Genetic and Molecular Analysis of Mitotic Recombination in Saccharomyces cerevisiae

Gómez-González, Belén; Ruiz, José F.; Aguilera, Andrés

doi:10.1007/978-1-61779-129-1_10

Cited by 20 publications

(33 citation statements)

References 33 publications

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“…The use of a minimal 24-bp endonuclease HO site (mini HO r) [20] provides a unique tool to mimic a natural situation in which DSBs appear as a consequence of replication failures [6, 21]. At this site, the HO endonuclease produces mainly single-stranded DNA nicks on any of the strands that are converted into DSBs when they are encountered by the RFs [6].…”

Section: Resultsmentioning

confidence: 99%

“…The chromosomal system, instead, detects only intermolecular recombination (Fig 1C). In addition, Leu+ recombinants can arise by either gene conversion or reciprocal exchange in the TINV system [21], and as a result of gene conversion in the chromosomal system (Fig 1C), since reciprocal exchange would lead to an unviable dicentric chromosome. Note that the leu2-k and the leu2 Δ 5’ alleles used as donors of repair of the HO break are genetically equivalent for our purpose, since leu2 Δ 5’ is truncated at the Cla I site, close by the Kpn I site mutated in leu2-k [5, 22].…”

Section: Resultsmentioning

confidence: 99%

“…When cells containing the HO endonuclease gene under the GAL1 promoter grow in galactose media, replication-born DSBs can be observed by Southern-blot as 2.4 and 1.4 Kb bands [5, 21]. At the same time, DSB repair leads to the formation of new 4.7- and 2.9-Kb bands, the first of which is exclusively a consequence of unequal Sister Chromatid Exchange (SCE) events [5, 21]. It has been shown that this is an accurate indicator of the proficiency in total SCR [5, 6, 27].…”

Section: Resultsmentioning

confidence: 99%

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

et al. 2017

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Replication forks stall at different DNA obstacles such as those originated by transcription. Fork stalling can lead to DNA double-strand breaks (DSBs) that will be preferentially repaired by homologous recombination when the sister chromatid is available. The Rrm3 helicase is a replisome component that promotes replication upon fork stalling, accumulates at highly transcribed regions and prevents not only transcription-induced replication fork stalling but also transcription-associated hyper-recombination. This led us to explore the possible role of Rrm3 in the repair of DSBs when originating at the passage of the replication fork. Using a mini-HO system that induces mainly single-stranded DNA breaks, we show that rrm3Δ cells are defective in DSB repair. The defect is clearly seen in sister chromatid recombination, the major repair pathway of replication-born DSBs. Our results indicate that Rrm3 recruitment to replication-born DSBs is crucial for viability, uncovering a new role for Rrm3 in the repair of broken replication forks.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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

et al. 2017

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“…Indeed, recombination between two leu2 direct repeats in the plasmid pLYDNS and the chromosomal leu2-kTADE2-URA3Tleu2-k (Lk-AU) (Gomez-Gonzalez et al 2011b) systems was slightly but significantly increased with respect to wild-type levels (Fig. 1C,D).…”

Section: Genome Instability and Recombination-dependent Viability In mentioning

confidence: 96%

“…We assayed recombination between two leu2 direct repeats separated by the GC-rich lacZ gene under the inducible GAL1,10 promoter (P GAL ) or the constitutive LEU2 promoter (P LEU2 ) (Gomez-Gonzalez et al 2011b). When transcription was low (P GAL in glucose), recombination levels in spt16-11, spt16-G132D, and pob3-Q308K were indistinguishable from the wild type ( Fig.…”

Section: Genome Instability In Yeast Fact Mutants Is Transcription-dementioning

confidence: 99%

The yeast and human FACT chromatin-reorganizing complexes solve R-loop-mediated transcription–replication conflicts

et al. 2014

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FACT (facilitates chromatin transcription) is a chromatin-reorganizing complex that swaps nucleosomes around the RNA polymerase during transcription elongation and has a role in replication that is not fully understood yet. Here we show that recombination factors are required for the survival of yeast FACT mutants, consistent with an accumulation of DNA breaks that we detected by Rad52 foci and transcription-dependent hyperrecombination. Breaks also accumulate in FACT-depleted human cells, as shown by gH2AX foci and single-cell electrophoresis. Furthermore, FACT-deficient yeast and human cells show replication impairment, which in yeast we demonstrate by ChIP-chip (chromatin immunoprecipitation [ChIP] coupled with microarray analysis) of Rrm3 to occur genome-wide but preferentially at highly transcribed regions. Strikingly, in yeast FACT mutants, high levels of Rad52 foci are suppressed by RNH1 overexpression; R loops accumulate at high levels, and replication becomes normal when global RNA synthesis is inhibited in FACT-depleted human cells. The results demonstrate a key function of FACT in the resolution of R-loop-mediated transcription-replication conflicts, likely associated with a specific chromatin organization.

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Intrachromosomal Recombination in Yeast

Epshtein

Klein

2020

Homologous Recombination

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Genetic and Molecular Analysis of Mitotic Recombination in Saccharomyces cerevisiae

Cited by 20 publications

References 33 publications

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

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

The yeast and human FACT chromatin-reorganizing complexes solve R-loop-mediated transcription–replication conflicts

Intrachromosomal Recombination in Yeast

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