Dynamique des chromosomes et recombinaison

Publications de l’équipe

Année de publication : 2018

Adam C, Guérois R, Citarella A, Verardi L, Adolphe F Béneut C, Sommermeyer V, Ramus C, Govin J, Couté Y, Borde V (2018 Feb 1)

The PHD finger protein Spp1 has distinct functions in the Set1 and the meiotic DSB formation complexes

PLoS Genetics : 14(2) : DOI : 10.1371/journal.pgen.1007223 En savoir plus

Histone H3K4 methylation is a feature of meiotic recombination hotspots shared by many organisms including plants and mammals. Meiotic recombination is initiated by programmed double-strand break (DSB) formation that in budding yeast takes place in gene promoters and is promoted by histone H3K4 di/trimethylation. This histone modification is recognized by Spp1, a PHD finger containing protein that belongs to the conserved histone H3K4 methyltransferase Set1 complex. During meiosis, Spp1 binds H3K4me3 and interacts with a DSB protein, Mer2, to promote DSB formation close to gene promoters. How Set1 complex- and Mer2- related functions of Spp1 are connected is not clear. Here, combining genome-wide localization analyses, biochemical approaches and the use of separation of function mutants, we show that Spp1 is present within two distinct complexes in meiotic cells, the Set1 and the Mer2 complexes. Disrupting the Spp1-Set1 interaction mildly decreases H3K4me3 levels and does not affect meiotic recombination initiation. Conversely, the Spp1-Mer2 interaction is required for normal meiotic recombination initiation, but dispensable for Set1 complex-mediated histone H3K4 methylation. Finally, we provide evidence that Spp1 preserves normal H3K4me3 levels independently of the Set1 complex. We propose a model where Spp1 works in three ways to promote recombination initiation: first by depositing histone H3K4 methylation (Set1 complex), next by « reading » and protecting histone H3K4 methylation, and finally by making the link with the chromosome axis (Mer2-Spp1 complex). This work deciphers the precise roles of Spp1 in meiotic recombination and opens perspectives to study its functions in other organisms where H3K4me3 is also present at recombination hotspots.

De Muyt A, Pyatnitskaya A, Andréani J, Ranjha L, Ramus C, Laureau R, Fernandez-Vega A, Holoch D, Girard E, Govin J, Margueron R, Couté Y, Cejka P, Guérois R, Borde V. (2018 Feb 1)

A meiotic XPF-ERCC1-like complex recognizes joint molecule recombination intermediates to promote crossover formation

Genes & Development : DOI : 10.1101/gad.308510.117 En savoir plus

Meiotic crossover formation requires the stabilization of early recombination intermediates by a set of proteins and occurs within the environment of the chromosome axis, a structure important for the regulation of meiotic recombination events. The molecular mechanisms underlying and connecting crossover recombination and axis localization are elusive. Here, we identified the ZZS (Zip2-Zip4-Spo16) complex, required for crossover formation, which carries two distinct activities: one provided by Zip4, which acts as hub through physical interactions with components of the chromosome axis and the crossover machinery, and the other carried by Zip2 and Spo16, which preferentially bind branched DNA molecules in vitro. We found that Zip2 and Spo16 share structural similarities to the structure-specific XPF-ERCC1 nuclease, although it lacks endonuclease activity. The XPF domain of Zip2 is required for crossover formation, suggesting that, together with Spo16, it has a noncatalytic DNA recognition function. Our results suggest that the ZZS complex shepherds recombination intermediates toward crossovers as a dynamic structural module that connects recombination events to the chromosome axis. The identification of the ZZS complex improves our understanding of the various activities required for crossover implementation and is likely applicable to other organisms, including mammals.


Année de publication : 2017

Yann Duroc, Rajeev Kumar, Lepakshi Ranjha, Céline Adam, Raphaël Guérois, Khan Md Muntaz, Marie-Claude Marsolier-Kergoat, Florent Dingli, Raphaëlle Laureau, Damarys Loew, Bertrand Llorente, Jean-Baptiste Charbonnier, Petr Cejka, Valérie Borde (2017 Jan 5)

Concerted action of the MutLβ heterodimer and Mer3 helicase regulates the global extent of meiotic gene conversion.

eLife : DOI : 10.7554/eLife.21900 En savoir plus

Gene conversions resulting from meiotic recombination are critical in shaping genome diversification and evolution. How the extent of gene conversions is regulated is unknown. Here we show that the budding yeast mismatch repair related MutLβ complex, Mlh1-Mlh2, specifically interacts with the conserved meiotic Mer3 helicase, which recruits it to recombination hotspots, independently of mismatch recognition. This recruitment is essential to limit gene conversion tract lengths genome-wide, without affecting crossover formation. Contrary to expectations, Mer3 helicase activity, proposed to extend the displacement loop (D-loop) recombination intermediate, does not influence the length of gene conversion events, revealing non-catalytical roles of Mer3. In addition, both purified Mer3 and MutLβ preferentially recognize D-loops, providing a mechanism for limiting gene conversion in vivo. These findings show that MutLβ is an integral part of a new regulatory step of meiotic recombination, which has implications to prevent rapid allele fixation and hotspot erosion in populations.


Année de publication : 2016

Vijayalakshmi V Subramanian, Amy J MacQueen, Gerben Vader, Miki Shinohara, Aurore Sanchez, Valérie Borde, Akira Shinohara, Andreas Hochwagen (2016 Feb 13)

Chromosome Synapsis Alleviates Mek1-Dependent Suppression of Meiotic DNA Repair.

PLoS biology : e1002369 : DOI : 10.1371/journal.pbio.1002369 En savoir plus

Faithful meiotic chromosome segregation and fertility require meiotic recombination between homologous chromosomes rather than the equally available sister chromatid, a bias that in Saccharomyces cerevisiae depends on the meiotic kinase, Mek1. Mek1 is thought to mediate repair template bias by specifically suppressing sister-directed repair. Instead, we found that when Mek1 persists on closely paired (synapsed) homologues, DNA repair is severely delayed, suggesting that Mek1 suppresses any proximal repair template. Accordingly, Mek1 is excluded from synapsed homologues in wild-type cells. Exclusion requires the AAA+-ATPase Pch2 and is directly coupled to synaptonemal complex assembly. Stage-specific depletion experiments further demonstrate that DNA repair in the context of synapsed homologues requires Rad54, a repair factor inhibited by Mek1. These data indicate that the sister template is distinguished from the homologue primarily by its closer proximity to inhibitory Mek1 activity. We propose that once pairing or synapsis juxtaposes homologues, exclusion of Mek1 is necessary to avoid suppression of all templates and accelerate repair progression.


Année de publication : 2015

Elsa Brachet, Claire Béneut, Maria-Elisabetta Serrentino, Valérie Borde (2015 May 5)

The CAF-1 and Hir Histone Chaperones Associate with Sites of Meiotic Double-Strand Breaks in Budding Yeast.

PloS one : e0125965 : DOI : 10.1371/journal.pone.0125965 En savoir plus

In the meiotic prophase, programmed DNA double-strand breaks (DSB) are introduced along chromosomes to promote homolog pairing and recombination. Although meiotic DSBs usually occur in nucleosome-depleted, accessible regions of chromatin, their repair by homologous recombination takes place in a nucleosomal environment. Nucleosomes may represent an obstacle for the recombination machinery and their timely eviction and reincorporation into chromatin may influence the outcome of recombination, for instance by stabilizing recombination intermediates. Here we show in budding yeast that nucleosomes flanking a meiotic DSB are transiently lost during recombination, and that specific histone H3 chaperones, CAF-1 and Hir, are mobilized at meiotic DSBs. However, the absence of these chaperones has no effect on meiotic recombination, suggesting that timely histone reincorporation following their eviction has no influence on the recombination outcome, or that redundant pathways are activated. This study is the first example of the involvement of histone H3 chaperones at naturally occurring, developmentally programmed DNA double-strand breaks.

Valérie Borde, Bernard de Massy (2015 Mar 26)

Meiosis: early DNA double-strand breaks pave the way for inter-homolog repair.

Developmental cell : 663-4 : DOI : 10.1016/j.devcel.2015.03.011 En savoir plus

During meiotic prophase, the repair of induced DNA double-strand breaks (DSBs) promotes interactions between homologous chromosomes (homologs). A study by Joshi et al. (2015) now highlights how the global DSB activity in a nucleus influences the choice between the homolog and the sister chromatid for DSB repair.

Hardeep Kaur, Arnaud De Muyt, Michael Lichten (2015 Feb 21)

Top3-Rmi1 DNA single-strand decatenase is integral to the formation and resolution of meiotic recombination intermediates.

Molecular cell : 583-94 : DOI : 10.1016/j.molcel.2015.01.020 En savoir plus

The topoisomerase III (Top3)-Rmi1 heterodimer, which catalyzes DNA single-strand passage, forms a conserved complex with the Bloom’s helicase (BLM, Sgs1 in budding yeast). This complex has been proposed to regulate recombination by disassembling double Holliday junctions in a process called dissolution. Top3-Rmi1 has been suggested to act at the end of this process, resolving hemicatenanes produced by earlier BLM/Sgs1 activity. We show here that, to the contrary, Top3-Rmi1 acts in all meiotic recombination functions previously associated with Sgs1, most notably as an early recombination intermediate chaperone, promoting regulated crossover and noncrossover recombination and preventing aberrant recombination intermediate accumulation. In addition, we show that Top3-Rmi1 has important Sgs1-independent functions that ensure complete recombination intermediate resolution and chromosome segregation. These findings indicate that Top3-Rmi1 activity is important throughout recombination to resolve strand crossings that would otherwise impede progression through both early steps of pathway choice and late steps of intermediate resolution.


Année de publication : 2014

Valérie Borde, Michael Lichten (2014 Aug 16)

A timeless but timely connection between replication and recombination.

Cell : 697-8 : DOI : 10.1016/j.cell.2014.07.029 En savoir plus

Initiation of meiotic recombination by DNA double-strand break formation is temporally coordinated with replication. Murakami and Keeney show that this coordination requires recruitment of the Dbf4-dependent kinase to the replication fork by the conserved TIM-TIPIN complex. The same mechanism may regulate other important replication-associated processes.


Année de publication : 2013

Jesús A Carballo, Silvia Panizza, Maria Elisabetta Serrentino, Anthony L Johnson, Marco Geymonat, Valérie Borde, Franz Klein, Rita S Cha (2013 Jul 5)

Budding yeast ATM/ATR control meiotic double-strand break (DSB) levels by down-regulating Rec114, an essential component of the DSB-machinery.

PLoS genetics : e1003545 : DOI : 10.1371/journal.pgen.1003545 En savoir plus

An essential feature of meiosis is Spo11 catalysis of programmed DNA double strand breaks (DSBs). Evidence suggests that the number of DSBs generated per meiosis is genetically determined and that this ability to maintain a pre-determined DSB level, or « DSB homeostasis », might be a property of the meiotic program. Here, we present direct evidence that Rec114, an evolutionarily conserved essential component of the meiotic DSB-machinery, interacts with DSB hotspot DNA, and that Tel1 and Mec1, the budding yeast ATM and ATR, respectively, down-regulate Rec114 upon meiotic DSB formation through phosphorylation. Mimicking constitutive phosphorylation reduces the interaction between Rec114 and DSB hotspot DNA, resulting in a reduction and/or delay in DSB formation. Conversely, a non-phosphorylatable rec114 allele confers a genome-wide increase in both DSB levels and in the interaction between Rec114 and the DSB hotspot DNA. These observations strongly suggest that Tel1 and/or Mec1 phosphorylation of Rec114 following Spo11 catalysis down-regulates DSB formation by limiting the interaction between Rec114 and DSB hotspots. We also present evidence that Ndt80, a meiosis specific transcription factor, contributes to Rec114 degradation, consistent with its requirement for complete cessation of DSB formation. Loss of Rec114 foci from chromatin is associated with homolog synapsis but independent of Ndt80 or Tel1/Mec1 phosphorylation. Taken together, we present evidence for three independent ways of regulating Rec114 activity, which likely contribute to meiotic DSBs-homeostasis in maintaining genetically determined levels of breaks.

Maria-Elisabetta Serrentino, Emmanuel Chaplais, Vérane Sommermeyer, Valérie Borde (2013 Apr 9)

Differential association of the conserved SUMO ligase Zip3 with meiotic double-strand break sites reveals regional variations in the outcome of meiotic recombination.

PLoS genetics : e1003416 : DOI : 10.1371/journal.pgen.1003416 En savoir plus

During the first meiotic prophase, programmed DNA double-strand breaks (DSBs) are distributed non randomly at hotspots along chromosomes, to initiate recombination. In all organisms, more DSBs are formed than crossovers (CO), the repair product that creates a physical link between homologs and allows their correct segregation. It is not known whether all DSB hotspots are also CO hotspots or if the CO/DSB ratio varies with the chromosomal location. Here, we investigated the variations in the CO/DSB ratio by mapping genome-wide the binding sites of the Zip3 protein during budding yeast meiosis. We show that Zip3 associates with DSB sites that are engaged in repair by CO, and Zip3 enrichment at DSBs reflects the DSB tendency to be repaired by CO. Moreover, the relative amount of Zip3 per DSB varies with the chromosomal location, and specific chromosomal features are associated with high or low Zip3 per DSB. This work shows that DSB hotspots are not necessarily CO hotspots and suggests that different categories of DSB sites may fulfill different functions.

Valérie Borde, Bernard de Massy (2013 Apr 2)

Programmed induction of DNA double strand breaks during meiosis: setting up communication between DNA and the chromosome structure.

Current opinion in genetics & development : 147-55 : DOI : 10.1016/j.gde.2012.12.002 En savoir plus

During the first meiotic prophase, hundreds of DNA double strand breaks (DSBs) are deliberately self-inflicted along chromosomes in order to promote homologous recombination between homologs. These DSBs, catalyzed by the evolutionary conserved Spo11 protein, are highly regulated. Recent studies in yeast and mammals have identified key components involved in meiotic DSB formation. In mammals, the DNA binding specificity of PRDM9 determines where DSB occur, whereas in yeast, Spo11 acts in regions which one important feature is chromatin accessibility. However, DSB formation requires additional proteins located on chromosome axes, and the Saccharomyces cerevisiae protein, Spp1 has been recently identified to make the link between axes and DSB sites. These recent findings open exciting routes to understanding how the requirement to regulate DSBs along and between homologs is achieved.

Vérane Sommermeyer, Claire Béneut, Emmanuel Chaplais, Maria Elisabetta Serrentino, Valérie Borde (2013 Jan 10)

Spp1, a member of the Set1 Complex, promotes meiotic DSB formation in promoters by tethering histone H3K4 methylation sites to chromosome axes.

Molecular cell : 43-54 : DOI : 10.1016/j.molcel.2012.11.008 En savoir plus

Meiotic chromosomes are organized into arrays of loops that are anchored to the chromosome axis structure. Programmed DNA double-strand breaks (DSBs) that initiate meiotic recombination, catalyzed by Spo11 and accessory DSB proteins, form in loop sequences in promoters, whereas the DSB proteins are located on chromosome axes. Mechanisms bridging these two chromosomal regions for DSB formation have remained elusive. Here we show that Spp1, a conserved member of the histone H3K4 methyltransferase Set1 complex, is required for normal levels of DSB formation and is associated with chromosome axes during meiosis, where it physically interacts with the Mer2 DSB protein. The PHD finger module of Spp1, which reads H3K4 methylation close to promoters, promotes DSB formation by tethering these regions to chromosome axes and activating cleavage by the DSB proteins. This paper provides the molecular mechanism linking DSB sequences to chromosome axes and explains why H3K4 methylation is important for meiotic recombination.


Année de publication : 2012

Maria-Elisabetta Serrentino, Valérie Borde (2012 Apr 11)

The spatial regulation of meiotic recombination hotspots: are all DSB hotspots crossover hotspots?

Experimental cell research : 1347-52 : DOI : 10.1016/j.yexcr.2012.03.025 En savoir plus

A key step for the success of meiosis is programmed homologous recombination, during which crossovers, or exchange of chromosome arms, take place. Crossovers increase genetic diversity but their main function is to ensure accurate chromosome segregation. Defects in crossover number and position produce aneuploidies that represent the main cause of miscarriages and chromosomal abnormalities such as Down’s syndrome. Recombination is initiated by the formation of programmed double strand breaks (DSBs), which occur preferentially at places called DSB hotspots. Among all DSBs generated, only a small fraction is repaired by crossover, the other being repaired by other homologous recombination pathways. Crossover maps have been generated in a number of organisms, defining crossover hotspots. With the availability of genome-wide maps of DSBs as well as the ability to measure genetically the repair outcome at several hotspots, it is becoming more and more clear that not all DSB hotspots behave the same for crossover formation, suggesting that chromosomal features distinguish different types of hotspots.

Elsa Brachet, Vérane Sommermeyer, Valérie Borde (2012 Feb 23)

Interplay between modifications of chromatin and meiotic recombination hotspots.

Biology of the cell / under the auspices of the European Cell Biology Organization : 51-69 : DOI : 10.1111/boc.201100113 En savoir plus

Meiotic recombination lies at the heart of sexual reproduction. It is essential for producing viable gametes with a normal haploid genomic content and its dysfunctions can be at the source of aneuploidies, such as the Down syndrome, or many genetic disorders. Meiotic recombination also generates genetic diversity that is transmitted to progeny by shuffling maternal and paternal alleles along chromosomes. Recombination takes place at non-random chromosomal sites called ‘hotspots’. Recent evidence has shown that their location is influenced by properties of chromatin. In addition, many studies in somatic cells have highlighted the need for changes in chromatin dynamics to allow the process of recombination. In this review, we discuss how changes in the chromatin landscape may influence the recombination map, and reciprocally, how recombination events may lead to epigenetic modifications at sites of recombination, which could be transmitted to progeny.


Année de publication : 2011

Emmanuelle Martini, Valérie Borde, Matthieu Legendre, Stéphane Audic, Béatrice Regnault, Guillaume Soubigou, Bernard Dujon, Bertrand Llorente (2011 Apr 28)

Genome-wide analysis of heteroduplex DNA in mismatch repair-deficient yeast cells reveals novel properties of meiotic recombination pathways.

PLoS genetics : e1002305 : DOI : 10.1371/journal.pgen.1002305 En savoir plus

Meiotic DNA double-strand breaks (DSBs) initiate crossover (CO) recombination, which is necessary for accurate chromosome segregation, but DSBs may also repair as non-crossovers (NCOs). Multiple recombination pathways with specific intermediates are expected to lead to COs and NCOs. We revisited the mechanisms of meiotic DSB repair and the regulation of CO formation, by conducting a genome-wide analysis of strand-transfer intermediates associated with recombination events. We performed this analysis in a SK1 × S288C Saccharomyces cerevisiae hybrid lacking the mismatch repair (MMR) protein Msh2, to allow efficient detection of heteroduplex DNAs (hDNAs). First, we observed that the anti-recombinogenic activity of MMR is responsible for a 20% drop in CO number, suggesting that in MMR-proficient cells some DSBs are repaired using the sister chromatid as a template when polymorphisms are present. Second, we observed that a large fraction of NCOs were associated with trans-hDNA tracts constrained to a single chromatid. This unexpected finding is compatible with dissolution of double Holliday junctions (dHJs) during repair, and it suggests the existence of a novel control point for CO formation at the level of the dHJ intermediate, in addition to the previously described control point before the dHJ formation step. Finally, we observed that COs are associated with complex hDNA patterns, confirming that the canonical double-strand break repair model is not sufficient to explain the formation of most COs. We propose that multiple factors contribute to the complexity of recombination intermediates. These factors include repair of nicks and double-stranded gaps, template switches between non-sister and sister chromatids, and HJ branch migration. Finally, the good correlation between the strand transfer properties observed in the absence of and in the presence of Msh2 suggests that the intermediates detected in the absence of Msh2 reflect normal intermediates.