RNAi promotes heterochromatic silencing through replication-coupled release of RNA pol II (ChIP-seq)
Ontology highlight
ABSTRACT:
ORGANISM(S): Schizosaccharomyces pombe
PROVIDER: GSE31016 | GEO | 2011/07/29
SECONDARY ACCESSION(S): PRJNA154613
REPOSITORIES: GEO
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Project description:Heterochromatin comprises tightly compacted repetitive regions of eukaryotic chromosomes. The inheritance of heterochromatin through mitosis requires RNA interference (RNAi), which guides histone modification during the DNA replication phase of the cell cycle. Here, we show that the alternating arrangement of origins of replication and non-coding RNA in pericentromeric heterochromatin results in competition between transcription and replication. Co-transcriptional RNAi releases RNA polymerase II (PolII), allowing completion of DNA replication by the leading strand DNA polymerase, and associated histone modifying enzymes which spread heterochromatin with the replication fork. In the absence of RNAi, stalled forks are repaired by homologous recombination without histone modification. small RNA cloning from wild type and dcr1-deleted strains
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Project description:Heterochromatin comprises tightly compacted repetitive regions of eukaryotic chromosomes. The inheritance of heterochromatin through mitosis requires RNA interference (RNAi), which guides histone modification during the DNA replication phase of the cell cycle. Here, we show that the alternating arrangement of origins of replication and non-coding RNA in pericentromeric heterochromatin results in competition between transcription and replication. Co-transcriptional RNAi releases RNA polymerase II (PolII), allowing completion of DNA replication by the leading strand DNA polymerase, and associated histone modifying enzymes which spread heterochromatin with the replication fork. In the absence of RNAi, stalled forks are repaired by homologous recombination without histone modification.
2011-07-21 | GSE30827 | GEO
Project description:Heterochromatin comprises tightly compacted repetitive regions of eukaryotic chromosomes. The inheritance of heterochromatin through mitosis requires RNA interference (RNAi), which guides histone modification 1 during the DNA replication phase of the cell cycle2. Here, we show that the alternating arrangement of origins of replication and non-coding RNA in pericentromeric heterochromatin results in competition between transcription and replication. Co-transcriptional RNAi releases RNA polymerase II (PolII), allowing completion of DNA replication by the leading strand DNA polymerase, and associated histone modifying enzymes3 which spread heterochromatin with the replication fork. In the absence of RNAi, stalled forks are repaired by homologous recombination without histone modification. RNA PolII ChIP from wild type and dcr1-deleted samples in exponentially growing and HU-arrested cultures.
2011-07-29 | E-GEOD-31016 | biostudies-arrayexpress
Project description:Nucleosome positioning is both active and passive and regulates access to the genome for replication, transcription and repair. Here we report that Mit1, a subunit of the fission yeast SHREC complex similar to Mi-2/NuRD, regulates transcription at regions of heterochromatin by positioning nucleosomes to preclude access to RNA Polymerase II. Purified Mit1 is a nucleosome remodeling factor capable of mobilizing histone octamers on short DNA fragments and requires ATP hydrolysis and chromatin tethering domains to remodel nucleosomes and silence transcription. We propose that SHREC is recruited to heterochromatin to mobilize nucleosomes onto unfavorable positions to prevent spurious transcription within heterochromatin. M-bM-^@M-^C RNA samples were prepared from biological duplicates of WT and mit1M-bM-^HM-^F::NatMX6 fission yeast to compare transcript levels using the Affymetrix GeneChip S.pombe Tiling 1.0FR microarray.
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Project description:Nucleosome positioning is both active and passive and regulates access to the genome for replication, transcription and repair. Here we report that Mit1, a subunit of the fission yeast SHREC complex similar to Mi-2/NuRD, regulates transcription at regions of heterochromatin by positioning nucleosomes to preclude access to RNA Polymerase II. Purified Mit1 is a nucleosome remodeling factor capable of mobilizing histone octamers on short DNA fragments and requires ATP hydrolysis and chromatin tethering domains to remodel nucleosomes and silence transcription. We propose that SHREC is recruited to heterochromatin to mobilize nucleosomes onto unfavorable positions to prevent spurious transcription within heterochromatin.
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Project description:Heterochromatin regions of the genome are transcriptionally repressive in nature. However, constitutive heterochromatin, such as at centromeres and telomeres, coats regions harboring multiple repetitive DNA elements that are preferentially transcribed during S-phase by RNA polymerase II. RNAi machinery processes the transcripts to generate small RNAs (siRNAs) that are crucial for heterochromatin assembly. Despite significant mechanistic advances in the field, the contribution of transcription factor (TF) based mechanisms to repeat element transcription and siRNA production is unclear. Here, we discover that the conserved TF PhpCNF-Y, which contains a histone fold motif, collaborates with a Zn-finger domain-containing factor, Moc3, to access specific sites within the heterochromatin region, regardless of the presence of the repressive histone methylation mark. The cooperative function of the TFs is essential for gaining access to the heterochromatin region, where they bind to specific CCAAT motif sites within the heterochromatic repeat elements cenH and dh. Perturbation of heterochromatin has minimal effect on the binding of the TFs to these specific sites, and they remain bound even outside of the S-phase. We find that PhpCNF-Y and Moc3 mediate the transcription of the repeat elements originating from a bottom strand of cenH, which the spliceosome targets to generate siRNAs in a Swi6HP1-independent manner. This pathway for generating the initial transcripts is crucial for de novo heterochromatin assembly. Collectively, our study uncovers a fundamental transcription factor-based mechanism that gains access to the heterochromatin to enforce transcriptional silencing of repeat elements. Our findings provide significant insights into de novo heterochromatin assembly.
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Project description:Heterochromatin regions of the genome are transcriptionally repressive in nature. However, constitutive heterochromatin, such as at centromeres and telomeres, coats regions harboring multiple repetitive DNA elements that are preferentially transcribed during S-phase by RNA polymerase II. RNAi machinery processes the transcripts to generate small RNAs (siRNAs) that are crucial for heterochromatin assembly. Despite significant mechanistic advances in the field, the contribution of transcription factor (TF) based mechanisms to repeat element transcription and siRNA production is unclear. Here, we discover that the conserved TF PhpCNF-Y, which contains a histone fold motif, collaborates with a Zn-finger domain-containing factor, Moc3, to access specific sites within the heterochromatin region, regardless of the presence of the repressive histone methylation mark. The cooperative function of the TFs is essential for gaining access to the heterochromatin region, where they bind to specific CCAAT motif sites within the heterochromatic repeat elements cenH and dh. Perturbation of heterochromatin has minimal effect on the binding of the TFs to these specific sites, and they remain bound even outside of the S-phase. We find that PhpCNF-Y and Moc3 mediate the transcription of the repeat elements originating from a bottom strand of cenH, which the spliceosome targets to generate siRNAs in a Swi6HP1-independent manner. This pathway for generating the initial transcripts is crucial for de novo heterochromatin assembly. Collectively, our study uncovers a fundamental transcription factor-based mechanism that gains access to the heterochromatin to enforce transcriptional silencing of repeat elements. Our findings provide significant insights into de novo heterochromatin assembly.
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Project description:Heterochromatin regions of the genome are transcriptionally repressive in nature. However, constitutive heterochromatin, such as at centromeres and telomeres, coats regions harboring multiple repetitive DNA elements that are preferentially transcribed during S-phase by RNA polymerase II. RNAi machinery processes the transcripts to generate small RNAs (siRNAs) that are crucial for heterochromatin assembly. Despite significant mechanistic advances in the field, the contribution of transcription factor (TF) based mechanisms to repeat element transcription and siRNA production is unclear. Here, we discover that the conserved TF PhpCNF-Y, which contains a histone fold motif, collaborates with a Zn-finger domain-containing factor, Moc3, to access specific sites within the heterochromatin region, regardless of the presence of the repressive histone methylation mark. The cooperative function of the TFs is essential for gaining access to the heterochromatin region, where they bind to specific CCAAT motif sites within the heterochromatic repeat elements cenH and dh. Perturbation of heterochromatin has minimal effect on the binding of the TFs to these specific sites, and they remain bound even outside of the S-phase. We find that PhpCNF-Y and Moc3 mediate the transcription of the repeat elements originating from a bottom strand of cenH, which the spliceosome targets to generate siRNAs in a Swi6HP1-independent manner. This pathway for generating the initial transcripts is crucial for de novo heterochromatin assembly. Collectively, our study uncovers a fundamental transcription factor-based mechanism that gains access to the heterochromatin to enforce transcriptional silencing of repeat elements. Our findings provide significant insights into de novo heterochromatin assembly.
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2010-07-21 | E-GEOD-22411 | biostudies-arrayexpress
Project description:One key aspect of epigenetic inheritance is that chromatin structure can be stably inherited through generations even after removal of the initial signal that establishes such structure. In fission yeast, the RNA interference (RNAi) machinery is critical for the targeting of histone methyltransferase Clr4 to repetitive DNA elements to establish a large domain of heterochromatin marked with histone H3 lysine 9 methylation (H3K9me). Subsequently during DNA replication, the deposition of parental histones containing H3K9me into daughter DNA strands is expected to recruit Clr4 to modify neighboring new histones, resulting in the inheritance of heterochromatin in both daughter cells. However, pericentric heterochromatin cannot be properly inherited in the absence of RNAi, suggesting the existence of mechanisms that counteract chromatin-based inheritance. Here we show that in the absence of certain components of the INO80 chromatin remodeling complex, pericentric heterochromatin can be properly inherited in RNAi mutants. The ability of INO80 to counter heterochromatin inheritance is attributed to one accessory subunit Iec5, which promotes histone turn over at heterochromatin regions, but has little effects on the ability of INO80 in regulating nucleosome positioning at heterochromatin, gene expression, or the DNA damage response. Slow histone turnover preserves parental histones at repeat regions to enhance epigenetic inheritance of heterochromatin not only in RNAi mutants but also in wild type cells. Our analyses identified a separation-of-function mutation of INO80 in regulating histone turnover at heterochromatin and demonstrate the important role of histone turnover in controlling heterochromatin inheritance and maintaining the proper heterochromatin landscape of the genome.
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