Schizosaccharomyces pombe
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PROVIDER: PRJNA177319 | ENA |
REPOSITORIES: ENA
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Project description:Hrp3_Purification from Schizosaccharomyces pombe 972h- Eukaryotic genome is composed of repeating units of nucleosomes to form chromatin arrays. A canonical gene is marked by nucleosome free region (NFR) at its 5’ end followed by uniformly spaced arrays of nucleosomes. In fission yeast we show both biochemically and in vivo that both Hrp1 and Hrp3 are key determinants of uniform spacing of genic arrays.
2014-05-16 | PXD000041 | Pride
Project description:Nucleosome positioning governs access to eukaryotic genomes. Many genes show a stereotypic organisation at their 5’ end: a nucleosome free region just upstream of the transcription start site (TSS) followed by a regular nucleosomal array over the coding region. The determinants for this pervasive pattern are unclear, but nucleosome remodeling ATPases likely are critical. Now we employ deletion mutants to study the role of nucleosome remodeling ATPases in global nucleosome positioning in S. pombe and the corresponding changes in expression patterns. We find a striking evolutionary shift in remodeling enzyme usage between budding and fission yeast. The S. pombe RSC remodeling complex seems not involved in nucleosome positioning, despite its prominent role in S. cerevisiae. While lacking ISWI-type remodelers, S. pombe has two CHD1-type ATPases, Hrp1 and Hrp3. We demonstrate nucleosome spacing activity for both in vitro, and together they are essential for linking regular genic arrays to most TSSs in vivo. Impaired chromatin may but need not lead to changes in transcription. The absence of both causes changed expression for about 20% and increased antisense transcription for 15% of all annotated elements.
2012-10-11 | GSE41024 | GEO
Project description:Chromatin remodelers are ATP-dependent enzymes that reorganize nucleosomes within all eukaryotic genomes. The Chd1 remodeler specializes in shifting nucleosomes into evenly spaced arrays, a defining characteristic of chromatin in gene bodies that blocks spurious transcription initiation. Linked to some forms of autism and commonly mutated in prostate cancer, Chd1 is essential for maintaining pluripotency in stem cells. Here we report a complex of yeast Chd1 bound to a nucleosome in a nucleotide-free state, determined by cryo-electron microscopy (cryo-EM) to 2.6 Å resolution. The structure shows a bulge of the DNA tracking strand where the ATPase motor engages the nucleosome, consistent with an initial stage in DNA translocation. Unlike other remodeler-nucleosome complexes, nucleosomal DNA compensates for the remodeler-induced bulge with a bulge of the complementary DNA strand one helical turn downstream from the ATPase motor. Unexpectedly, the structure also reveals an N-terminal binding motif, called ChEx, which binds on the exit-side acidic patch of the nucleosome. The ChEx motif can displace a LANA-based peptide from the acidic patch, which suggests a means by which Chd1 remodelers may block competing chromatin remodelers from acting on the opposite side of the nucleosome.
2022-02-23 | PXD025287 | Pride
Project description:Positioned nucleosomes limit the access of proteins to DNA and implement regulatory features encoded in eukaryotic genomes. Here we generated the first genome-wide nucleosome positioning map for Schizosaccharomyces pombe and annotated transcription start and termination sites genome-wide. Using this resource we found surprising differences compared to the nucleosome organization in the distantly related yeast Saccharomyces cerevisiae [the cerevisiae data has been published by others (PMID: 17873876) and the raw data is deposited at ArrayExpress(E-MEXP-1172)]. DNA sequence guides nucleosome positioning differently, e.g., poly(dA:dT) elements are not enriched in S. pombe nucleosome-depleted regions (NDRs). Regular nucleosomal arrays emanate more asymmetrically, i.e., mainly co-directionally with transcription, from promoter NDRs, but promoters harbouring the histone variant H2A.Z show regular arrays also upstream. Regular nucleosome phasing in S. pombe has a very short repeat length of 154 base pairs, and requires a remodeler, Mit1, conserved in humans but not found in S. cerevisiae. Nucleosome positioning mechanisms are evidently not universal but evolutionarily plastic.
2010-02-01 | GSE16040 | GEO
Project description:Nucleosome positioning governs access to eukaryotic genomes. Many genes show a stereotypic organisation at their 5M-bM-^@M-^Y end: a nucleosome free region just upstream of the transcription start site (TSS) followed by a regular nucleosomal array over the coding region. The determinants for this pervasive pattern are unclear, but nucleosome remodeling ATPases likely are critical. Now we employ deletion mutants to study the role of nucleosome remodeling ATPases in global nucleosome positioning in S. pombe and the corresponding changes in expression patterns. We find a striking evolutionary shift in remodeling enzyme usage between budding and fission yeast. The S. pombe RSC remodeling complex seems not involved in nucleosome positioning, despite its prominent role in S. cerevisiae. While lacking ISWI-type remodelers, S. pombe has two CHD1-type ATPases, Hrp1 and Hrp3. We demonstrate nucleosome spacing activity for both in vitro, and together they are essential for linking regular genic arrays to most TSSs in vivo. Impaired chromatin may but need not lead to changes in transcription. The absence of both causes changed expression for about 20% and increased antisense transcription for 15% of all annotated elements. For RNA expression: total RNA from hrp1D, hrp3D, hrp1Dhrp3D and wt (with actinomycin D) and total RNA from snf21ts at 25C and 34C, snf21ts swr1D at 25C and 34C, pht1D swr1D (without actinomycin D). For nucleosome mapping: Nucleosomal DNA in pht1M-NM-^T swr1M-NM-^T mutant, snf21- ts mutant, snf21- ts swr1M-NM-^T mutant, mit1M-NM-^T mutant, hrp1M-NM-^T mutant, hrp3M-NM-^T mutant and hrp1M-NM-^T hrp3M-NM-^T mutant S.pombe vs. Genomic Input DNA in wildtype and mit1M-NM-^T mutant S.pombe.
2012-10-11 | E-GEOD-41024 | biostudies-arrayexpress
Project description:Arrays of regularly spaced nucleosomes dominate chromatin and are often phased, i.e., aligned at reference sites like active promoters. How distances between nucleosomes and distances between phasing sites and nucleosomes are determined remained unclear, specifically, the role of ATP dependent chromatin remodelers in it. Here, we used a genome-wide reconstitution system to probe how yeast remodelers generate phased nucleosome arrays. We find that remodelers bear a structural element named the ‘ruler’ that sets nucleosome spacing, in the order Chd1 < ISW1a < ISW2 < INO80. Structure-based mutagenesis confirmed the functional significance of the ruler element in INO80. Differences in the ruler elements of different remodelers explain the observed nucleosome array features. More generally, we propose that remodelers use their rulers to regulate the direction of nucleosome sliding in response to nucleosome density and environment, leading to nucleosome positioning relative to other nucleosomes, DNA bound factors or DNA sequence elements.
2020-02-10 | GSE140614 | GEO
Project description:A key element for defining the centromere identity is the incorporation of a specific histone H3, CENP-A, known as Cnp1p in S. pombe. Previous studies have suggested that functional S. pombe centromeres lack nucleosome arrays and may involve chromatin remodeling as a key step of kinetochore assembly. We used tiling microarrays to show that nucleosomes are in fact positioned in regular intervals in the core of centromere 2, providing the first high resolution map of regional centromere chromatin. Nucleosome locations are not disrupted by mutations in kinetochore proteins cnp1, mis18, mis12, nuf2, mal2, overexpression of Cnp1p, or deletion of ams2. Bioinformatic analysis of the centromere sequence indicates certain enriched motifs in linker regions between nucleosomes and reveals a sequence-bias in nucleosome positioning. We conclude that centromeric nucleosome positions are stable and may be derived from the underlying DNA sequence. In addition, sequence analysis of nucleosome-free regions identifies novel binding sites for the GATA-like protein Ams2p, which participates in CENP-A incorporation. Keywords: Nucleosome Mapping Study
2008-05-30 | GSE10742 | GEO
Project description:Nucleosome arrays begin at nucleosome-free promoter regions (NFRs) and regulate gene expression. Reconstituting such organization throughout a genome with purified proteins is a critical challenge in establishing biochemical mechanisms for chromosome assembly. Here we establish a four-step hierarchical building plan for yeast genomic nucleosome organization using only purified components: genomic DNA, histones, site-specific organizing factors Abf1 and Reb1, and the chromatin remodelers RSC, ISW2, INO80, and ISW1a. First, RSC makes NFRs by translating promoter poly(dA:dT) tracts into directional nucleosome removal. Second, +1 nucleosomes are positioned by INO80 at most genes potentially involving DNA shape, or by ISW2 using gene-specific Abf1 and Reb1. Third, INO80 or ISW2 create arrays with wide spacing. Fourth, ISW1a tightens the spacing and creates properly positioned arrays. We conclude that entire genomes use a simple set of rules and proteins, without transcription, to build a common chromatin architecture. In this study, nucleosomes were assembled using Salt Gradient Dialysis (SGD) on yeast genomic DNA library. Assembled nucleosomes were either left untreated (labelled as "SGD", control), treated with whole cell extract (WCE), mutant extracts (rsc3ts WCE, isw1 isw2 chd1 WCE), purified remodelers; singly or in combinations (RSC, ISW1a, ISW1b, ISW2, INO80, CHD1, SWI/SNF), combinations of mutant extracts and chromatin remodelers or combination of General Regulatory Factors (Abf1, Reb1) and chromatin remodelers. The resulting nucleosome positions were mapped genome-wide using MNase-(anti-H3-ChIP)-Seq.
2016-08-14 | E-GEOD-72106 | biostudies-arrayexpress
Project description:A key element for defining the centromere identity is the incorporation of a specific histone H3, CENP-A, known as Cnp1p in S. pombe. Previous studies have suggested that functional S. pombe centromeres lack nucleosome arrays and may involve chromatin remodeling as a key step of kinetochore assembly. We used tiling microarrays to show that nucleosomes are in fact positioned in regular intervals in the core of centromere 2, providing the first high resolution map of regional centromere chromatin. Nucleosome locations are not disrupted by mutations in kinetochore proteins cnp1, mis18, mis12, nuf2, mal2, overexpression of Cnp1p, or deletion of ams2. Bioinformatic analysis of the centromere sequence indicates certain enriched motifs in linker regions between nucleosomes and reveals a sequence-bias in nucleosome positioning. We conclude that centromeric nucleosome positions are stable and may be derived from the underlying DNA sequence. In addition, sequence analysis of nucleosome-free regions identifies novel binding sites for the GATA-like protein Ams2p, which participates in CENP-A incorporation. Keywords: Nucleosome Mapping Study Entire cnt regions and histone-related genes were tiled at 1-5 bp spacing using 60-mer probes.
2008-05-30 | E-GEOD-10742 | biostudies-arrayexpress
Project description:Knowing the exact positions of nucleosomes not only advances our understanding of their role in gene regulation, but also the mechanisms that underlie between-species variation in chromatin structure. We have generated a chemical map of nucleosomes in vivo in Schizosaccharomyces pombe at base pair resolution. This new map reveals that S.pombe genome shares a similar periodic linker length distribution with Saccharomyces cerevisiae, but with major distinctions in nucleosomal/linker DNA sequence features. In S.pombe, A/T rich sequences are enriched in the nucleosome core region, particularly +/-20 bp of dyad, while they are disfavored in S.cerevisiae nucleosomes. The poly (dA-dT) tracts only slightly affect the nucleosome occupancy in S.pombe; and they possess preferential rotational positions within the nucleosome core with significant enrichment in the 10-30 bp region from the dyad for longer tracts. S.pombe does not have well-defined nucleosome free region immediately upstream of most transcription start sites (TSS), instead the -1 nucleosome is positioned with regular distance to the +1 nucleosome, and its occupancy is negatively correlated with gene expression. The nucleosomes around TSS show more pronounced bidirectional phasing when the intergenic distance is relatively short, and the downstream nucleosome positioning is strongly correlated with DNA sequence features. We discovered that heterochromatin regions tend to have sparse nucleosome positioning, mixed with both well-positioned and fuzzy nucleosomes. The S.pombe map suggests that some of nucleosome positioning codes, formerly thought to be intrinsic, may largely depend on species-specific extrinsic factors including linker histone, chromatin remodelers and other DNA-binding proteins. 2 samples were analyzed with high throughput paired-end parallel sequencing. Both samples were created using the same chemical mapping protocol
2013-11-12 | E-GEOD-46975 | biostudies-arrayexpress