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The Scc2NIPBL/Scc4MAU2 complex acts in sister chromatid cohesion and transcriptional regulation by maintaining nucleosome-free regions [ChIP-seq]

ABSTRACT: 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.

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Lidia Lopez-Serra

PROVIDER: E-GEOD-56993 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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The Scc2NIPBL/Scc4MAU2 complex acts in sister chromatid cohesion and transcriptional regulation by maintaining nucleosome-free regions [Mnase-seq]

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 MNase digestion of purified DNA followed by paired-end sequencing was performed in order to investigate genome-wide nucleosomal positioning in S. cerevisiae under multiple experimental conditions.

2014-08-01 | E-GEOD-56992 | biostudies-arrayexpress

Releasing activity disengages cohesinâs Smc3/Scc1 interface in a process blocked by acetylation

Project description:Sister chromatid cohesion conferred by entrapment of sister DNAs within a tripartite ring formed between cohesinâs Scc1, Smc1, and Smc3 subunits is generated during S and eventually destroyed at anaphase through cleavage of Scc1 by separase. Throughout the cell cycle, cohesinâs association with chromosomes is controlled by opposing activities: loading by the Scc2/4 complex and release by a separase independent releasing activity. Co-entrapment of sister DNAs during replication is accompanied by acetylation of Smc3 by Eco1, which blocks releasing activity and ensures that sisters remain stably connected. Because fusion of Smc3 to Scc1 prevents release and bypasses the requirement for Eco1, we suggested that release is mediated by disengagement of the Smc3/Scc1 interface. We now show that all mutations capable of bypassing Eco1, be they in cohesinâs Smc1, Smc3, Scc1,Wapl, Pds5, or Scc3 subunits, greatly reduce dissociation of N-terminal cleavage fragments of Scc1 (NScc1) from Smc3. We show that this process involves interaction between Smc ATPase heads and is inhibited by Smc3 acetylation Effect of mutations QQ and EQ in Smc3 on cohesin loading onto chromosomes

2016-02-18 | E-GEOD-76890 | biostudies-arrayexpress

Genome-wide nucleosome, TFIIIC and TFIIIB occupancies in Saccharomyces cerevisiae

Project description:Paired-end sequencing study of nucleosomes and immuno-precipitated Tfc1 and Brf1 complexes from MNase-digested nuclei. Nucleosomal DNA, input DNA and DNA from immunopurified TFIIIB and TFIIIC complexes (IP) were subjected to paired-end sequencing.

2013-07-17 | E-GEOD-44586 | biostudies-arrayexpress

Structure of the Cohesin Loader Scc2-Scc4 and its Binding to a Folded Cohesin

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

Structural evidence for Scc4-dependent localization of cohesin loading

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

Inactivation of the Budding Yeast Cohesin Loader Scc2 alters Gene Expression both Globally and in Response to a Single DNA Double Strand Break

Project description:Genome integrity is fundamental for cell survival and cell cycle progression. Important mechanisms for keeping the genome intact are proper sister chromatid segregation, correct gene regulation and efficient repair of damaged DNA. Cohesin and its DNA loader, the Scc2/4 complex have been implicated in all these cellular actions. The gene regulation role has been described in several organisms. In yeast it has been suggested that the proteins in the cohesin network would effect transcription based on its role as insulator. More recently, data are emerging indicating direct roles for gene regulation also in yeast. Here we extend these studies by investigating whether the cohesin loader Scc2 is involved in regulation of gene expression. We performed global gene expression profiling in the absence and presence of DNA damage, in wild type and Scc2 deficient G2/M arrested cells, when it is known that Scc2 is important for DNA double strand break repair and formation of damage induced cohesion. We found that not only the DNA damage specific transcriptional response is distorted after inactivation of Scc2, but also the overall transcription profile. Interestingly, these alterations did not correlate with changes in cohesin binding. We investigated the gene transcription profiles using GeneChip Yeast Genome 2.0 Array in WT and Scc2 deficient cells in the absence and presence of DNA damage.

2014-10-16 | E-GEOD-61530 | biostudies-arrayexpress

RNA polymerase II collision interrupts convergent transcription

Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series

2012-11-30 | E-GEOD-38384 | biostudies-arrayexpress

localization of Scc2 between wild type and scc4-m35 cells in S-phase

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

Phosphorylation of Cohesin's Deposition Factor, Scc2 Regulates its Deposition Activity

Project description:Phosphorylated residues of the budding yeast Scc2 protein were identified using phosphopeptide enrichment and mass spectrometry.

2015-09-10 | PXD001830 | Pride

RNA polymerase II collision interrupts convergent transcription (ChIP-seq)

Project description:Anti-sense non-coding transcripts, genes-within-genes, and convergent gene pairs are prevalent among eukaryotes. The existence of such transcription units raises the question of what happens when RNA polymerase II (RNAPII) molecules collide head-to-head. Here we use a combination of biochemical and genetic approaches in yeast to show that polymerases transcribing opposite DNA strands cannot bypass each other. RNAPII stops, but does not dissociate upon head-to-head collision in vitro, suggesting that opposing polymerases represent insurmountable obstacles for each other. Head-to-head collision in vivo results in RNAPII stopping as well, and removal of collided RNAPII from the DNA template can be achieved via ubiquitylation-directed proteolysis. Indeed, in cells lacking efficient RNAPII poly-ubiquitylation, the half-life of collided polymerases increases, so that these can be detected between convergent genes by ChIP-Seq. These results provide new insight into fundamental mechanisms of gene traffic control, and point to an unexplored effect of anti-sense transcription on gene regulation via polymerase collision. ChIP-Seq of RNA polymerase II was performed with WT and Elongain C deletion mutant (elc1M-bM-^HM-^F) cells. 4H8 antibody against the Rpb1 C-terminal domain was used for RNA polymerase II immunoprecipitation, whilst mouse IgG antibody was used for control immunoprecipitations. Two biological replicates were performed for both WT and elc1M-bM-^HM-^F.

2012-11-30 | E-GEOD-38382 | biostudies-arrayexpress

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