Aspects of the regulation of DNA repair by replication induced by a single molecule end

Les cassures double brin de l’ADN (DSBs) sont considérées comme des lésions délétères puisqu'une seule DSB non réparée suffit à entraîner la mort cellulaire. Les DSBs peuvent être réparées par simple religature ou bien par recombinaison homologue (HR) qui copie une molécule d’ADN homologue intacte pour réparer la lésion initiale. Le dysfonctionnement de l'une ou l’autre conduit à une instabilité du génome, qui est une caractéristique de nombreux cancers. Le BIR est une sous-voie de la HR chez les eucaryotes qui vise à réparer les DSBs ne possédant qu’une seule extrémité, comme les cassures provenant des fourches de réplication de l'ADN cassées et des télomères érodés. À la recherche de facteurs agissant en cis et régulant l'efficacité du BIR, nous avons constaté que l'efficacité du BIR est la plus élevée près des extrémités des chromosomes. L'efficacité du BIR en fonction de la longueur de l'ADN à répliquer peut être décrit par une combinaison de deux fonctions exponentielles décroissantes. Cette propriété résulte probablement des cycles répétés d'invasion, d'allongement et de dissociation des brins qui caractérisent le BIR. La processivité apparente du BIR dépend de la longueur de l'ADN déjà synthétisé. Le BIR est plus susceptible d'être perturbé lors de la synthèse des premiers ~60 kb d'ADN que plus tard, notamment lorsque la chromatide matrice est répliquée, transcrite ou hétérochromatique.Nous avons montré que l'hélicase Srs2 favorise le BIR dans les cellules diploïdes et seulement à proximité des télomères dans les cellules haploïdes. Dans l'ensemble, nous apportons un nouvel éclairage sur la façon dont une voie de réparation de l'ADN de dernier recours est régulée.DNA double strand breaks (DSB) are considered particularly deleterious lesions since one unrepaired DSB is sufficient to cause cell death. DSBs can be repaired by simple religation or by homologous recombination (HR) which copies an intact homologous DNA molecule to repair the initial lesion. Dysfunction of either of these two pathways leads to genome instability, which is a hallmark of many cancers. Here, the objective is to better understand the mechanism of break-induced replication (BIR) and its regulation. BIR is a sub-pathway of HR in eukaryotes that aims to repair DSBs with only one end, such as breaks from broken DNA replication forks and eroded telomeres. Looking for cis-acting factors that regulate the efficiency of BIR along the chromosomal arms of baker's yeast, we found that the efficiency of BIR is highest near the ends of chromosomes. In addition, we have found that the efficiency of BIR as a function of the length of the DNA to be replicated can be described by a combination of two decreasing exponential functions. This property probably results from the repeated cycles of invasion, elongation and dissociation of the strands which characterize BIR. Interestingly, the apparent processivity of BIR depends on the length of the DNA already synthesized. BIR is more likely to be disrupted during the synthesis of the first ~ 60 kb of DNA than later, especially when the template chromatid is replicated, transcribed, or heterochromatic. Finally, we have shown that the Srs2 helicase promotes BIR in diploid cells and only near telomeres in haploid cells. Overall, we shed new light on how a last resort DNA repair pathway is regulated

Comprehensive analysis of cis- and trans-acting factors affecting ectopic Break-Induced Replication

International audienceBreak-induced replication (BIR) is a highly mutagenic eukaryotic homologous DNA recombination pathway that repairs one-ended DNA double strand breaks such as broken DNA replication forks and eroded telomeres. While searching for cis-acting factors regulating ectopic BIR efficiency, we found that ectopic BIR efficiency is the highest close to chromosome ends. The variations of ectopic BIR efficiency as a function of the length of DNA to replicate can be described as a combination of two decreasing exponential functions, a property in line with repeated cycles of strand invasion, elongation and dissociation that characterize BIR. Interestingly, the apparent processivity of ectopic BIR depends on the length of DNA already synthesized. Ectopic BIR is more susceptible to disruption during the synthesis of the first ~35–40 kb of DNA than later, notably when the template chromatid is being transcribed or heterochromatic. Finally, we show that the Srs2 helicase promotes ectopic BIR from both telomere proximal and telomere distal regions in diploid cells but only from telomere proximal sites in haploid cells. Altogether, we bring new light on the factors impacting a last resort DNA repair pathway