Meiotic cohesin REC8 marks the axial elements of rat synaptonemal complexes before cohesins SMC1β and SMC3

In meiotic prophase, the sister chromatids of each chromosome develop a common axial element (AE) that is integrated into the synaptonemal complex (SC). We analyzed the incorporation of sister chromatid cohesion proteins (cohesins) and other AE components into AEs. Meiotic cohesin REC8 appeared shortly before premeiotic S phase in the nucleus and formed AE-like structures (REC8-AEs) from premeiotic S phase on. Subsequently, meiotic cohesin SMC1β, cohesin SMC3, and AE proteins SCP2 and SCP3 formed dots along REC8-AEs, which extended and fused until they lined REC8-AEs along their length. In metaphase I, SMC1β, SMC3, SCP2, and SCP3 disappeared from the chromosome arms and accumulated around the centromeres, where they stayed until anaphase II. In striking contrast, REC8 persisted along the chromosome arms until anaphase I and near the centromeres until anaphase II. We propose that REC8 provides a basis for AE formation and that the first steps in AE assembly do not require SMC1β, SMC3, SCP2, and SCP3. Furthermore, SMC1β, SMC3, SCP2, and SCP3 cannot provide arm cohesion during metaphase I. We propose that REC8 then provides cohesion. RAD51 and/or DMC1 coimmunoprecipitates with REC8, suggesting that REC8 may also provide a basis for assembly of recombination complexes

Cohesin Smc1β determines meiotic chromatin axis loop organization

Meiotic chromosomes consist of proteinaceous axial structures from which chromatin loops emerge. Although we know that loop density along the meiotic chromosome axis is conserved in organisms with different genome sizes, the basis for the regular spacing of chromatin loops and their organization is largely unknown. We use two mouse model systems in which the postreplicative meiotic chromosome axes in the mutant oocytes are either longer or shorter than in wild-type oocytes. We observe a strict correlation between chromosome axis extension and a general and reciprocal shortening of chromatin loop size. However, in oocytes with a shorter chromosome axis, only a subset of the chromatin loops is extended. We find that the changes in chromatin loop size observed in oocytes with shorter or longer chromosome axes depend on the structural maintenance of chromosomes 1β (Smc1β), a mammalian chromosome–associated meiosis-specific cohesin. Our results suggest that in addition to its role in sister chromatid cohesion, Smc1β determines meiotic chromatin loop organization

Cohesin SMC1β protects telomeres in meiocytes

Telomeres fail to attach to the nuclear envelope and lose structural integrity in cells lacking SMC1β

Molecular basis for SMC rod formation and its dissolution upon DNA binding.

SMC condensin complexes are central modulators of chromosome superstructure in all branches of life. Their SMC subunits form a long intramolecular coiled coil, which connects a constitutive "hinge" dimerization domain with an ATP-regulated "head" dimerization module. Here, we address the structural arrangement of the long coiled coils in SMC complexes. We unequivocally show that prokaryotic Smc-ScpAB, eukaryotic condensin, and possibly also cohesin form rod-like structures, with their coiled coils being closely juxtaposed and accurately anchored to the hinge. Upon ATP-induced binding of DNA to the hinge, however, Smc switches to a more open configuration. Our data suggest that a long-distance structural transition is transmitted from the Smc head domains to regulate Smc-ScpAB's association with DNA. These findings uncover a conserved architectural theme in SMC complexes, provide a mechanistic basis for Smc's dynamic engagement with chromosomes, and offer a molecular explanation for defects in Cornelia de Lange syndrome

The Coiled Coils of Cohesin Are Conserved in Animals, but Not In Yeast

The SMC proteins are involved in DNA repair, chromosome condensation, and sister chromatid cohesion throughout Eukaryota. Long, anti-parallel coiled coils are a prominent feature of SMC proteins, and are thought to serve as spacer rods to provide an elongated structure and to separate domains. We reported recently that the coiled coils of mammalian condensin (SMC2/4) showed moderate sequence divergence (approximately 10-15%) consistent with their functioning as spacer rods. The coiled coils of mammalian cohesins (SMC1/3), however, were very highly constrained, with amino acid sequence divergence typically <0.5%. These coiled coils are among the most highly conserved mammalian proteins, suggesting that they make extensive contacts over their entire surface.Here, we broaden our initial analysis of condensin and cohesin to include additional vertebrate and invertebrate organisms and multiple species of yeast. We found that the coiled coils of SMC1/3 are highly constrained in Drosophila and other insects, and more generally across all animal species. However, in yeast they are no more constrained than the coils of SMC2/4 and Ndc80/Nuf2p, suggesting that they are serving primarily as spacer rods.SMC1/3 functions for sister chromatid cohesion in all species. Since its coiled coils apparently serve only as spacer rods in yeast, it is likely that this is sufficient for sister chromatid cohesion in all species. This suggests an additional function in animals that constrains the sequence of the coiled coils. Several recent studies have demonstrated that cohesin has a role in gene expression in post-mitotic neurons of Drosophila, and other animal cells. Some variants of human Cornelia de Lange Syndrome involve mutations in human SMC1/3. We suggest that the role of cohesin in gene expression may involve intimate contact of the coiled coils of SMC1/3, and impose the constraint on sequence divergence

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The cohesin complex mediates DNA-DNA interactions both between (sister chromatid cohesion) and within chromosomes (DNA looping). It has been suggested that intra-chromosome loops are generated by extrusion of DNAs through the lumen of cohesin's ring. Scc2 (Nipbl) stimulates cohesin's ABC-like ATPase and is essential for loading cohesin onto chromosomes. However, it is possible that the stimulation of cohesin's ATPase by Scc2 also has a post-loading function, for example driving loop extrusion. Using fluorescence recovery after photobleaching (FRAP) and single- molecule tracking, we show that Scc2 binds dynamically to chromatin, principally through an association with cohesin. Scc2's movement within chromatin is consistent with a 'stop-and-go' or 'hopping' motion. We suggest that a low diffusion coefficient, a low stoichiometry relative to cohesin, and a high affinity for chromosomal cohesin enables Scc2 to move rapidly from one chromosomal cohesin complex to another, performing a function distinct from loading

A molecular model for the role of SYCP3 in meiotic chromosome organisation.

The synaptonemal complex (SC) is an evolutionarily-conserved protein assembly that holds together homologous chromosomes during prophase of the first meiotic division. Whilst essential for meiosis and fertility, the molecular structure of the SC has proved resistant to elucidation. The SC protein SYCP3 has a crucial but poorly understood role in establishing the architecture of the meiotic chromosome. Here we show that human SYCP3 forms a highly-elongated helical tetramer of 20 nm length. N-terminal sequences extending from each end of the rod-like structure bind double-stranded DNA, enabling SYCP3 to link distant sites along the sister chromatid. We further find that SYCP3 self-assembles into regular filamentous structures that resemble the known morphology of the SC lateral element. Together, our data form the basis for a model in which SYCP3 binding and assembly on meiotic chromosomes leads to their organisation into compact structures compatible with recombination and crossover formation. DOI: http://dx.doi.org/10.7554/eLife.02963.00

corona Is Required for Higher-Order Assembly of Transverse Filaments into Full-Length Synaptonemal Complex in Drosophila Oocytes

The synaptonemal complex (SC) is an intricate structure that forms between homologous chromosomes early during the meiotic prophase, where it mediates homolog pairing interactions and promotes the formation of genetic exchanges. In Drosophila melanogaster, C(3)G protein forms the transverse filaments (TFs) of the SC. The N termini of C(3)G homodimers localize to the Central Element (CE) of the SC, while the C-termini of C(3)G connect the TFs to the chromosomes via associations with the axial elements/lateral elements (AEs/LEs) of the SC. Here, we show that the Drosophila protein Corona (CONA) co-localizes with C(3)G in a mutually dependent fashion and is required for the polymerization of C(3)G into mature thread-like structures, in the context both of paired homologous chromosomes and of C(3)G polycomplexes that lack AEs/LEs. Although AEs assemble in cona oocytes, they exhibit defects that are characteristic of c(3)G mutant oocytes, including failure of AE alignment and synapsis. These results demonstrate that CONA, which does not contain a coiled coil domain, is required for the stable ‘zippering’ of TFs to form the central region of the Drosophila SC. We speculate that CONA's role in SC formation may be similar to that of the mammalian CE proteins SYCE2 and TEX12. However, the observation that AE alignment and pairing occurs in Tex12 and Syce2 mutant meiocytes but not in cona oocytes suggests that the SC plays a more critical role in the stable association of homologs in Drosophila than it does in mammalian cells

Genome-wide DNA methylation analysis in cohesin mutant human cell lines

The cohesin complex has recently been shown to be a key regulator of eukaryotic gene expression, although the mechanisms by which it exerts its effects are poorly understood. We have undertaken a genome-wide analysis of DNA methylation in cohesin-deficient cell lines from probands with Cornelia de Lange syndrome (CdLS). Heterozygous mutations in NIPBL, SMC1A and SMC3 genes account for ∼65% of individuals with CdLS. SMC1A and SMC3 are subunits of the cohesin complex that controls sister chromatid cohesion, whereas NIPBL facilitates cohesin loading and unloading. We have examined the methylation status of 27 578 CpG dinucleotides in 72 CdLS and control samples. We have documented the DNA methylation pattern in human lymphoblastoid cell lines (LCLs) as well as identified specific differential DNA methylation in CdLS. Subgroups of CdLS probands and controls can be classified using selected CpG loci. The X chromosome was also found to have a unique DNA methylation pattern in CdLS. Cohesin preferentially binds to hypo-methylated DNA in control LCLs, whereas the differential DNA methylation alters cohesin binding in CdLS. Our results suggest that in addition to DNA methylation multiple mechanisms may be involved in transcriptional regulation in human cells and in the resultant gene misexpression in CdLS