (2004) Recombination proteins in yeast

    Annual Review of Genetics 38:233–271.

    https://doi.org/10.1146/annurev.genet.38.072902.091500

    • PubMed
    • Google Scholar
32. 1. Lambert S
    2. Watson A
    3. Sheedy DM
    4. Martin B
    5. Carr AM

    (2005) Gross chromosomal rearrangements and elevated recombination at an inducible site-specific replication fork barrier

    Cell 121:689–702.

    https://doi.org/10.1016/j.cell.2005.03.022

    • PubMed
    • Google Scholar
33. 1. Lambert S
    2. Mizuno K
    3. Blaisonneau J
    4. Martineau S
    5. Chanet R
    6. Fréon K
    7. Murray JM
    8. Carr AM
    9. Baldacci G

    (2010) Homologous recombination restarts blocked replication forks at the expense of genome rearrangements by template exchange

    Molecular Cell 39:346–359.

    https://doi.org/10.1016/j.molcel.2010.07.015

    • PubMed
    • Google Scholar
34. 1. Lambert S
    2. Carr AM

    (2013) Impediments to replication fork movement: stabilisation, reactivation and genome instability

    Chromosoma 122:33–45.

    https://doi.org/10.1007/s00412-013-0398-9

    • PubMed
    • Google Scholar
35. 1. Larsen NB
    2. Hickson ID

    (2013) RecQ helicases: conserved guardians of genomic integrity

    Advances in Experimental Medicine and Biology 767:161–184.

    https://doi.org/10.1007/978-1-4614-5037-5_8

    • PubMed
    • Google Scholar
36. 1. LeRoy G
    2. Carroll R
    3. Kyin S
    4. Seki M
    5. Cole MD

    (2005) Identification of RecQL1 as a holliday junction processing enzyme in human cell lines

    Nucleic Acids Research 33:6251–6257.

    https://doi.org/10.1093/nar/gki929

    • PubMed
    • Google Scholar
37. 1. Liu J
    2. Ede C
    3. Wright WD
    4. Gore SK
    5. Jenkins SS
    6. Freudenthal BD
    7. Todd Washington M
    8. Veaute X
    9. Heyer WD

    (2017) Srs2 promotes synthesis-dependent strand annealing by disrupting DNA polymerase δ-extending D-loops

    eLife 6:e22195.

    https://doi.org/10.7554/eLife.22195

    • PubMed
    • Google Scholar
38. 1. Lorenz A
    2. Osman F
    3. Folkyte V
    4. Sofueva S
    5. Whitby MC

    (2009) Fbh1 limits Rad51-dependent recombination at blocked replication forks

    Molecular and Cellular Biology 29:4742–4756.

    https://doi.org/10.1128/MCB.00471-09

    • PubMed
    • Google Scholar
39. 1. Lydeard JR
    2. Jain S
    3. Yamaguchi M
    4. Haber JE

    (2007) Break-induced replication and telomerase-independent telomere maintenance require Pol32

    Nature 448:820–823.

    https://doi.org/10.1038/nature06047

    • PubMed
    • Google Scholar
40. 1. Lydeard JR
    2. Lipkin-Moore Z
    3. Sheu YJ
    4. Stillman B
    5. Burgers PM
    6. Haber JE

    (2010) Break-induced replication requires all essential DNA replication factors except those specific for pre-RC assembly

    Genes & Development 24:1133–1144.

    https://doi.org/10.1101/gad.1922610

    • PubMed
    • Google Scholar
41. 1. Macheret M
    2. Halazonetis TD

    (2015) DNA replication stress as a hallmark of cancer

    Annual Review of Pathology: Mechanisms of Disease 10:425–448.

    https://doi.org/10.1146/annurev-pathol-012414-040424

    • PubMed
    • Google Scholar
42. 1. Machwe A
    2. Karale R
    3. Xu X
    4. Liu Y
    5. Orren DK

    (2011) The Werner and bloom syndrome proteins help resolve replication blockage by converting (regressed) holliday junctions to functional replication forks

    Biochemistry 50:6774–6788.

    https://doi.org/10.1021/bi2001054

    • PubMed
    • Google Scholar
43. 1. Marini V
    2. Krejci L

    (2010) Srs2: the "Odd-Job Man" in DNA repair

    DNA Repair 9:268–275.

    https://doi.org/10.1016/j.dnarep.2010.01.007

    • PubMed
    • Google Scholar
44. 1. Mayle R
    2. Campbell IM
    3. Beck CR
    4. Yu Y
    5. Wilson M
    6. Shaw CA
    7. Bjergbaek L
    8. Lupski JR
    9. Ira G

    (2015) Mus81 and converging forks limit the mutagenicity of replication fork breakage

    Science 349:742–747.

    https://doi.org/10.1126/science.aaa8391

    • PubMed
    • Google Scholar
45. 1. McFarlane RJ
    2. Whitehall SK

    (2009) tRNA genes in eukaryotic genome organization and reorganization

    Cell Cycle 8:3102–3106.

    https://doi.org/10.4161/cc.8.19.9625

    • PubMed
    • Google Scholar
46. 1. Miyabe I
    2. Mizuno K
    3. Keszthelyi A
    4. Daigaku Y
    5. Skouteri M
    6. Mohebi S
    7. Kunkel TA
    8. Murray JM
    9. Carr AM

    (2015) Polymerase δ replicates both strands after homologous recombination-dependent fork restart

    Nature Structural & Molecular Biology 22:932–938.

    https://doi.org/10.1038/nsmb.3100

    • PubMed
    • Google Scholar
47. 1. Mizuno K
    2. Lambert S
    3. Baldacci G
    4. Murray JM
    5. Carr AM

    (2009) Nearby inverted repeats fuse to generate acentric and dicentric palindromic chromosomes by a replication template exchange mechanism

    Genes & Development 23:2876–2886.

    https://doi.org/10.1101/gad.1863009

    • PubMed
    • Google Scholar
48. 1. Mizuno K
    2. Miyabe I
    3. Schalbetter SA
    4. Carr AM
    5. Murray JM

    (2013) Recombination-restarted replication makes inverted chromosome fusions at inverted repeats

    Nature 493:246–249.

    https://doi.org/10.1038/nature11676

    • PubMed
    • Google Scholar
49. 1. Mohebi S
    2. Mizuno Ken’Ichi
    3. Watson A
    4. Carr AM
    5. Murray JM

    (2015) Checkpoints are blind to replication restart and recombination intermediates that result in gross chromosomal rearrangements

    Nature Communications 6:e6357.

    https://doi.org/10.1038/ncomms7357

    • Google Scholar
50. 1. Morishita T
    2. Furukawa F
    3. Sakaguchi C
    4. Toda T
    5. Carr AM
    6. Iwasaki H
    7. Shinagawa H

    (2005) Role of the schizosaccharomyces pombe F-Box DNA helicase in processing recombination intermediates

    Molecular and Cellular Biology 25:8074–8083.

    https://doi.org/10.1128/MCB.25.18.8074-8083.2005

    • PubMed
    • Google Scholar
51. 1. Morrow CA
    2. Nguyen MO
    3. Fower A
    4. Wong IN
    5. Osman F
    6. Bryer C
    7. Whitby MC

    (2017) Inter-Fork strand annealing causes genomic deletions during the termination of DNA replication

    eLife 6:e25490.

    https://doi.org/10.7554/eLife.25490

    • PubMed
    • Google Scholar
52. 1. Muñoz-Galván S
    2. García-Rubio M
    3. Ortega P
    4. Ruiz JF
    5. Jimeno S
    6. Pardo B
    7. Gómez-González B
    8. Aguilera A

    (2017) A new role for Rrm3 in repair of replication-born DNA breakage by sister chromatid recombination

    PLOS Genetics 13:e1006781.

    https://doi.org/10.1371/journal.pgen.1006781

    • PubMed
    • Google Scholar
53. 1. Nguyen MO
    2. Jalan M
    3. Morrow CA
    4. Osman F
    5. Whitby MC

    (2015) Recombination occurs within minutes of replication blockage by RTS1 producing restarted forks that are prone to collapse

    eLife 4:e04539.

    https://doi.org/10.7554/eLife.04539

    • PubMed
    • Google Scholar
54. 1. Niu H
    2. Klein HL

    (2017) Multifunctional roles of saccharomyces cerevisiae Srs2 protein in replication, recombination and repair

    FEMS FEMS Yeast Res 17:efow111.

    https://doi.org/10.1093/femsyr/fow111

    • Google Scholar
55. 1. Osman F
    2. Dixon J
    3. Barr AR
    4. Whitby MC

    (2005) The F-Box DNA helicase Fbh1 prevents Rhp51-dependent recombination without mediator proteins

    Molecular and Cellular Biology 25:8084–8096.

    https://doi.org/10.1128/MCB.25.18.8084-8096.2005

    • PubMed
    • Google Scholar
56. 1. Osman F
    2. Whitby MC

    (2007) Exploring the roles of Mus81-Eme1/Mms4 at perturbed replication forks

    DNA Repair 6:1004–1017.

    https://doi.org/10.1016/j.dnarep.2007.02.019

    • PubMed
    • Google Scholar
57. 1. Osmundson JS
    2. Kumar J
    3. Yeung R
    4. Smith DJ

    (2017) Pif1-family helicases cooperatively suppress widespread replication-fork arrest at tRNA genes

    Nature Structural & Molecular Biology 24:162–170.

    https://doi.org/10.1038/nsmb.3342

    • PubMed
    • Google Scholar
58. 1. Pietrobon V
    2. Fréon K
    3. Hardy J
    4. Costes A
    5. Iraqui I
    6. Ochsenbein F
    7. Lambert SA

    (2014) The chromatin assembly factor 1 promotes Rad51-dependent template switches at replication forks by counteracting D-loop disassembly by the RecQ-type helicase Rqh1

    PLOS Biology 12:e1001968.

    https://doi.org/10.1371/journal.pbio.1001968

    • PubMed
    • Google Scholar
59. 1. Pinter SF
    2. Aubert SD
    3. Zakian VA

    (2008) The schizosaccharomyces pombe Pfh1p DNA helicase is essential for the maintenance of nuclear and mitochondrial DNA

    Molecular and Cellular Biology 28:6594–6608.

    https://doi.org/10.1128/MCB.00191-08

    • PubMed
    • Google Scholar
60. 1. Pryce DW
    2. Ramayah S
    3. Jaendling A
    4. McFarlane RJ

    (2009) Recombination at DNA replication fork barriers is not universal and is differentially regulated by Swi1

    PNAS 106:4770–4775.

    https://doi.org/10.1073/pnas.0807739106

    • PubMed
    • Google Scholar
61. 1. Roumelioti FM
    2. Sotiriou SK
    3. Katsini V
    4. Chiourea M
    5. Halazonetis TD
    6. Gagos S

    (2016) Alternative lengthening of human telomeres is a conservative DNA replication process with features of break-induced replication

    EMBO Reports 17:1731–1737.

    https://doi.org/10.15252/embr.201643169

    • PubMed
    • Google Scholar
62. 1. Sabouri N
    2. McDonald KR
    3. Webb CJ
    4. Cristea IM
    5. Zakian VA

    (2012) DNA replication through hard-to-replicate sites, including both highly transcribed RNA pol II and pol III genes, requires the S. pombe Pfh1 helicase

    Genes & Development 26:581–593.

    https://doi.org/10.1101/gad.184697.111

    • PubMed
    • Google Scholar
63. 1. Sabouri N

    (2017) The functions of the multi-tasking Pfh1^Pif1 helicase

    Current Genetics 63:621–626.

    https://doi.org/10.1007/s00294-016-0675-2

    • PubMed
    • Google Scholar
64. 1. Saini N
    2. Ramakrishnan S
    3. Elango R
    4. Ayyar S
    5. Zhang Y
    6. Deem A
    7. Ira G
    8. Haber JE
    9. Lobachev KS
    10. Malkova A

    (2013) Migrating bubble during break-induced replication drives conservative DNA synthesis

    Nature 502:389–392.

    https://doi.org/10.1038/nature12584

    • PubMed
    • Google Scholar
65. 1. Sakofsky CJ
    2. Roberts SA
    3. Malc E
    4. Mieczkowski PA
    5. Resnick MA
    6. Gordenin DA
    7. Malkova A

    (2014) Break-induced replication is a source of mutation clusters underlying kataegis

    Cell Reports 7:1640–1648.

    https://doi.org/10.1016/j.celrep.2014.04.053

    • PubMed
    • Google Scholar
66. 1. Sakofsky CJ
    2. Malkova A

    (2017) Break induced replication in eukaryotes: mechanisms, functions, and consequences

    Critical Reviews in Biochemistry and Molecular Biology 52:395–413.

    https://doi.org/10.1080/10409238.2017.1314444

    • PubMed
    • Google Scholar
67. 1. Schmidt KH
    2. Wu J
    3. Kolodner RD

    (2006) Control of translocations between highly diverged genes by Sgs1, the saccharomyces cerevisiae homolog of the Bloom's syndrome protein

    Molecular and Cellular Biology 26:5406–5420.

    https://doi.org/10.1128/MCB.00161-06

    • PubMed
    • Google Scholar
68. 1. Simandlova J
    2. Zagelbaum J
    3. Payne MJ
    4. Chu WK
    5. Shevelev I
    6. Hanada K
    7. Chatterjee S
    8. Reid DA
    9. Liu Y
    10. Janscak P
    11. Rothenberg E
    12. Hickson ID

    (2013) FBH1 helicase disrupts RAD51 filaments in vitro and modulates homologous recombination in mammalian cells

    Journal of Biological Chemistry 288:34168–34180.

    https://doi.org/10.1074/jbc.M113.484493

    • PubMed
    • Google Scholar
69. 1. Sipiczki M

    (2000) Where does fission yeast sit on the tree of life?

    Genome Biology 1: reviews1011.1.

    https://doi.org/10.1186/gb-2000-1-2-reviews1011

    • Google Scholar
70. 1. Smith CE
    2. Llorente B
    3. Symington LS

    (2007) Template switching during break-induced replication

    Nature 447:102–105.

    https://doi.org/10.1038/nature05723

    • PubMed
    • Google Scholar
71. 1. Sommer D
    2. Stith CM
    3. Burgers PM
    4. Lahue RS

    (2008) Partial reconstitution of DNA large loop repair with purified proteins from saccharomyces cerevisiae

    Nucleic Acids Research 36:4699–4707.

    https://doi.org/10.1093/nar/gkn446

    • PubMed
    • Google Scholar
72. 1. Stafa A
    2. Donnianni RA
    3. Timashev LA
    4. Lam AF
    5. Symington LS

    (2014) Template switching during break-induced replication is promoted by the Mph1 helicase in saccharomyces cerevisiae

    Genetics 196:1017–1028.

    https://doi.org/10.1534/genetics.114.162297

    • PubMed
    • Google Scholar
73. 1. Steinacher R
    2. Osman F
    3. Dalgaard JZ
    4. Lorenz A
    5. Whitby MC

    (2012) The DNA helicase Pfh1 promotes fork merging at replication termination sites to ensure genome stability

    Genes & Development 26:594–602.

    https://doi.org/10.1101/gad.184663.111

    • PubMed
    • Google Scholar
74. 1. Sun W
    2. Nandi S
    3. Osman F
    4. Ahn JS
    5. Jakovleska J
    6. Lorenz A
    7. Whitby MC

    (2008) The FANCM ortholog Fml1 promotes recombination at stalled replication forks and limits crossing over during DNA double-strand break repair

    Molecular Cell 32:118–128.

    https://doi.org/10.1016/j.molcel.2008.08.024

    • PubMed
    • Google Scholar
75. 1. Teixeira-Silva A
    2. Ait Saada A
    3. Hardy J
    4. Iraqui I
    5. Nocente MC
    6. Fréon K
    7. Lambert SAE

    (2017) The end-joining factor ku acts in the end-resection of double strand break-free arrested replication forks

    Nature Communications 8:e1982.

    https://doi.org/10.1038/s41467-017-02144-5

    • Google Scholar
76. 1. Tran PLT
    2. Pohl TJ
    3. Chen CF
    4. Chan A
    5. Pott S
    6. Zakian VA

    (2017) PIF1 family DNA helicases suppress R-loop mediated genome instability at tRNA genes

    Nature Communications 8:15025.

    https://doi.org/10.1038/ncomms15025

    • PubMed
    • Google Scholar
77. 1. Tsang E
    2. Miyabe I
    3. Iraqui I
    4. Zheng J
    5. Lambert SA
    6. Carr AM

    (2014) The extent of error-prone replication restart by homologous recombination is controlled by Exo1 and checkpoint proteins

    Journal of Cell Science 127:2983–2994.

    https://doi.org/10.1242/jcs.152678

    • PubMed
    • Google Scholar
78. 1. Tsutsui Y
    2. Kurokawa Y
    3. Ito K
    4. Siddique MS
    5. Kawano Y
    6. Yamao F
    7. Iwasaki H

    (2014) Multiple regulation of Rad51-mediated homologous recombination by fission yeast Fbh1

    PLOS Genetics 10:e1004542.

    https://doi.org/10.1371/journal.pgen.1004542

    • PubMed
    • Google Scholar
79. 1. van Brabant AJ
    2. Ye T
    3. Sanz M
    4. German III JL
    5. Ellis NA
    6. Holloman WK

    (2000) Binding and melting of D-loops by the bloom syndrome helicase

    Biochemistry 39:14617–14625.

    https://doi.org/10.1021/bi0018640

    • PubMed
    • Google Scholar
80. 1. Veaute X
    2. Jeusset J
    3. Soustelle C
    4. Kowalczykowski SC
    5. Le Cam E
    6. Fabre F

    (2003) The Srs2 helicase prevents recombination by disrupting Rad51 nucleoprotein filaments

    Nature 423:309–312.

    https://doi.org/10.1038/nature01585

    • PubMed
    • Google Scholar
81. 1. Wilson MA
    2. Kwon Y
    3. Xu Y
    4. Chung WH
    5. Chi P
    6. Niu H
    7. Mayle R
    8. Chen X
    9. Malkova A
    10. Sung P
    11. Ira G

    (2013) Pif1 helicase and polδ promote recombination-coupled DNA synthesis via bubble migration

    Nature 502:393–396.

    https://doi.org/10.1038/nature12585

    • PubMed
    • Google Scholar

Author details

1. Manisha Jalan

   Department of Biochemistry, University of Oxford, Oxford, United Kingdom

   Present address

   Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, United States

   Contribution

   Conceptualization, Data curation, Formal analysis, Validation, Investigation, Visualization, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-4467-4934

2. Judith Oehler

   Department of Biochemistry, University of Oxford, Oxford, United Kingdom

   Contribution

   Formal analysis, Investigation, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-8397-6492

3. Carl A Morrow

   Department of Biochemistry, University of Oxford, Oxford, United Kingdom

   Present address

   Department of Oncology, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, United Kingdom

   Contribution

   Formal analysis, Investigation, Writing—review and editing

   Competing interests

   No competing interests declared

4. Fekret Osman

   Department of Biochemistry, University of Oxford, Oxford, United Kingdom

   Contribution

   Formal analysis, Supervision, Investigation, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

5. Matthew C Whitby

   Department of Biochemistry, University of Oxford, Oxford, United Kingdom

   Contribution

   Conceptualization, Resources, Supervision, Funding acquisition, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing

   For correspondence

   matthew.whitby@bioch.ox.ac.uk

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0003-0951-3374

• 2,859

  views

• 421

  downloads

• 43

  citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.