Localization of Scc2 between wild type and scc4-m35 cells in S-phase
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE70316 | GEO | 2016/07/21
SECONDARY ACCESSION(S): PRJNA288305
REPOSITORIES: GEO
Similar Datasets
Project description:Wild-type (AM15307) and scc4-m35 (AM15311) cells carrying SCC2-3FLAG were fixed for 2 hr, 15 min following an a-factor arrest. Samples were harvested, anti-HA ChIP was performed and both input and IP samples were sequenced for both strains. ChIP-seq for S. cerevisiae Scc2-3FLAG in S-phase cells in Wild type and in scc4-m35 mutant Vasso Makrantoni: experiments and Alastair Kerr: ChIP Seq Data analysis
2016-07-21 | E-GEOD-70316 | biostudies-arrayexpress
Project description:Purpose: The Goal of this study is to look at 1) Differences in genome wide localization of Scc2 between wild type and scc4-m35 cells in S-phase 2) Differences in genome wide localisation of Scc1 cohesin between wild type and scc4-m35 in metaphase arrested cells. Method: 1) Wild-type (AM15307) and scc4-m35 (AM15311) cells carrying SCC2-3FLAG were fixed for 2 hr, 15 min following an a-factor arrest. Samples were harvested, anti-HA ChIP was performed and both input and IP samples were sequenced for both strains. 2) Wild-type (AM1145), wild type constructed in the same way as the mutant strain (carrying a HIS3 marker) (AM15540) and scc4-m35 mutant (AM15537) cells carrying SCC1-6HA were synchronised with a-factor and arrested in mitosis by treatment with nocodazole/benomyl for 2 hr. Samples were harvested, anti-HA ChIP was performed and input and IP samples were sequenced for the three strains. Results: 1) This experiment did not yield meaningful data. 2) All 16 centromeres were specifically depleted of Scc1 in the scc4-m35 background. Scc1 depletion extended roughly 10 kilobases to either side of the core centromere but not to chromosome arms 1) ChIP-seq for S. cerevisiae Scc2-3FLAG in S-phase cells in Wild type and in scc4-m35 mutant (samples VM1-VM4). 2) ChIP-seq for Scc1-6HA in metaphase-arrested cells in Wild type and in scc4-m35 mutant (samples VM5-VM10) Vasso Makrantoni: experiments and Alastair Kerr: ChIP Seq Data analysis https://github.com/AlastairKerr/Hinshaw2015
2015-06-05 | E-GEOD-68573 | biostudies-arrayexpress
Project description:The functions of cohesin are central to genome integrity, chromosome organization, and transcription regulation through its prevention of premature sister-chromatid separation and the formation of DNA loops. The loading of cohesin onto chromatin depends on the Scc2-Scc4 complex, however, little is known about how it stimulates the cohesin loading activity. Here we determine the large “hook” structure of Scc2 responsible for catalyzing cohesin loading. We identify key Scc2 surfaces that are crucial for DNA binding and for cohesin loading in vivo. Using previously determined structures and modeling, we derive a pseudo-atomic structure of the full-length Scc2-Scc4 complex. Finally, our crosslinking and electron microscopy analyses reveal that Scc2-Scc4 utilizes its modular structure to make multiple contacts with a folded cohesin at an interface created by the cohesin head-hinge interaction.
2017-01-11 | PXD004692 | Pride
Project description:The ring-like cohesin complex plays an essential role in chromosome segregation, organization, and double-strand break repair through its ability to bring two DNA double helices together. Scc2 (NIPBL in humans) together with Scc4 function as the loader of cohesin onto chromosomes. Chromatin adapters such as the RSC complex facilitate localization of the Scc2-Scc4 cohesin loader. Here we identify a broad range of Scc2- chromatin protein interactions that are evolutionarily conserved and reveal a role for one complex, Mediator, in recruitment of the cohesin loader. We identified budding yeast Med14, a subunit of the Mediator complex, as a high copy suppressor of poor growth in Scc2 mutant strains. Physical and genetic interactions between Scc2 and Mediator are functionally substantiated in direct recruitment and cohesion assays. Depletion of Med14 results in defective sister chromatid cohesion and decreased binding of Scc2 at RNA Pol II transcribed genes. Previous work has suggested that Mediator, Nipbl, and cohesin connect enhancers and promoters of active mammalian genes. Our studies suggest an evolutionarily conserved fundamental role for Mediator in direct recruitment of Scc2 to RNA pol II transcribed genes. We identified two mutations in the evolutionarily conserved HEAT domain of SCC2 that result in significantly reduced growth, scc2R787G and scc2G1242V. This experiment uses RNA-Seq analysis to study the effect of these mutations on gene expression.
2022-05-25 | GSE201192 | GEO
Project description:The ring-like cohesin complex plays an essential role in chromosome segregation, organization, and double-strand break repair through its ability to bring two DNA double helices together. Scc2 (NIPBL in humans) together with Scc4 function as the loader of cohesin onto chromosomes. Chromatin adapters such as the RSC complex facilitate localization of the Scc2-Scc4 cohesin loader. Here we identify a broad range of Scc2- chromatin protein interactions that are evolutionarily conserved and reveal a role for one complex, Mediator, in recruitment of the cohesin loader. We identified budding yeast Med14, a subunit of the Mediator complex, as a high copy suppressor of poor growth in Scc2 mutant strains. Physical and genetic interactions between Scc2 and Mediator are functionally substantiated in direct recruitment and cohesion assays. Depletion of Med14 results in defective sister chromatid cohesion and decreased binding of Scc2 at RNA Pol II transcribed genes. Previous work has suggested that Mediator, Nipbl, and cohesin connect enhancers and promoters of active mammalian genes. Our studies suggest an evolutionarily conserved fundamental role for Mediator in direct recruitment of Scc2 to RNA pol II transcribed genes. We identified two mutations in the evolutionarily conserved HEAT domain of SCC2 that result in significantly reduced growth, scc2R787G and scc2G1242V. This experiment uses RNA-Seq analysis to study the effect of these mutations on gene expression.
2022-05-25 | GSE201191 | GEO
Project description:Purpose: The goal of this study is to look at differences in genome wide localisation of Scc1 cohesin between wild type and scc4-m35 in metaphase arrested cells. Method: Wild-type (AM1145), wild type constructed in the same way as the mutant strain (carrying a HIS3 marker) (AM15540) and scc4-m35 mutant (AM15537) cells carrying SCC1-6HA were synchronised with a-factor and arrested in mitosis by treatment with nocodazole/benomyl for 2 hr. Samples were harvested, anti-HA ChIP was performed and input and IP samples were sequenced for the three strains. Results: All 16 centromeres were specifically depleted of Scc1 in the scc4-m35 background. Scc1 depletion extended roughly 10 kilobases to either side of the core centromere but not to chromosome arms
2015-06-05 | GSE68573 | GEO
Project description:The ring-like cohesin complex plays an essential role in chromosome segregation, organization, and double-strand break repair through its ability to bring two DNA double helices together. Scc2 (NIPBL in humans) together with Scc4 function as the loader of cohesin onto chromosomes. Chromatin adapters such as the RSC complex facilitate localization of the Scc2-Scc4 cohesin loader. Here we identify a broad range of Scc2- chromatin protein interactions that are evolutionarily conserved and reveal a role for one complex, Mediator, in recruitment of the cohesin loader. We identified budding yeast Med14, a subunit of the Mediator complex, as a high copy suppressor of poor growth in Scc2 mutant strains. Physical and genetic interactions between Scc2 and Mediator are functionally substantiated in direct recruitment and cohesion assays. Depletion of Med14 results in defective sister chromatid cohesion and decreased binding of Scc2 at RNA Pol II transcribed genes. Previous work has suggested that Mediator, Nipbl, and cohesin connect enhancers and promoters of active mammalian genes. Our studies suggest an evolutionarily conserved fundamental role for Mediator in direct recruitment of Scc2 to RNA pol II transcribed genes. We identified two mutations in the evolutionarily conserved HEAT domain of SCC2 that result in significantly reduced growth, scc2R787G and scc2G1242V. This experiment uses ChIP Seq to examine global localization of Scc2 in the presence or absence of MED14.
2022-05-25 | GSE201193 | GEO
Project description:Cohesin’s association with and translocation along chromosomal DNAs depend on an ATP hydrolysis cycle driving the association and subsequent release of DNA. This involves DNA being ‘clamped’ by Scc2 and ATP-dependent engagement of cohesin’s Smc1 and Smc3 head domains. Scc2’s replacement by Pds5 abrogates cohesin’s ATPase and has an important role in halting DNA loop extrusion. The ATPase domains of all SMC proteins are separated from their hinge dimerisation domains by 50 nm long coiled coils, which have been observed to zip up along their entire length and fold around an elbow, thereby greatly shortening the distance between hinges and ATPase heads. Whether folding exists in vivo or has any physiological importance is not known. We present here a cryo-EM structure of the apo form of cohesin that reveals the structure of folded and zipped up coils in unprecedented detail and shows that Scc2 can associate with Smc1’s ATPase head even when it is fully disengaged from that of Smc3. Using cysteine-specific cross-linking, we show that cohesin’s coiled coils are frequently folded in vivo, including when cohesin holds sister chromatids together. Moreover, we describe a mutation (SMC1D588Y) within Smc1’s hinge that alters how Scc2 and Pds5 interact with Smc1’s hinge and that enables Scc2 to support loading in the absence of its normal partner Scc4. The mutant phenotype of loading without Scc4 is only explicable if loading depends on an association between Scc2/4 and cohesin’s hinge, which in turn requires coiled coil folding.
2021-07-21 | GSE167318 | GEO
Project description:In addition to sharing with condensin an ability to organize DNA into chromatids, cohesin regulates enhancer-promoter interactions and confers sister chromatid cohesion. Association with chromosomes is regulated by hook-shaped HEAT repeat proteins that Associate With its Kleisin (Scc1) subunit (HAWKs), namely Scc3, Pds5, and Scc2. Unlike Pds5, Scc2 is not a stable cohesin constituent but, as shown here, transiently displaces Pds5 during loading. Scc1 mutations that compromise its interaction with Scc2 adversely affect cohesin’s ATPase activity, loading, and translocation while Scc2 mutations that alter how the ATPase responds to DNA abolish loading despite cohesin’s initial association with loading sites. Lastly, Scc2 mutations that permit loading in the absence of Scc4 increase Scc2’s association with chromosomal cohesin and reduce that of Pds5. We suggest that cohesin switches between two states, one with Pds5 bound to Scc1 that is not able to hydrolyse ATP efficiently but is capable of release from chromosomes and another in which Scc2, transiently replacing Pds5, stimulates the ATP hydrolysis necessary for loading and translocation away from loading sites.
2018-07-17 | GSE106182 | GEO
Project description:The Scc2/Scc4 complex binds to broad nucleosome-free regions in the promoters of highly expressed genes. The cohesin loader is recruited to these sites by the RSC chromatin remodeling complex ChIp- Sequencing analysis of the distribution of Scc2 and Scc4 on budding yeast chromosomes.An input sample was analysed as a control.
2014-08-01 | E-GEOD-56993 | biostudies-arrayexpress