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The three-dimensional organization of the genome supports regulated gene expression, recombination, DNA repair, and chromosome segregation during mitosis. Chromosome conformation capture (Hi-C) 1 , 2 analysis has revealed a complex genomic landscape of internal chromosomal structures in vertebrate cells 3 – 7 , but the identical sequence of sister chromatids has made it difficult to determine how they topologically interact in replicated chromosomes. Here we describe sister-chromatid-sensitive Hi-C (scsHi-C), which is based on labelling of nascent DNA with 4-thio-thymidine and nucleoside conversion chemistry. Genome-wide conformation maps of human chromosomes reveal that sister-chromatid pairs interact most frequently at the boundaries of topologically associating domains (TADs). Continuous loading of a dynamic cohesin pool separates sister-chromatid pairs inside TADs and is required to focus sister-chromatid contacts at TAD boundaries. We identified a subset of TADs that are overall highly paired and are characterized by facultative heterochromatin and insulated topological domains that form separately within individual sister chromatids. The rich pattern of sister-chromatid topologies and our scsHi-C technology will make it possible to investigate how physical interactions between identical DNA molecules contribute to DNA repair, gene expression, chromosome segregation, and potentially other biological processes. Modified chromosome conformation capture (Hi-C) technology is used to characterize the interactions between sister chromatids, despite their identical DNA sequences.

CYREN is a direct inhibitor of classical non-homologous end joining that promotes error-free repair by homologous recombination during the S and G2 phases of the cell cycle. Ku succumbs to a CYREN call A broken DNA molecule can be repaired in two distinct ways: homologous recombination, or classical non-homologous end joining (cNHEJ). Both processes function in the S and G2 phases of the cell cycle, but why cNHEJ is not always used is unknown. Jan Karlseder and colleagues have found that cNHEJ is suppressed in S and G2 by CYREN (cell cycle regulator of NHEJ), which binds to the Ku complex to inhibit the initial steps of end joining. The absence of CYREN results in genomic instability. CYREN thus biases repair in S and G2, when sister chromatids are available, to a recombination process that, unlike cNHEJ, is error-free. Classical non-homologous end joining 1 (cNHEJ) and homologous recombination 2 compete for the repair of double-stranded DNA breaks during the cell cycle. Homologous recombination is inhibited during the G1 phase of the cell cycle, but both pathways are active in the S and G2 phases. However, it is unclear why cNHEJ does not always outcompete homologous recombination during the S and G2 phases. Here we show that CYREN (cell cycle regulator of NHEJ) is a cell-cycle-specific inhibitor of cNHEJ. Suppression of CYREN allows cNHEJ to occur at telomeres and intrachromosomal breaks during the S and G2 phases, and cells lacking CYREN accumulate chromosomal aberrations upon damage induction, specifically outside the G1 phase. CYREN acts by binding to the Ku70/80 heterodimer and preferentially inhibits cNHEJ at breaks with overhangs by protecting them. We therefore propose that CYREN is a direct cell-cycle-dependent inhibitor of cNHEJ that promotes error-free repair by homologous recombination during cell cycle phases when sister chromatids are present.

Sister chromatids are tethered together by the cohesin complex from the time they are made until their separation at anaphase. The ability of cohesin to tether sister chromatids together depends on acetylation of its Smc3 subunit by members of the Eco1 family of cohesin acetyltransferases. Vertebrates express two orthologs of Eco1, called Esco1 and Esco2, both of which are capable of modifying Smc3, but their relative contributions to sister chromatid cohesion are unknown. We therefore set out to determine the precise contributions of Esco1 and Esco2 to cohesion in vertebrate cells. Herewe show that cohesion establishment is critically dependent upon Esco2. Although most Smc3 acetylation is Esco1 dependent, inactivation of the ESCO1 gene has little effect on mitotic cohesion. The unique ability of Esco2 to promote cohesion is mediated by sequences in the N terminus of the protein. We propose that Esco1-dependent modification of Smc3 regulates almost exclusively the noncohesive activities of cohesin, such as DNA repair, transcriptional control, chromosome loop formation, and/or stabilization. Collectively, our data indicate that Esco1 and Esco2 contribute to distinct and separable activities of cohesin in vertebrate cells.

Genotoxic DNA double-strand breaks (DSBs) can be repaired by error-free homologous recombination (HR) or mutagenic non-homologous end-joining 1 . HR supresses tumorigenesis 1 , but is restricted to the S and G2 phases of the cell cycle when a sister chromatid is present 2 . Breast cancer type 1 susceptibility protein (BRCA1) promotes HR by antagonizing the anti-resection factor TP53-binding protein 1(53BP1) (refs. 2 , 3 , 4 – 5 ), but it remains unknown how BRCA1 function is limited to the S and G2 phases. We show that BRCA1 recruitment requires recognition of histone H4 unmethylated at lysine 20 (H4K20me0), linking DSB repair pathway choice directly to sister chromatid availability. We identify the ankyrin repeat domain of BRCA1-associated RING domain protein 1 (BARD1)—the obligate BRCA1 binding partner 3 —as a reader of H4K20me0 present on new histones in post-replicative chromatin 6 . BARD1 ankyrin repeat domain mutations disabling H4K20me0 recognition abrogate accumulation of BRCA1 at DSBs, causing aberrant build-up of 53BP1, and allowing anti-resection activity to prevail in S and G2. Consequently, BARD1 recognition of H4K20me0 is required for HR and resistance to poly (ADP-ribose) polymerase inhibitors. Collectively, this reveals that BRCA1–BARD1 monitors the replicative state of the genome to oppose 53BP1 function, routing only DSBs within sister chromatids to HR. Nakamura et al. show that DNA repair pathway choice and initiation of homologous recombination is guided by the recruitment of BRCA1 to post-replicative chromatin by BARD1 recognition of histone H4 tails unmethylated on lysine 20.

The ring-like cohesin complex mediates sister-chromatid cohesion by encircling pairs of sister chromatids. Cohesin also extrudes loops along chromatids. Whether the two activities involve similar mechanisms of DNA engagement is not known. We implemented an experimental approach based on isolated nuclei carrying engineered cleavable RAD21 proteins to precisely control cohesin ring integrity so that its role in chromatin looping could be studied under defined experimental conditions. This approach allowed us to identify cohesin complexes with distinct biochemical, and possibly structural, properties that mediate different sets of chromatin loops. When RAD21 is cleaved and the cohesin ring is opened, cohesin complexes at CTCF sites are released from DNA and loops at these elements are lost. In contrast, cohesin-dependent loops within chromatin domains that are not anchored at pairs of CTCF sites are more resistant to RAD21 cleavage. The results show that the cohesin complex mediates loops in different ways depending on the genomic context and suggests that it undergoes structural changes as it dynamically extrudes and encounters CTCF sites. Liu and Dekker test the importance of cohesin ring integrity for genome architecture: cohesin ring opening via Rad21 cleavage causes loss of CTCF–CTCF loops but maintains dynamic intra-domain loops, suggesting distinct cohesin engagement modes.

Cell division is a crucial process, and one of its essential steps involves copying the genetic material, which is organized into structures called chromosomes. Before a cell can divide into two, it needs to ensure that each newly copied chromosome is paired tightly with its identical twin. This pairing is maintained by a protein complex known as cohesin, which is conserved in various organisms, from single-celled ones to humans. Cohesin essentially encircles the DNA, creating a ring-like structure to handcuff, to keep the newly synthesized sister chromosomes together in pairs. Therefore, chromosomal cohesion and separation are fundamental processes governing the attachment and segregation of sister chromatids during cell division. Metaphase-to-anaphase transition requires dissolution of cohesins by the enzyme Separase. The tight regulation of these processes is vital for safeguarding genomic stability. Dysregulation in chromosomal cohesion and separation resulting in aneuploidy, a condition characterized by an abnormal chromosome count in a cell, is strongly associated with cancer. Aneuploidy is a recurring hallmark in many cancer types, and abnormalities in chromosomal cohesion and separation have been identified as significant contributors to various cancers, such as acute myeloid leukemia, myelodysplastic syndrome, colorectal, bladder, and other solid cancers. Mutations within the cohesin complex have been associated with these cancers, as they interfere with chromosomal segregation, genome organization, and gene expression, promoting aneuploidy and contributing to the initiation of malignancy. In summary, chromosomal cohesion and separation processes play a pivotal role in preserving genomic stability, and aberrations in these mechanisms can lead to aneuploidy and cancer. Gaining a deeper understanding of the molecular intricacies of chromosomal cohesion and separation offers promising prospects for the development of innovative therapeutic approaches in the battle against cancer.

The mechanism by which chromatids and chromosomes are segregated during mitosis and meiosis is a major puzzle of biology and biophysics. Using polymer simulations of chromosome dynamics, we show that a single mechanism of loop extrusion by condensins can robustly compact, segregate and disentangle chromosomes, arriving at individualized chromatids with morphology observed in vivo. Our model resolves the paradox of topological simplification concomitant with chromosome 'condensation', and explains how enzymes a few nanometers in size are able to control chromosome geometry and topology at micron length scales. We suggest that loop extrusion is a universal mechanism of genome folding that mediates functional interactions during interphase and compacts chromosomes during mitosis.

Aneuploidy in eggs is a leading cause of miscarriages or viable developmental syndromes. Aneuploidy rates differ between individual chromosomes. For instance, chromosome 21 frequently missegregates, resulting in Down Syndrome. What causes chromosome-specific aneuploidy in meiosis is unclear. Chromosome 21 belongs to the class of acrocentric chromosomes, whose centromeres are located close to the chromosome end, resulting in one long and one short chromosome arm. We demonstrate that acrocentric chromosomes are generally more often aneuploid than metacentric chromosomes in porcine eggs. Kinetochores of acrocentric chromosomes are often partially covered by the short chromosome arm during meiosis I in human and porcine oocytes and orient less efficiently toward the spindle poles. These partially covered kinetochores are more likely to be incorrectly attached to the spindle. Additionally, sister chromatids of acrocentric chromosomes are held together by lower levels of cohesin, making them more vulnerable to age-dependent cohesin loss. Chromosome architecture and low cohesion therefore bias acrocentric chromosomes toward aneuploidy during mammalian meiosis. Bellou et al. show that acrocentric chromosomes, such as chromosome 21, are more prone to aneuploidy in oocytes due to their unique architecture and lower cohesion levels, which lead to inefficient alignment and increased mis-segregation during meiosis.

Cohesin mediates sister chromatid cohesion to enable chromosome segregation and DNA damage repair. To perform these functions, cohesin needs to be protected from WAPL, which otherwise releases cohesin from DNA. It has been proposed that cohesin is protected from WAPL by SORORIN. However, in vivo evidence for this antagonism is missing and SORORIN is only known to exist in vertebrates and insects. It is therefore unknown how important and widespread SORORIN’s functions are. Here we report the identification of SORORIN orthologs in Schizosaccharomyces pombe (Sor1) and Arabidopsis thaliana (AtSORORIN). sor1Δ mutants display cohesion defects, which are partially alleviated by wpl1Δ . Atsororin mutant plants display dwarfism, tissue specific cohesion defects and chromosome mis-segregation. Furthermore, Atsororin mutant plants are sterile and separate sister chromatids prematurely at anaphase I. The somatic, but not the meiotic deficiencies can be alleviated by loss of WAPL. These results provide in vivo evidence for SORORIN antagonizing WAPL, reveal that SORORIN is present in organisms beyond the animal kingdom and indicate that it has acquired tissue specific functions in plants. Cohesin functions must be tightly regulated. Here Prusén Mota et al. show that the cohesin regulator SORORIN is conserved beyond the animal kingdom and they provide the first organismal in vivo evidence that SORORIN antagonizes WAPL.

During meiotic prophase, sister chromatids are organized into axial element (AE), which underlies the structural framework for the meiotic events such as meiotic recombination and homolog synapsis. HORMA domain-containing proteins (HORMADs) localize along AE and play critical roles in the regulation of those meiotic events. Organization of AE is attributed to two groups of proteins: meiotic cohesins REC8 and RAD21L; and AE components SYCP2 and SYCP3. It has been elusive how these chromosome structural proteins contribute to the chromatin loading of HORMADs prior to AE formation. Here we newly generated Sycp2 null mice and showed that initial chromatin loading of HORMAD1 was mediated by meiotic cohesins prior to AE formation. HORMAD1 interacted not only with the AE components SYCP2 and SYCP3 but also with meiotic cohesins. Notably, HORMAD1 interacted with meiotic cohesins even in Sycp2-KO, and localized along cohesin axial cores independently of the AE components SYCP2 and SYCP3. Hormad1/Rad21L-double knockout (dKO) showed more severe defects in the formation of synaptonemal complex (SC) compared to Hormad1-KO or Rad21L-KO. Intriguingly, Hormad1/Rec8-dKO but not Hormad1/Rad21L-dKO showed precocious separation of sister chromatid axis. These findings suggest that meiotic cohesins REC8 and RAD21L mediate chromatin loading and the mode of action of HORMAD1 for synapsis during early meiotic prophase.