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Recovery of arrested replication forks by homologous recombination is error-prone.

Iraqui I, Chekkal Y, Jmari N, Pietrobon V, Fréon K, Costes A, Lambert SA - PLoS Genet. (2012)

Bottom Line: The mutations caused are small insertions/duplications between short tandem repeats (micro-homology) indicative of replication slippage.Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage.We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Institut Curie, Centre de Recherche, Orsay, France.

ABSTRACT
Homologous recombination is a universal mechanism that allows repair of DNA and provides support for DNA replication. Homologous recombination is therefore a major pathway that suppresses non-homology-mediated genome instability. Here, we report that recovery of impeded replication forks by homologous recombination is error-prone. Using a fork-arrest-based assay in fission yeast, we demonstrate that a single collapsed fork can cause mutations and large-scale genomic changes, including deletions and translocations. Fork-arrest-induced gross chromosomal rearrangements are mediated by inappropriate ectopic recombination events at the site of collapsed forks. Inverted repeats near the site of fork collapse stimulate large-scale genomic changes up to 1,500 times over spontaneous events. We also show that the high accuracy of DNA replication during S-phase is impaired by impediments to fork progression, since fork-arrest-induced mutation is due to erroneous DNA synthesis during recovery of replication forks. The mutations caused are small insertions/duplications between short tandem repeats (micro-homology) indicative of replication slippage. Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage. The inaccuracy of DNA synthesis does not rely on PCNA ubiquitination or trans-lesion-synthesis DNA polymerases, and it is not counteracted by mismatch repair. We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

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Model of replication-stress-induced genetic instability at collapsed forks.Collapsed forks might arise from torsional stress, fork breakage (i.e. at nick, ICLs), or proteins tightly-bound to DNA. Replisome disassembly at collapsed forks may favour the unwinding of the nascent strand on which Rad51 nucleates. At this initial stage of fork resumption by recombination, homology-driven template exchange can promote intra- or inter-recombination resulting in GCRs. Fork recovery by recombination overcomes the initial replication block and allows an inaccurate replisome to form (see text for details).
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pgen-1002976-g006: Model of replication-stress-induced genetic instability at collapsed forks.Collapsed forks might arise from torsional stress, fork breakage (i.e. at nick, ICLs), or proteins tightly-bound to DNA. Replisome disassembly at collapsed forks may favour the unwinding of the nascent strand on which Rad51 nucleates. At this initial stage of fork resumption by recombination, homology-driven template exchange can promote intra- or inter-recombination resulting in GCRs. Fork recovery by recombination overcomes the initial replication block and allows an inaccurate replisome to form (see text for details).

Mentions: Using conditional fork arrest constructs, we studied the consequences for genome instability of impediments to replication forks progression. A single fork arrest results in large-scale genomic changes and mutations that occur during recombination-mediated fork recovery (Figure 6). Inappropriate ectopic recombination at arrested forks results in GCRs, whereas appropriate restarting of the fork on the initial template results in error-prone DNA synthesis. GCRs and mutations at collapsed forks are genetically separable: Rqh1 limits fork-arrest-induced GCRs but not mutations (Figure 2D and Figure 3B). We demonstrate here that collapsed forks whose progression resumes by recombination lose accuracy during DNA synthesis, resulting in frequent intra-template switches. Thus, homologous recombination contributes to completion of DNA replication when forks progression is impeded but also fuels genome modifications both at the chromosomal and nucleotide level.


Recovery of arrested replication forks by homologous recombination is error-prone.

Iraqui I, Chekkal Y, Jmari N, Pietrobon V, Fréon K, Costes A, Lambert SA - PLoS Genet. (2012)

Model of replication-stress-induced genetic instability at collapsed forks.Collapsed forks might arise from torsional stress, fork breakage (i.e. at nick, ICLs), or proteins tightly-bound to DNA. Replisome disassembly at collapsed forks may favour the unwinding of the nascent strand on which Rad51 nucleates. At this initial stage of fork resumption by recombination, homology-driven template exchange can promote intra- or inter-recombination resulting in GCRs. Fork recovery by recombination overcomes the initial replication block and allows an inaccurate replisome to form (see text for details).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3475662&req=5

pgen-1002976-g006: Model of replication-stress-induced genetic instability at collapsed forks.Collapsed forks might arise from torsional stress, fork breakage (i.e. at nick, ICLs), or proteins tightly-bound to DNA. Replisome disassembly at collapsed forks may favour the unwinding of the nascent strand on which Rad51 nucleates. At this initial stage of fork resumption by recombination, homology-driven template exchange can promote intra- or inter-recombination resulting in GCRs. Fork recovery by recombination overcomes the initial replication block and allows an inaccurate replisome to form (see text for details).
Mentions: Using conditional fork arrest constructs, we studied the consequences for genome instability of impediments to replication forks progression. A single fork arrest results in large-scale genomic changes and mutations that occur during recombination-mediated fork recovery (Figure 6). Inappropriate ectopic recombination at arrested forks results in GCRs, whereas appropriate restarting of the fork on the initial template results in error-prone DNA synthesis. GCRs and mutations at collapsed forks are genetically separable: Rqh1 limits fork-arrest-induced GCRs but not mutations (Figure 2D and Figure 3B). We demonstrate here that collapsed forks whose progression resumes by recombination lose accuracy during DNA synthesis, resulting in frequent intra-template switches. Thus, homologous recombination contributes to completion of DNA replication when forks progression is impeded but also fuels genome modifications both at the chromosomal and nucleotide level.

Bottom Line: The mutations caused are small insertions/duplications between short tandem repeats (micro-homology) indicative of replication slippage.Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage.We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Institut Curie, Centre de Recherche, Orsay, France.

ABSTRACT
Homologous recombination is a universal mechanism that allows repair of DNA and provides support for DNA replication. Homologous recombination is therefore a major pathway that suppresses non-homology-mediated genome instability. Here, we report that recovery of impeded replication forks by homologous recombination is error-prone. Using a fork-arrest-based assay in fission yeast, we demonstrate that a single collapsed fork can cause mutations and large-scale genomic changes, including deletions and translocations. Fork-arrest-induced gross chromosomal rearrangements are mediated by inappropriate ectopic recombination events at the site of collapsed forks. Inverted repeats near the site of fork collapse stimulate large-scale genomic changes up to 1,500 times over spontaneous events. We also show that the high accuracy of DNA replication during S-phase is impaired by impediments to fork progression, since fork-arrest-induced mutation is due to erroneous DNA synthesis during recovery of replication forks. The mutations caused are small insertions/duplications between short tandem repeats (micro-homology) indicative of replication slippage. Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage. The inaccuracy of DNA synthesis does not rely on PCNA ubiquitination or trans-lesion-synthesis DNA polymerases, and it is not counteracted by mismatch repair. We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

Show MeSH
Related in: MedlinePlus