Functional characterization of NSE4A subunit of SMC5/6 complex in Arabidopsis
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ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE113310 | GEO | 2018/06/06
REPOSITORIES: GEO
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Project description:During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Diminished Smc5/6 function causes severe defects in nuclear division, but the underlying causes of these defects remain unclear. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of joint-molecule intermediates. Furthermore, we find that Smc5/6 specifically promotes resolution of joint molecules via the XPF-family endonuclease, Mus81-Mms4Eme1. We propose that Smc5/6 acts as a chaperone for M-bM-^@M-^XmitoticM-bM-^@M-^Y-like recombination processes during meiosis. ChIP-chip was used to compare Smc5 localization in wild-type and spo11 strains
2013-12-27 | E-GEOD-44852 | biostudies-arrayexpress
Project description:Structural Maintenance of Chromosomes (SMC) complexes - cohesin, condensin and SMC5/6 - are important for many aspects of the chromosomal organization. These complexes are composed of Smc and non-Smc element (Nse) subunits. The Nse1 subunit of SMC5/6 contains a variant RING domain, characteristic of ubiquitin ligases. Human NSE1 has been shown to possess ubiquitin ligase activity in vitro; however, other studies were unable to show such activity. Here, we confirm Nse1 ubiquitin-ligase activity in vitro using purified Schizosaccharomyces pombe proteins. We demonstrate that the Nse1 ligase activity is stimulated by Nse3 and Nse4, it specifically utilizes Ubc13/Mms2 E2 enzyme and Nse1 interacts directly with ubiquitin. We identify Nse1 mutation (R188E) that specifically disrupts its E3 activity, and show that the Nse1-dependent ubiquitination is particularly important under replication stress. Moreover, we identify Nse4 (lysine K181) as the first known SMC5/6-associated Nse1 substrate. Interestingly, abolition of Nse4 modification at K181 leads to suppression of DNA-damage sensitivity of other SMC5/6 mutants. Altogether, this study brings new evidence for Nse1 ubiquitin ligase activity, significantly advancing our under-standing of this enigmatic SMC5/6 function.
2022-01-24 | PXD029573 | Pride
Project description:During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Diminished Smc5/6 function causes severe defects in nuclear division, but the underlying causes of these defects remain unclear. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of joint-molecule intermediates. Furthermore, we find that Smc5/6 specifically promotes resolution of joint molecules via the XPF-family endonuclease, Mus81-Mms4Eme1. We propose that Smc5/6 acts as a chaperone for ‘mitotic’-like recombination processes during meiosis.
2013-12-27 | GSE44852 | GEO
Project description:The Structural Maintenance of Chromosome (SMC) protein complexes cohesin, condensin and the Smc5/6 complex (Smc5/6) are essential for chromosome function. At the molecular level, these complexes fold DNA by loop extrusion. Accordingly, cohesin creates chromosome loops in interphase, and condensin compacts mitotic chromosomes. However, the role of Smc5/6’s recently discovered DNA loop extrusion activity is unknown. Here, we uncover that Smc5/6 controls the spatial organization of supercoiled chromosomal regions. The results show that Smc5/6 associates with transcription-induced positively supercoiled chromosomal DNA at cohesin-dependent chromosome loop boundaries. Mechanistically, single-molecule imaging reveals that dimers of Smc5/6 specifically recognize the tip of positively supercoiled DNA plectonemes, and efficiently initiates loop extrusion to gather the supercoiled DNA into a large plectonemic loop. Finally, Hi-C analysis shows that Smc5/6 links chromosomal regions containing transcription-induced positive supercoiling in cis. Altogether, our findings indicate that Smc5/6 controls the three-dimensional organization of chromosomes by recognizing and initiating loop extrusion on positively supercoiled DNA.
2024-01-30 | GSE235559 | GEO
Project description:The Structural Maintenance of Chromosome (SMC) protein complexes cohesin, condensin and the Smc5/6 complex (Smc5/6) are essential for chromosome function. At the molecular level, these complexes fold DNA by loop extrusion. Accordingly, cohesin creates chromosome loops in interphase, and condensin compacts mitotic chromosomes. However, the role of Smc5/6’s recently discovered DNA loop extrusion activity is unknown. Here, we uncover that Smc5/6 controls the spatial organization of supercoiled chromosomal regions. The results show that Smc5/6 associates with transcription-induced positively supercoiled chromosomal DNA at cohesin-dependent chromosome loop boundaries. Mechanistically, single-molecule imaging reveals that dimers of Smc5/6 specifically recognize the tip of positively supercoiled DNA plectonemes, and efficiently initiates loop extrusion to gather the supercoiled DNA into a large plectonemic loop. Finally, Hi-C analysis shows that Smc5/6 links chromosomal regions containing transcription-induced positive supercoiling in cis. Altogether, our findings indicate that Smc5/6 controls the three-dimensional organization of chromosomes by recognizing and initiating loop extrusion on positively supercoiled DNA.
2024-01-30 | GSE235558 | GEO
Project description:The Structural Maintenance of Chromosome (SMC) protein complexes cohesin, condensin and the Smc5/6 complex (Smc5/6) are essential for chromosome function. At the molecular level, these complexes fold DNA by loop extrusion. Accordingly, cohesin creates chromosome loops in interphase, and condensin compacts mitotic chromosomes. However, the role of Smc5/6’s recently discovered DNA loop extrusion activity is unknown. Here, we uncover that Smc5/6 controls the spatial organization of supercoiled chromosomal regions. The results show that Smc5/6 associates with transcription-induced positively supercoiled chromosomal DNA at cohesin-dependent chromosome loop boundaries. Mechanistically, single-molecule imaging reveals that dimers of Smc5/6 specifically recognize the tip of positively supercoiled DNA plectonemes, and efficiently initiates loop extrusion to gather the supercoiled DNA into a large plectonemic loop. Finally, Hi-C analysis shows that Smc5/6 links chromosomal regions containing transcription-induced positive supercoiling in cis. Altogether, our findings indicate that Smc5/6 controls the three-dimensional organization of chromosomes by recognizing and initiating loop extrusion on positively supercoiled DNA.
2024-01-30 | GSE235557 | GEO
Project description:A family of Structural Maintenance of Chromosome (SMC) complexes is essential for key cellular processes ensuring proper cohesion, condensation and replication. They share common SMC-kleisin architecture allowing them to embrace DNA. The prokaryotic and eukaryotic SMC5/6 complexes contain additional kleisin-interacting KITE subunits which regulate SMC-kleisin dynamics. In SMC5/6, the Nse1 and Nse3 KITE proteins and Nse4 kleisin subunits form stable subcomplex which binds DNA and regulates essential processes. In addition, non-essential Nse5 and Nse6 subunits associate with the core SMC5/6 complex and recruit it to DNA repair sites. The architecture of the SMC5/6 complex is crucial for its proper functioning and mutations within the human SMC5/6 subunits result in severe syndromes. Therefore, we aimed to analyse interactions within the human SMC5/6 complex and determine its architecture at detail levels. First, we analysed the human hNSE1-hNSE3 dimer and the hNSE1-hNSE3-hNSE4 trimer by crosslinking and MS/MS analysis. Our data suggested domain arrangements of hNSE1-hNSE3 and orientation of hNSE4 within the trimer. The crosslinking and electron microscopy analysis of the SMC5/6 core complex showed its rod-like shaped architecture with juxtaposed hSMC5-hSMC6 arms. In addition, we observed fully or partially opened hSMC5-hSMC6 shapes with the hNSE1-hNSE3-hNSE4 trimer localized at the SMC head domains. To complete mapping of the human SMC5/6 complex architecture, we analysed positions of hNSE5-hNSE6 at the hSMC5-hSMC6 arms. Interestingly, hNSE6 bound both hSMC5-hSMC6 arms, suggesting that hNSE6 may lock the arms and regulate their dynamics. All together, we gained new insights into the architecture and dynamics of the human SMC5/6 complex.
2023-08-18 | PXD017379 | Pride
Project description:This data is from BS3 crosslinked condensin tetramer How protein complexes of the SMC family fold DNA into the large loops that are fundamental for the 3D organization of genomes is a central unresolved question of chromosome biology. We used electron cryomicroscopy to investigate the reaction cycle of the SMC complex condensin, which is a key determinant of chromosome morphology and behavior during mitosis. Our structures of the Saccharomyces cerevisiae condensin holo complex at different functional stages suggest that ATP binding induces the transition from a folded-rod SMC conformation into an open architecture and triggers the exchange of the two HEAT-repeat subunits at the SMC ATPase head domains. We propose that these steps result in the interconversion of DNA binding sites in the catalytic core that form the basis of the DNA translocation and loop-extrusion activities of condensin.
2020-07-28 | PXD019275 | Pride
Project description:Structural maintenance of chromosomes (SMC) complexes, cohesin, condensin and Smc5/6, are essential for viability and participate in multiple processes, including sister chromatid cohesion, chromosome condensation, and DNA repair. Here we show that SUMO chains target all three SMC complexes and are antagonized by the SUMO protease Ulp2 to prevent their turnover. We uncover that the essential role of the cohesin-associated subunit Pds5 is to counteract SUMO chains jointly with Ulp2. Importantly, fusion of Ulp2 to kleisin Scc1 supports viability of PDS5 null cells and protects cohesin from proteasomal degradation mediated by the SUMO-targeted ubiquitin ligase Slx5/Slx8. The lethality of PDS5 deleted cells can also be bypassed by simultaneous loss of the PCNA unloader, Elg1, and the cohesin releaser, Wpl1, but only when Ulp2 is functional. Condensin and Smc5/6 complex are similarly guarded by Ulp2 against unscheduled SUMO-chain assembly, which we propose to time the availability of SMC complexes on chromatin.
2021-08-05 | PXD022717 | Pride
Project description:This data is from sulfo-SDA crosslinked condensin pentamer. Two datsets, one without atp aand one with ATP. How protein complexes of the SMC family fold DNA into the large loops that are fundamental for the 3D organization of genomes is a central unresolved question of chromosome biology. We used electron cryomicroscopy to investigate the reaction cycle of the SMC complex condensin, which is a key determinant of chromosome morphology and behavior during mitosis. Our structures of the Saccharomyces cerevisiae condensin holo complex at different functional stages suggest that ATP binding induces the transition from a folded-rod SMC conformation into an open architecture and triggers the exchange of the two HEAT-repeat subunits at the SMC ATPase head domains. We propose that these steps result in the interconversion of DNA binding sites in the catalytic core that form the basis of the DNA translocation and loop-extrusion activities of condensin.
2020-07-28 | PXD019274 | Pride