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Isw2 and Ino80 chromatin remodeling factors regulate chromatin, replication, and copy number at the yeast ribosomal DNA locus

ABSTRACT: Isw2 and Ino80 chromatin remodeling factors regulate chromatin, replication, and copy number at the yeast ribosomal DNA locus

PROVIDER: PRJNA446650 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR6908150.fastq.gz	Fastqsanger.gz
	SRR6908160.fastq.gz	Fastqsanger.gz
	SRR6908170.fastq.gz	Fastqsanger.gz
	SRR6908190.fastq.gz	Fastqsanger.gz
	SRR6908200.fastq.gz	Fastqsanger.gz

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Chromatin Remodeling Factors Isw2 and Ino80 Regulate Chromatin, Replication, and Copy Number of the Saccharomyces cerevisiae Ribosomal DNA Locus.

Project description:In the budding yeast Saccharomyces cerevisiae, ribosomal RNA genes are encoded in a highly repetitive tandem array referred to as the ribosomal DNA (rDNA) locus. The yeast rDNA is the site of a diverse set of DNA-dependent processes, including transcription of ribosomal RNAs by RNA polymerases I and III, transcription of noncoding RNAs by RNA polymerase II, DNA replication initiation, replication fork blocking, and recombination-mediated regulation of rDNA repeat copy number. All of this takes place in the context of chromatin, but little is known about the roles played by ATP-dependent chromatin remodeling factors at the yeast rDNA. In this work, we report that the Isw2 and Ino80 chromatin remodeling factors are targeted to this highly repetitive locus. We characterize for the first time their function in modifying local chromatin structure, finding that loss of these factors decreases the fraction of actively transcribed 35S ribosomal RNA genes and the positioning of nucleosomes flanking the ribosomal origin of replication. In addition, we report that Isw2 and Ino80 promote efficient firing of the ribosomal origin of replication and facilitate the regulated increase of rDNA repeat copy number. This work significantly expands our understanding of the importance of ATP-dependent chromatin remodeling for rDNA biology.

| S-EPMC6283175 | biostudies-literature

Genome-wide maps of chromatin remodeling factors Isw2 and Ino80 in asynchronously growing yeast cells

Project description:ChIP-seq of Isw2 (subunit of Isw2 complex) and Nhp10 (subunit of Ino80 complex) was performed on two independent isolates of FLAG-tagged Isw2 or Nhp10 Sample naming convention: <ChIP target>_<growth condition>_<genotype-replicate>_<strain name>

2018-10-30 | GSE112423 | GEO

Isw2 and Ino80 chromatin remodeling factors regulate chromatin, replication, and copy number at the yeast ribosomal DNA locus

Project description:This SuperSeries is composed of the SubSeries listed below.

2018-10-30 | GSE112465 | GEO

Genome-wide maps of chromatin remodeling factors Isw2 and Ino80 in asynchronously growing yeast cells

Project description:Genome-wide maps of chromatin remodeling factors Isw2 and Ino80 in asynchronously growing yeast cells

| PRJNA445972 | ENA

Yeast replication in the presence of MMS: WT and isw2 nhp10 strains

Project description:The eukaryotic DNA replication machinery must traverse every nucleosome present in the eukaryotic genome during S phase. Since nucleosomes are generally inhibitory to DNA-dependent processes, it is thought that chromatin structure must undergo extensive reorganization to facilitate DNA synthesis. However, the identity of chromatin-remodeling factors involved in replication and how they affect DNA synthesis is largely unknown. Here we show that two ATP-dependent chromatin-remodeling complexes in Saccharomyces cerevisiae, Isw2 and Ino80, function in parallel to promote DNA replication, especially during periods of replication stress. The rate of replication-fork progression is slowed when both chromatin-remodeling pathways are compromised. Both Isw2 and Ino80 complexes are recruited to actively replicating chromatin suggesting that these chromatin-remodeling complexes act directly to promote replication-fork progression. These findings support an important role for ATP-dependent chromatin-remodeling complexes in promoting DNA replication, and define specific stages of replication that require remodeling activity for normal function. A critical part of this study was the use of microarrays to measure DNA replication progression throughout the genome during MMS treatment. These samples are from experiments where we competed newly replicated DNA against unreplicated DNA on the same microarrays. Keywords: Time course

2007-11-30 | GSE9122 | GEO

Mapping chromatin remodelling in delta isw2 cells

Project description:To define chromatin structure changes along the yeast genome using microarrays. Nucleosomal DNA from WT and delta isw2 yeast were hybridized and differences in signals were calculated. Keywords: Nucleosomal DNA hybridization

2007-09-10 | GSE8813 | GEO

Histone octamer structure is altered early in ISW2 ATP-dependent remodeling of nucleosomes

Project description:Histone octamers are thought to be a rigid part of nucleosomes that shape chromatin and block cellular machinery from accessing DNA. ATP-dependent chromatin remodelers like ISW2 mobilize nucleosomes to provide DNA access. We find evidence for histone octamer distortion preceding DNA being moved into nucleosomes and processive movement of the ATPase motor of ISW2 on nucleosomal DNA. DNA entering nucleosome is uncoupled from the ATPase activity of ISW2 and alterations of the histone octamer structure mediated by ISW2 by deletion of the SANT domain from the C-terminus of the Isw2 catalytic subunit. We also find that restricting histone movement by chemical crosslinking traps remodeling intermediates resembling those seen by loss of the SANT domain, further supporting the importance of changes in histone octamer structure early in ISW2 remodeling. Transient octamer distortions are stabilized by H3-H4 tetramer disulfide crosslinking, thereby linking intrinsic histone octamer flexibility to chromatin remodeling.

2020-05-27 | PXD013967 | Pride

Stepwise reconstitution of genomic nucleosome organization with pure proteins

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

Chromatin remodeling factors Isw2 and Ino80 regulate checkpoint activity and chromatin structure in S phase.

Project description:When cells undergo replication stress, proper checkpoint activation and deactivation are critical for genomic stability and cell survival and therefore must be highly regulated. Although mechanisms of checkpoint activation are well studied, mechanisms of checkpoint deactivation are far less understood. Previously, we reported that chromatin remodeling factors Isw2 and Ino80 attenuate the S-phase checkpoint activity in Saccharomyces cerevisiae, especially during recovery from hydroxyurea. In this study, we found that Isw2 and Ino80 have a more pronounced role in attenuating checkpoint activity during late S phase in the presence of methyl methanesulfonate (MMS). We therefore screened for checkpoint factors required for Isw2 and Ino80 checkpoint attenuation in the presence of MMS. Here we demonstrate that Isw2 and Ino80 antagonize checkpoint activators and attenuate checkpoint activity in S phase in MMS either through a currently unknown pathway or through RPA. Unexpectedly, we found that Isw2 and Ino80 increase chromatin accessibility around replicating regions in the presence of MMS through a novel mechanism. Furthermore, through growth assays, we provide additional evidence that Isw2 and Ino80 partially counteract checkpoint activators specifically in the presence of MMS. Based on these results, we propose that Isw2 and Ino80 attenuate S-phase checkpoint activity through a novel mechanism.

| S-EPMC4391558 | biostudies-literature

Hap2-Ino80 mediated transcription facilitates de novo establishment of CENP-A chromatin

Project description:Centromeres are maintained epigenetically by the presence of CENP-A, an evolutionarily-conserved histone H3 variant, which directs kinetochore assembly and hence, centromere function. To identify factors that promote assembly of CENP-A chromatin, we affinity selected solubilised fission yeast CENP-ACnp1 chromatin. All subunits of the Ino80 complex were enriched, including the auxiliary subunit Hap2. In addition to a role in maintenance of CENP-ACnp1 chromatin integrity at endogenous centromeres, Hap2 is required for de novo assembly of CENP-ACnp1 chromatin on naïve centromere DNA and promotes H3 turnover on centromere regions and other loci prone to CENP-ACnp1 deposition. Prior to CENP-ACnp1 chromatin assembly, Hap2 is required for transcription from centromere DNA. These analyses suggest that Hap2-Ino80 destabilises H3 nucleosomes on centromere DNA through transcription-mediated histone H3 turnover, driving the replacement of resident H3 nucleosomes with CENP-ACnp1 nucleosomes. These inherent properties define centromere DNA by directing a program that mediates CENP-ACnp1 assembly on appropriate sequences.

2020-05-27 | PXD016602 | Pride

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