Nucleosome positioning under perturbation conditions in yeast
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE56095 | GEO | 2014/10/23
SECONDARY ACCESSION(S): PRJNA242410
REPOSITORIES: GEO
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Project description:Proper chromatin organization is essential for defining transcription units and maintaining genomic integrity in eukaryotes. Mutations affecting the chromatin structure can lead to increased cryptic transcription and genomic instability. In this study we found that deletion of the Schizosaccharomyces pombe Chd1-type chromatin remodelers, hrp1 and hrp3, causes strong, genome-wide accumulation of antisense transcripts, while the amount of coding mRNA transcripts is mostly unaffected. Nucleosome mapping revealed a specific role for Chd1-remodelers in the positioning of nucleosomes in gene coding regions. While the arrangement of nucleosomes in promoter regions was similar to WT, nucleosome organization within coding regions was remarkably irregular in hrp1∆hrp3∆ strain. We extended our analysis to other mutations associated with enhanced cryptic transcription activity, such as set2∆, alp13∆, and FACT complex subunit pob3∆. While nucleosomes were severely depleted in the pob3∆ strain, nucleosome positioning was less affected. In sharp contrast, nucleosome organization in the alp13∆ and set2∆ strains was indistinguishable from WT. These data indicate multiple mechanisms in the repression of cryptic promoter activity in eukaryotic cells. Genome-wide profiling of H3K9/K14 acetylation Genome-wide expression analysis of either Alp13-, Set2-, Hrp3 or Hrp1 and Hrp3-deficient cells Genome-wide expression analysis of either Hrp1, Hrp3, or Hrp1 and Hrp3-deficient cells Nucleosome mapping experiments
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Project description:To measure nucleosome positions on a genomic scale, we developed a high-throughput DNA microarray method to identify nucleosomal and linker DNA sequences based on susceptibility of linker DNA to micrococcal nuclease. Nucleosomal DNA was isolated and labeled with Cy3 fluorescent dye (green), and mixed with Cy5-labeled total genomic DNA (red). This mixture was hybridized to microarrays printed with overlapping 50mer oligonucleotide probes tiled across chromosomal regions of interest. Keywords = nucleosome Keywords = yeast Keywords = tiling microrray Keywords: repeat sample
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Project description:Proper chromatin organization is essential for defining transcription units and maintaining genomic integrity in eukaryotes. Mutations affecting the chromatin structure can lead to increased cryptic transcription and genomic instability. In this study we found that deletion of the Schizosaccharomyces pombe Chd1-type chromatin remodelers, hrp1 and hrp3, causes strong, genome-wide accumulation of antisense transcripts, while the amount of coding mRNA transcripts is mostly unaffected. Nucleosome mapping revealed a specific role for Chd1-remodelers in the positioning of nucleosomes in gene coding regions. While the arrangement of nucleosomes in promoter regions was similar to WT, nucleosome organization within coding regions was remarkably irregular in hrp1M-bM-^HM-^Fhrp3M-bM-^HM-^F strain. We extended our analysis to other mutations associated with enhanced cryptic transcription activity, such as set2M-bM-^HM-^F, alp13M-bM-^HM-^F, and FACT complex subunit pob3M-bM-^HM-^F. While nucleosomes were severely depleted in the pob3M-bM-^HM-^F strain, nucleosome positioning was less affected. In sharp contrast, nucleosome organization in the alp13M-bM-^HM-^F and set2M-bM-^HM-^F strains was indistinguishable from WT. These data indicate multiple mechanisms in the repression of cryptic promoter activity in eukaryotic cells. Genome-wide profiling of H3K9/K14 acetylation Genome-wide expression analysis of either Alp13-, Set2-, Hrp3 or Hrp1 and Hrp3-deficient cells Genome-wide expression analysis of either Hrp1, Hrp3, or Hrp1 and Hrp3-deficient cells Nucleosome mapping experiments ChIP for the detection of the genome-wide acetylation profile of H3K9/K14 was performed for a wildtype strain and the deletion strains of Set2, Alp13 and the double knock-out of Hrp1 and Hrp3. Each experiment was performed twice in biological replicates All experiments were performed twice in biological replicates, except for the expression array of set2M-NM-^T. The replicates of hrp3M-NM-^T and hrp1M-NM-^Thrp3M-NM-^T were performed with a slightly different array design All experiments were performed twice in biological replicates. The replicates of hrp3M-NM-^T and hrp1M-NM-^Thrp3M-NM-^T were performed with a slightly different array design MNase treated sample were comparatively hybridized with genomic DNA of corresponding strain, WT, hrp1d hrp3d, pob3d in biological duplicates, set2d, alp13d, mit1d, hrp1d, hrp3d analysis was performed once
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Project description:To measure nucleosome positions on a genomic scale, we developed a high-throughput DNA microarray method to identify nucleosomal and linker DNA sequences based on susceptibility of linker DNA to micrococcal nuclease. Nucleosomal DNA was isolated and labeled with Cy3 fluorescent dye (green), and mixed with Cy5-labeled total genomic DNA (red). This mixture was hybridized to microarrays printed with overlapping 50mer oligonucleotide probes tiled across chromosomal regions of interest. Keywords = nucleosome Keywords = yeast Keywords = tiling microrray
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Project description:We report nucleosome poisitioning in Yeast Mono-nucleosome DNA was mapped
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