Sister Chromatids Are Preferred over Homologs as Substrates for Recombinational Repair in Saccharomyces Cerevisiae

A diploid Saccharomyces cerevisiae strain was constructed in which the products of both homolog recombination and unequal sister chromatid recombination events could be selected. This strain was synchronized in G1 or in G2, irradiated with X-rays to induce DNA damage, and monitored for levels of recombination. Cells irradiated in G1 were found to repair recombinogenic damage primarily by homolog recombination, whereas those irradiated in G2 repaired such damage preferentially by sister chromatid recombination. We found, as have others, that G1 diploids were much more sensitive to the lethal effects of X-ray damage than were G2 diploids, especially at higher doses of irradiation. The following possible explanations for this observation were tested: G2 cells have more potential templates for repair than G1 cells; G2 cells are protected by the RAD9-mediated delay in G2 following DNA damage; sister chromatids may share more homology than homologous chromosomes. All these possibilities were ruled out by appropriate tests. We propose that, due to a special relationship they share, sister chromatids are not only preferred over homologous chromatids as substrates for recombinational repair, but have the capacity to repair more DNA damage than do homologs.     

Sister chromatid cohesion is required for postreplicative double-strand break repair in Saccharomyces cerevisiae.

The repair of DNA double-strand breaks by recombination requires the presence of an undamaged copy that is used as a template during the repair process. Because cells acquire resistance to gamma irradiation during DNA replication and because sister chromatids are the preferred partner for double-strand break repair in mitotic diploid yeast cells, it has long been suspected that cohesion between sister chromatids might be crucial for efficient repair. This hypothesis is consistent with the sensitivity to gamma irradiation of mutants defective in the cohesin complex that holds sister chromatids together from DNA replication until the onset of anaphase (reviewed in) . It is also in accordance with the finding that surveillance mechanisms (checkpoints) that sense DNA damage arrest cell cycle progression in yeast by causing stabilization of the securin Pds1, thereby blocking sister chromatid separation. The hypersensitivity to irradiation of cohesin mutants could, however, be due to a more direct involvement of the cohesin complex in the process of DNA repair. We show here that passage through S phase in the presence of cohesin, and not cohesin per se, is essential for efficient double-strand break repair during G2 in yeast. Proteins needed to load cohesin onto chromosomes (Scc2) and to generate cohesion during S phase (Eco1) are also shown to be required for repair. Our results confirm what has long been suspected but never proven, that cohesion between sister chromatids is essential for efficient double-strand break repair in mitotic cells.     

Sister Chromatid Cohesion

During S phase, not only does DNA have to be replicated, but also newly synthesized DNA molecules have to be connected with each other. This sister chromatid cohesion is essential for the biorientation of chromosomes on the mitotic or meiotic spindle, and is thus an essential prerequisite for chromosome segregation. Cohesion is mediated by cohesin complexes that are thought to embrace sister chromatids as large rings. Cohesin binds to DNA dynamically before DNA replication and is converted into a stably DNA-bound form during replication. This conversion requires acetylation of cohesin, which in vertebrates leads to recruitment of sororin. Sororin antagonizes Wapl, a protein that is able to release cohesin from DNA, presumably by opening the cohesin ring. Inhibition of Wapl by sororin therefore “locks” cohesin rings on DNA and allows them to maintain cohesion for long periods of time in mammalian oocytes, possibly for months or even years.DNA replication during the synthesis (S) phase generates identical DNA molecules, which, in their chromatinized form, are called sister chromatids. The pairs of sister chromatids remain united as part of one chromosome during the subsequent gap (G₂) phase and during early mitosis, in prophase, prometaphase, and metaphase. During these stages of mitosis chromosomes condense, in most eukaryotes the nuclear envelope breaks down, and in all species chromosomes are ultimately attached to both poles of the mitotic spindle. Only once this biorientation has been achieved for all chromosomes, the sister chromatids are separated from each other in anaphase and transported toward opposite spindle poles of the mother cell, enabling its subsequent division into two genetically identical daughter cells.This series of events critically depends on the fact that sister chromatids remain physically connected with each other from S phase until metaphase. This physical connection, called sister chromatid cohesion, opposes the pulling forces that are generated by microtubules that attach to kinetochores and thereby enables the biorientation of chromosomes on the mitotic spindle (Tanaka et al. 2000b). Without cohesion, sister chromatids could therefore not be segregated symmetrically between the forming daughter cells, resulting in aneuploidy. For the same reasons, cohesion is essential for chromosome segregation in meiosis I and meiosis II. Cohesion defects in human oocytes can lead to aneuploidy, which is thought to be the major cause of spontaneous abortion, because only a few types of aneuploidy are compatible with viability, such as trisomy 21 (Down syndrome), trisomy 18 (Edwards syndrome), and trisomy 13 (Patau syndrome) (Hunt and Hassold 2010). Studying the mechanisms of cohesion is therefore essential for understanding how the genome is passed properly from one cell generation to the next.In addition, sister chromatid cohesion facilitates the repair of DNA double-strand breaks in cells that have replicated their DNA, where such breaks can be repaired by a homologous recombination mechanism that uses the undamaged sister chromatid as a template (for review, see Watrin et al. 2006). Furthermore, mutations in the proteins that are required for sister chromatid cohesion can cause defects in chromatin structure and gene regulation, and can in rare cases lead to congenital developmental disorders, called Cornelia de Lange syndrome, Roberts/SC Phocomelia syndrome, and Warsaw Breakage syndrome (for review, see Mannini et al. 2010).     

Recombinational repair of radiation-induced double-strand breaks occurs in the absence of extensive resection

Recombinational repair provides accurate chromosomal restitution after double-strand break (DSB) induction. While all DSB recombination repair models include 5′-3′ resection, there are no studies that directly assess the resection needed for repair between sister chromatids in G-2 arrested cells of random, radiation-induced ‘dirty’ DSBs. Using our Pulse Field Gel Electrophoresis-shift approach, we determined resection at IR-DSBs in WT and mutants lacking exonuclease1 or Sgs1 helicase. Lack of either reduced resection length by half, without decreased DSB repair or survival. In the exo1Δ sgs1Δ double mutant, resection was barely detectable, yet it only took an additional hour to achieve a level of repair comparable to WT and there was only a 2-fold dose-modifying effect on survival. Results with a Dnl4 deletion strain showed that remaining repair was not due to endjoining. Thus, similar to what has been shown for a single, clean HO-induced DSB, a severe reduction in resection tract length has only a modest effect on repair of multiple, dirty DSBs in G2-arrested cells. Significantly, this study provides the first opportunity to directly relate resection length at DSBs to the capability for global recombination repair between sister chromatids.     

Postreplicative recruitment of cohesin to double-strand breaks is required for DNA repair

Chromosome stability depends on accurate chromosome segregation and efficient DNA double-strand break (DSB) repair. Sister chromatid cohesion, established during S phase by the protein complex cohesin, is central to both processes. In the absence of cohesion, chromosomes missegregate and G2-phase DSB repair fails. Here, we demonstrate that G2-phase repair also requires the presence of cohesin at the damage site. Cohesin components are shown to be recruited to extended chromosome regions surrounding DNA breaks induced during G2. We find that in the absence of functional cohesin-loading proteins (Scc2/Scc4), the accumulation of cohesin at DSBs is abolished and repair is defective, even though sister chromatids are connected by S phase generated cohesion. Evidence is also provided that DSB induction elicits establishment of sister chromatid cohesion in G2, implicating that damage-recruited cohesin facilitates DNA repair by tethering chromatids.     

Cell-cycle dependent changes in sensitivity to γ-rays in synchronously dividing yeast culture

Changes in survival of yeast cells following γ-irradiation at different stages of the cell cycle were studied using a well synchronized culture. Maximum radioresistance occurs at the end of the S phase. Maximum radiosensitivity is observed just before entry into the S phase. The high degree of synchrony obtained allows more precise measurement of the extent of survival changes than has been achieved until now with partially synchronized cultures. Indeed, after a 60 krad irradiation we find a 100 % survival for cells which have just finished the S phase of the first cell cycle, against a 2 % survival for cells which are ready to enter the S phase of the second cell cycle. As the culture desynchronizes through successive cell cycles we have been able to follow the way in which survival curves are modified. We can extrapolate that with a perfectly synchronized culture the survival of ‘early S’ cells to a 60 krad irradiation would not be 2 % but 0.01 %. The high radioresistance observed at the end of S phase can hardly be explained simply in terms of DNA target or accumulation of radioprotectors. More likely the end of the S phase is a favourable stage for repair processes, at which time two genomes are able to recombine.     

Cohesin promotes the repair of ionizing radiation-induced DNA double-strand breaks in replicated chromatin

The cohesin protein complex holds sister chromatids together after synthesis until mitosis. It also contributes to post-replicative DNA repair in yeast and higher eukaryotes and accumulates at sites of laser-induced damage in human cells. Our goal was to determine whether the cohesin subunits SMC1 and Rad21 contribute to DNA double-strand break repair in X-irradiated human cells in the G2 phase of the cell cycle. RNA interference-mediated depletion of SMC1 sensitized HeLa cells to X-rays. Repair of radiation-induced DNA double-strand breaks, measured by γH2AX/53BP1 foci analysis, was slower in SMC1- or Rad21-depleted cells than in controls in G2 but not in G1. Inhibition of the DNA damage kinase DNA-PK, but not ATM, further inhibited foci loss in cohesin-depleted cells in G2. SMC1 depletion had no effect on DNA single-strand break repair in either G1 or late S/G2. Rad21 and SMC1 were recruited to sites of X-ray-induced DNA damage in G2-phase cells, but not in G1, and only when DNA damage was concentrated in subnuclear stripes, generated by partially shielded ultrasoft X-rays. Our results suggest that the cohesin complex contributes to cell survival by promoting the repair of radiation-induced DNA double-strand breaks in G2-phase cells in an ATM-dependent pathway.     

Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells

The repair of DNA double-strand breaks (DSBs) by homologous recombinational repair (HRR) underlies the high radioresistance and low mutability observed in S-phase mammalian cells. To evaluate the contributions of HRR and non-homologous end-joining (NHEJ) to overall DSB repair capacity throughout the cell cycle after γ-irradiation, we compared HRR-deficient RAD51D-knockout 51D1 to CgRAD51D-complemented 51D1 (51D1.3) CHO cells for survival and chromosomal aberrations (CAs). Asynchronous cultures were irradiated with 150 or 300 cGy and separated by cell size using centrifugal elutriation. Cell survival of each synchronous fraction (～20 fractions total from early G1 to late G2/M) was measured by colony formation. 51D1.3 cells were most resistant in S, while 51D1 cells were most resistant in early G1 (with survival and chromosome-type CA levels similar to 51D1.3) and became progressively more sensitive throughout S and G2. Both cell lines experienced significantly reduced survival from late S into G2. Metaphases were collected from every third elutriation fraction at the first post-irradiation mitosis and scored for CAs. 51D1 cells irradiated in S and G2 had ～2-fold higher chromatid-type CAs and a remarkable ～25-fold higher level of complex chromatid-type exchanges compared to 51D1.3 cells. Complex exchanges in 51D1.3 cells were only observed in G2. These results show an essential role for HRR in preventing gross chromosomal rearrangements in proliferating cells and, with our previous report of reduced survival of G2-phase NHEJ-deficient prkdc CHO cells [Hinz et al., DNA Repair 4, 782–792, 2005], imply reduced activity/efficiency of both HRR and NHEJ as cells transition from S to G2.     

Pathways of DNA double-strand break repair during the mammalian cell cycle

Little is known about the quantitative contributions of nonhomologous end joining (NHEJ) and homologous recombination (HR) to DNA double-strand break (DSB) repair in different cell cycle phases after physiologically relevant doses of ionizing radiation. Using immunofluorescence detection of gamma-H2AX nuclear foci as a novel approach for monitoring the repair of DSBs, we show here that NHEJ-defective hamster cells (CHO mutant V3 cells) have strongly reduced repair in all cell cycle phases after 1 Gy of irradiation. In contrast, HR-defective CHO irs1SF cells have a minor repair defect in G(1), greater impairment in S, and a substantial defect in late S/G(2). Furthermore, the radiosensitivity of irs1SF cells is slight in G(1) but dramatically higher in late S/G(2), while V3 cells show high sensitivity throughout the cell cycle. These findings show that NHEJ is important in all cell cycle phases, while HR is particularly important in late S/G(2), where both pathways contribute to repair and radioresistance. In contrast to DSBs produced by ionizing radiation, DSBs produced by the replication inhibitor aphidicolin are repaired entirely by HR. irs1SF, but not V3, cells show hypersensitivity to aphidicolin treatment. These data provide the first evaluation of the cell cycle-specific contributions of NHEJ and HR to the repair of radiation-induced versus replication-associated DSBs.     

Is there a relationship between hypoxia,contact resistance,and intercellular communication?

Summary This investigation addresses the shape of radiation survival curves of cells cultured as multicell spheroids. It is shown that spheroids of cells capable of intercellular communication by gap-junctions display survival curves lacking a radioresistant fraction of hypoxic cells. Compared to the corresponding monolayers, these spheroid survival curves exhibit a uniform increase in radioresistance due to the contact effect. In contrast, biphasic survival curves indicative of hypoxic cells are obtained with non-communicating spheroids, however, without indication of a contact effect. Evidence is presented that this relationship between intercellular communication, hypoxia, and contact effect may possibly also hold for survival curves of solid tumors.     

Genetic analysis of the DNA-dependent protein kinase reveals an inhibitory role of Ku in late S-G2 phase DNA double-strand break repair

Two major complementary double-strand break (DSB) repair pathways exist in vertebrates, homologous recombination (HR), which involves Rad54, and non-homologous end-joining, which requires the DNA-dependent protein kinase (DNA-PK). DNA-PK comprises a catalytic subunit (DNA-PKcs) and a DNA-binding Ku70 and Ku80 heterodimer. To define the activities of individual DNA-PK components in DSB repair, we targeted the DNA-PKcs gene in chicken DT40 cells. DNA-PKcs deficiency caused a DSB repair defect that was, unexpectedly, suppressed by KU70 disruption. We have shown previously that genetic ablation of Ku70 confers RAD54-dependent radioresistance on S-G(2) phase cells, when sister chromatids are available for HR repair. To test whether direct interference by Ku70 with HR might explain the Ku70(-/-)/DNA-PKcs(-/-/-) radioresistance, we monitored HR activities directly in Ku- and DNA-PKcs-deficient cells. The frequency of intrachromosomal HR induced by the I-SceI restriction enzyme was increased in the absence of Ku but not of DNA-PKcs. Significantly, abrogation of HR activity by targeting RAD54 in Ku70(-/-) or DNA-PKcs(-/-/-) cells caused extreme radiosensitivity, suggesting that the relative radioresistance seen with loss of Ku70 was because of HR-dependent repair pathways. Our findings suggest that Ku can interfere with HR-mediated DSB repair, perhaps competing with HR for DSB recognition.     

Genetic Control of Mitosis: Mutation ff3Causes Premature Beginning of Telophase Chromatin Reorganization in Drosophila melanogaster

The effect of cell cycle mutation ff3 on chromosome segregation was studied on fixed cells of neural ganglia of Drosophila melanogasterlarvae. The cell distributions by diameter of interphase nuclei and by distance between sister chromatid sets were compared at anaphase and telophase. In the control wild-type strain Lausenne, the cell distribution by distance between sister chromatids in anaphase was similar to their distribution by nuclear size. The mean distance between segregating chromatids at anaphase (l _av) coincided with the mean diameter of interphase nuclei (d _av) and was 8.3 m. Cells passed to telophase when chromatids were at least 10 m apart. The mutant ff3 strain differed from the control strain Lausenne in cell distribution by interphase nuclear diameter and distance between sister chromatids in anaphase; the mean nuclear diameter and mean distance between segregating chromatids similarly increased to 9.3 m. A specific feature of mitosis in mutant strain ff3 was a premature beginning of telophase chromatin reorganization. This caused the occurrence of cells with abnormally short (less then the interphase nuclear diameter) distance between sister chromatid sets in telophase but not in anaphase, as if these cells had passed from anaphase to telophase prematurely, during the chromatid movement toward poles in anaphase A.     

The CDK regulates repair of double-strand breaks by homologous recombination during the cell cycle

DNA double-strand breaks (DSBs) are dangerous lesions that can lead to genomic instability and cell death. Eukaryotic cells repair DSBs either by nonhomologous end-joining (NHEJ) or by homologous recombination. We investigated the ability of yeast cells (Saccharomyces cerevisiae) to repair a single, chromosomal DSB by recombination at different stages of the cell cycle. We show that cells arrested at the G1 phase of the cell cycle restrict homologous recombination, but are able to repair the DSB by NHEJ. Furthermore, we demonstrate that recombination ability does not require duplicated chromatids or passage through S phase, and is controlled at the resection step by Clb-CDK activity.     

Actinomycin D effects on mitosis and chromosomes: sticky chromatids and localized lesions

When Indian muntjac and Chinese hamster cells in culture were treated with Actinomycin D (1 g/ml) for 1–2 hours, the sister chromatids, especially the distal segments, appeared to have difficulty separating in anaphase. The separated proximal segments progressively became stretched. The nucleolus organizer regions seemed to be most susceptible to stretching, and breaks in these regions were frequently observed. Electron microscopic observations showed that the sticky chromatids (and less frequently sticky chromosomes) contain connecting submicroscopic chromosome strands. When the treated cells were allowed to grow in a drug-free medium for several days, a high frequency of endoreduplicated mitotic figures was found. Chromosome and chromatid breaks and other aberrations were common, mainly localized at G band negative areas particularly nucleolus organizer regions.     

Shugoshin1 may play important roles in separation of homologous chromosomes and sister chromatids during mouse oocyte meiosis

Background

Homologous chromosomes separate in meiosis I and sister chromatids separate in meiosis II, generating haploid gametes. To address the question why sister chromatids do not separate in meiosis I, we explored the roles of Shogoshin1 (Sgo1) in chromosome separation during oocyte meiosis.

Methodology/Principal Findings

Sgo1 function was evaluated by exogenous overexpression to enhance its roles and RNAi to suppress its roles during two meioses of mouse oocytes. Immunocytochemistry and chromosome spread were used to evaluate phenotypes. The exogenous Sgo1 overexpression kept homologous chromosomes and sister chromatids not to separate in meiosis I and meiosis II, respectively, while the Sgo1 RNAi promoted premature separation of sister chromatids.

Conclusions

Our results reveal that prevention of premature separation of sister chromatids in meiosis I requires the retention of centromeric Sgo1, while normal separation of sister chromatids in meiosis II requires loss of centromeric Sgo1.     

The level of induced DNA double-strand breaks does not correlate with cell killing in X-irradiated mitotic and G1-phase CHO cells

The neutral (pH 9.6) filter elution technique was used to evaluate DNA damage induced in CHO cells irradiated at mitosis or in G1-phase under various incubation and postirradiation treatment conditions. Mitotic and G1/S border cells were more sensitive to radiation than G1 cells with respect to cell killing, but showed similar (G1/S) or lower (M) DNA elution dose--response curves. Similar cell survival and DNA/elution dose--response curves were obtained with plateau-phase cultures containing mainly G1-cells, as well as with G1 cells obtained after division of mitotic cells in either fresh or conditioned medium. However, survival of plateau-phase cells could be modified substantially by delayed-plating or postirradiation treatment with araA. These results, together with previously published observations, indicate that induction of DNA dsb cannot be invoked as an explanation for the variations in radiosensitivity observed through the cycle, or as an explanation for the formation of the survival curve shoulder. It is proposed that repair and fixation of radiation-induced DNA damage, expressed at the cell survival level as repair and fixation of alpha-PLD, are responsible for these effects.     

Cohesin Is Limiting for the Suppression of DNA Damage–Induced Recombination between Homologous Chromosomes

Double-strand break (DSB) repair through homologous recombination (HR) is an evolutionarily conserved process that is generally error-free. The risk to genome stability posed by nonallelic recombination or loss-of-heterozygosity could be reduced by confining HR to sister chromatids, thereby preventing recombination between homologous chromosomes. Here we show that the sister chromatid cohesion complex (cohesin) is a limiting factor in the control of DSB repair and genome stability and that it suppresses DNA damage–induced interactions between homologues. We developed a gene dosage system in tetraploid yeast to address limitations on various essential components in DSB repair and HR. Unlike RAD50 and RAD51, which play a direct role in HR, a 4-fold reduction in the number of essential MCD1 sister chromatid cohesion subunit genes affected survival of gamma-irradiated G₂/M cells. The decreased survival reflected a reduction in DSB repair. Importantly, HR between homologous chromosomes was strongly increased by ionizing radiation in G₂/M cells with a single copy of MCD1 or SMC3 even at radiation doses where survival was high and DSB repair was efficient. The increased recombination also extended to nonlethal doses of UV, which did not induce DSBs. The DNA damage–induced recombinants in G₂/M cells included crossovers. Thus, the cohesin complex has a dual role in protecting chromosome integrity: it promotes DSB repair and recombination between sister chromatids, and it suppresses damage-induced recombination between homologues. The effects of limited amounts of Mcd1and Smc3 indicate that small changes in cohesin levels may increase the risk of genome instability, which may lead to genetic diseases and cancer.     

Gene conversion at different points in the mitotic cycle of Saccharomyces cerevisiae

Summary In the Saccharomyces cerevisiae mitotic cycle, the timing of radiation-induced gene conversion has been studied using thermosensitive cell division cycle mutants. The cells were found to perform conversion at different G1 or post-replication steps. A lower yield in induction is found during the G2 phase and is explained by the competition for recombinational repair between sister chromatids and homologous chromosomes. The results are discussed in relation to repair.     

Human Wapl is a cohesin-binding protein that promotes sister-chromatid resolution in mitotic prophase

BACKGROUND: The linkage between duplicated chromosomes (sister chromatids) is established during S phase by the action of cohesin, a multisubunit complex conserved from yeast to humans. Most cohesin dissociates from chromosome arms when the cell enters mitotic prophase, leading to the formation of metaphase chromosomes with two cytologically discernible chromatids. This process is known as sister-chromatid resolution. Although two mitotic kinases have been implicated in this process, it remains unknown exactly how the cohesin-mediated linkage is destabilized at a mechanistic level. RESULTS: The wings apart-like (Wapl) protein was originally identified as a gene product that potentially regulates heterochromatin organization in Drosophila melanogaster. We show that the human ortholog of Wapl is a cohesin-binding protein that facilitates cohesin's timely release from chromosome arms during prophase. Depletion of Wapl from HeLa cells causes transient accumulation of prometaphase-like cells with chromosomes that display poorly resolved sister chromatids with a high level of cohesin. Reduction of cohesin relieves the Wapl-depletion phenotype, and depletion of Wapl rescues premature sister separation observed in Sgo1-depleted or Esco2-depleted cells. Conversely, overexpression of Wapl causes premature separation of sister chromatids. Wapl physically associates with cohesin in HeLa-cell nuclear extracts. Remarkably, in vitro reconstitution experiments demonstrate that Wapl forms a stoichiometric, ternary complex with two regulatory subunits of cohesin, implicating its noncatalytic function in inactivating cohesin's ability to interact with chromatin. CONCLUSIONS: Wapl is a new regulator of sister chromatid resolution and promotes release of cohesin from chromosomes by directly interacting with its regulatory subunits.     

The STRUCTURAL MAINTENANCE OF CHROMOSOMES 5/6 Complex Promotes Sister Chromatid Alignment and Homologous Recombination after DNA Damage in Arabidopsis thaliana

Sister chromatids are often arranged as incompletely aligned entities in interphase nuclei of Arabidopsis thaliana. The STRUCTURAL MAINTENANCE OF CHROMOSOMES (SMC) 5/6 complex, together with cohesin, is involved in double-strand break (DSB) repair by sister chromatid recombination in yeasts and mammals. Here, we analyzed the function of genes in Arabidopsis. The wild-type allele of SMC5 is essential for seed development. Each of the two SMC6 homologs of Arabidopsis is required for efficient repair of DNA breakage via intermolecular homologous recombination in somatic cells. Alignment of sister chromatids is enhanced transiently after X-irradiation (and mitomycin C treatment) in wild-type nuclei. In the smc5/6 mutants, the x-ray–mediated increase in sister chromatid alignment is much lower and delayed. The reduced S phase–established cohesion caused by a knockout mutation in one of the α-kleisin genes, SYN1, also perturbed enhancement of sister chromatid alignment after irradiation, suggesting that the S phase–established cohesion is a prerequisite for correct DSB-dependent cohesion. The radiation-sensitive51 mutant, deficient in heteroduplex formation during DSB repair, showed wild-type frequencies of sister chromatid alignment after X-irradiation, implying that the irradiation-mediated increase in sister chromatid alignment is a prerequisite for, rather than a consequence of, DNA strand exchange between sister chromatids. Our results suggest that the SMC5/6 complex promotes sister chromatid cohesion after DNA breakage and facilitates homologous recombination between sister chromatids.