Chromatin Dynamics During DNA Replication
Ontology highlight
ABSTRACT:
PROVIDER: PRJEB13217 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR1354210_1.fastq.gz | Fastqsanger.gz | |||
| ERR1354210_2.fastq.gz | Fastqsanger.gz | |||
| ERR1354220_1.fastq.gz | Fastqsanger.gz | |||
| ERR1354220_2.fastq.gz | Fastqsanger.gz | |||
| ERR1354230_1.fastq.gz | Fastqsanger.gz |
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Project description:Replication of the eukaryotic genome occurs in the context of chromatin, a nucleoprotein packaging state consisting of repeating nucleosomes. Chromatin is commonly thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture that occur during genomic replication. Here, we sought to directly measure a key aspect of chromatin structure dynamics during replication â how rapidly nucleosome positions are established on the newly-replicated daughter genomes. By isolating newly-synthesized DNA marked with the nucleotide analogue EdU, we characterize nucleosome positions on both daughter genomes of budding yeast during a time course of chromatin maturation. We find that nucleosomes rapidly adopt their mid log positions at highly-transcribed genes, and that this process was impaired upon treatment with the transcription inhibitor thiolutin, consistent with a role for transcription in positioning nucleosomes in vivo. Additionally, experiments in the Hir1Î background reveal a role for HIR in nucleosome spacing. Using strand-specific EdU libraries, we characterize nucleosome positions on the leading and lagging strand daughter genomes, uncovering differences in chromatin maturation dynamics between the two daughter genomes at hundreds of genes. Our data define the maturation dynamics of newly-replicated chromatin, and support a role for transcription in sculpting the chromatin template. We have mapped changes in nucleosome positions on newly replicated DNA in a timecourse after genome replication. We have used Micrococcal Nuclease footprinting of cross linked chromatin to determine nucleosome positions and EdU (ethylene deoxy uridine) to mark nascent DNA strands. EdU incorporated into nascent DNA strands was biotinylated with Click chemistry and nascent DNA strand fragments were subsequently isolated using Streptavidin coated magnetic beads.
2016-08-25 | E-GEOD-74090 | biostudies-arrayexpress
Project description:Replication of the eukaryotic genome occurs in the context of chromatin, a nucleoprotein packaging state consisting of repeating nucleosomes. Chromatin is commonly thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture that occur during genomic replication. Here, we sought to directly measure a key aspect of chromatin structure dynamics during replication â?? how rapidly nucleosome positions are established on the newly-replicated daughter genomes. By isolating newly-synthesized DNA marked with the nucleotide analogue EdU, we characterize nucleosome positions on both daughter genomes of budding yeast during a time course of chromatin maturation. We find that nucleosomes rapidly adopt their mid log positions at highly-transcribed genes, and that this process was impaired upon treatment with the transcription inhibitor thiolutin, consistent with a role for transcription in positioning nucleosomes in vivo. Additionally, experiments in the Hir1-delta background reveal a role for HIR in nucleosome spacing. Using strand-specific EdU libraries, we characterize nucleosome positions on the leading and lagging strand daughter genomes, uncovering differences in chromatin maturation dynamics between the two daughter genomes at hundreds of genes. Our data define the maturation dynamics of newly-replicated chromatin, and support a role for transcription in sculpting the chromatin template. Gene expression array.
2016-08-25 | E-GEOD-79384 | biostudies-arrayexpress
Project description:Replication of the eukaryotic genome occurs in the context of chromatin, a nucleoprotein packaging state consisting of repeating nucleosomes. Chromatin is commonly thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture that occur during genomic replication. Here, we sought to directly measure a key aspect of chromatin structure dynamics during replication – how rapidly nucleosome positions are established on the newly-replicated daughter genomes. By isolating newly-synthesized DNA marked with the nucleotide analogue EdU, we characterize nucleosome positions on both daughter genomes of budding yeast during a time course of chromatin maturation. We find that nucleosomes rapidly adopt their mid log positions at highly-transcribed genes, and that this process was impaired upon treatment with the transcription inhibitor thiolutin, consistent with a role for transcription in positioning nucleosomes in vivo. Additionally, experiments in the Hir1Δ background reveal a role for HIR in nucleosome spacing. Using strand-specific EdU libraries, we characterize nucleosome positions on the leading and lagging strand daughter genomes, uncovering differences in chromatin maturation dynamics between the two daughter genomes at hundreds of genes. Our data define the maturation dynamics of newly-replicated chromatin, and support a role for transcription in sculpting the chromatin template.
2016-08-25 | GSE74090 | GEO
Project description:Replication of the eukaryotic genome occurs in the context of chromatin, a nucleoprotein packaging state consisting of repeating nucleosomes. Chromatin is commonly thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture that occur during genomic replication. Here, we sought to directly measure a key aspect of chromatin structure dynamics during replication – how rapidly nucleosome positions are established on the newly-replicated daughter genomes. By isolating newly-synthesized DNA marked with the nucleotide analogue EdU, we characterize nucleosome positions on both daughter genomes of budding yeast during a time course of chromatin maturation. We find that nucleosomes rapidly adopt their mid log positions at highly-transcribed genes, and that this process was impaired upon treatment with the transcription inhibitor thiolutin, consistent with a role for transcription in positioning nucleosomes in vivo. Additionally, experiments in the Hir1-delta background reveal a role for HIR in nucleosome spacing. Using strand-specific EdU libraries, we characterize nucleosome positions on the leading and lagging strand daughter genomes, uncovering differences in chromatin maturation dynamics between the two daughter genomes at hundreds of genes. Our data define the maturation dynamics of newly-replicated chromatin, and support a role for transcription in sculpting the chromatin template.
2016-08-25 | GSE79384 | GEO
Project description:Eukaryotic chromosomes are composed of chromatin, in which regularly spaced nucleosomes containing ~147 bp of DNA are separated by linker DNA. Most eukaryotic cells have a characteristic average nucleosome spacing of ~190 bp, corresponding to a ~45 bp linker. However, cortical neurons have a shorter average spacing of ~165 bp. The significance of this atypical global chromatin organization is unclear. We have compared the chromatin structures of purified mouse dorsal root ganglia (DRG) neurons, cortical oligodendrocyte precursor cells (OPCs) and cortical astrocytes. DRG neurons have short average spacing (~165 bp), whereas OPCs (~182 bp) and astrocytes (~183 bp) have longer spacing. We measured nucleosome positions by MNase-seq and gene expression by RNA-seq. Most genes in all three cell types have a promoter chromatin organization typical of active genes: a nucleosome-depleted region at the promoter flanked by regularly spaced nucleosomes phased relative to the transcription start site. In DRG neurons, the spacing of phased nucleosomes downstream of promoters (~178 bp) is longer than expected from the genomic average, whereas phased nucleosome spacing in OPCs and astrocytes is similar to the global average (~183 bp). Thus, the atypical nucleosome spacing of neuronal chromatin does not extend to promoter-proximal regions.
2019-10-07 | GSE133966 | GEO
Project description:Eukaryotic chromosomes are composed of chromatin, in which regularly spaced nucleosomes containing ~147 bp of DNA are separated by linker DNA. Most eukaryotic cells have a characteristic average nucleosome spacing of ~190 bp, corresponding to a ~45 bp linker. However, cortical neurons have a shorter average spacing of ~165 bp. The significance of this atypical global chromatin organization is unclear. We have compared the chromatin structures of purified mouse dorsal root ganglia (DRG) neurons, cortical oligodendrocyte precursor cells (OPCs) and cortical astrocytes. DRG neurons have short average spacing (~165 bp), whereas OPCs (~182 bp) and astrocytes (~183 bp) have longer spacing. We measured nucleosome positions by MNase-seq and gene expression by RNA-seq. Most genes in all three cell types have a promoter chromatin organization typical of active genes: a nucleosome-depleted region at the promoter flanked by regularly spaced nucleosomes phased relative to the transcription start site. In DRG neurons, the spacing of phased nucleosomes downstream of promoters (~178 bp) is longer than expected from the genomic average, whereas phased nucleosome spacing in OPCs and astrocytes is similar to the global average (~183 bp). Thus, the atypical nucleosome spacing of neuronal chromatin does not extend to promoter-proximal regions.
2019-10-07 | GSE133745 | GEO
Project description:We generated S.cerevisiae strains in which endogenous copies of candidate nucleosome spacing factors were replaced with the K.lactis copies. With this candidate approach, we found that K.lactis Chd1 directed longer nucleosome repeat length in S.cerevisiae. Generating chimeric proteins revealed that the strongest contribution to this differential spacing lies in the undercharacterised N-terminus of Chd1.
2015-03-18 | GSE66979 | 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:During every cell cycle, both the genome and the associated chromatin must be accurately replicated. Chromatin Assembly Factor-1 (CAF-1) is a key regulator of chromatin replication, but how CAF-1 functions in relation to the DNA replication machinery is unknown. Here, we reveal that this crosstalk differs between the leading and lagging strand at replication forks. Using biochemical reconstitution, we show that DNA and histones promote CAF-1 recruitment to its binding partner PCNA and reveal that two CAF-1 complexes are required for efficient nucleosome assembly under these conditions. Remarkably, in the context of the replisome, CAF-1 competes with the leading strand DNA polymerase epsilon (Pole) for PCNA binding. However, CAF-1 does not affect the activity of the lagging strand DNA polymerase Delta (Pold). Yet, in cells, CAF-1 deposits newly synthesized histones equally on both daughter strands. Thus, on the leading strand, chromatin assembly by CAF-1 cannot occur simultaneously to DNA synthesis, while on the lagging strand these processes may be coupled. We propose that these differences may facilitate distinct parental histone recycling mechanisms and accommodate the inherent asymmetry of DNA replication.
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