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Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions [Affymetrix]

ABSTRACT: Expression data from 3134 mouse mammary epithelial cell line -/+ Tet-inducible chromatin remodeler mutant variants ATP-dependent chromatin remodeling is an essential process required for the dynamic organization of chromatin structure. Here we describe the genome-wide location and activity of three remodeler proteins with diverse physiological functions in the mouse genome: Brg1, Chd4 and Snf2h. The localization patterns of all three proteins substantially overlap with one another and with regions of accessible chromatin. Furthermore, using inducible mutant variants, we demonstrate that the catalytic activity of these proteins contributes to the remodeling of chromatin genome wide and that each of these remodelers can independently regulate chromatin reorganization at distinct sites. Many regions require the activity of more than one remodeler to regulate accessibility. We also examined effects of mutant remodeler variants on selective gene expression by global analysis of RNA expression patterns and found more than 800 genes are deregulated by these variants. These findings provide a dynamic view of chromatin organization and highlight the differential contributions of remodelers to chromatin maintenance in higher eukaryotes.

ORGANISM(S): Mus musculus

PROVIDER: GSE66544 | GEO | 2015/03/05

SECONDARY ACCESSION(S): PRJNA277304

REPOSITORIES: GEO

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Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions [Affymetrix]

Project description:Expression data from 3134 mouse mammary epithelial cell line -/+ Tet-inducible chromatin remodeler mutant variants ATP-dependent chromatin remodeling is an essential process required for the dynamic organization of chromatin structure. Here we describe the genome-wide location and activity of three remodeler proteins with diverse physiological functions in the mouse genome: Brg1, Chd4 and Snf2h. The localization patterns of all three proteins substantially overlap with one another and with regions of accessible chromatin. Furthermore, using inducible mutant variants, we demonstrate that the catalytic activity of these proteins contributes to the remodeling of chromatin genome wide and that each of these remodelers can independently regulate chromatin reorganization at distinct sites. Many regions require the activity of more than one remodeler to regulate accessibility. We also examined effects of mutant remodeler variants on selective gene expression by global analysis of RNA expression patterns and found more than 800 genes are deregulated by these variants. These findings provide a dynamic view of chromatin organization and highlight the differential contributions of remodelers to chromatin maintenance in higher eukaryotes. Flag tagged dominant negative variants of the chromatin remodelers Brg1, Chd4, and Snf2h were each stably integrated into a cell line containg the tetracycline transactivator regulatory system (3134Tet (7110)). The 3134Tet control cells and each of the dominant-negative remodeler cell lines were grown in media with or without tetracycline for 48 hrs to induce expression of the tetracycline transactivator alone (3134Tet cells) or tetracycline transactivator and dominant negative remodeler variant. Following treatment, cells were harvested for RNA extraction and hybridization to Affymetrix Mouse Gene 1.0 ST arrays.

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Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions (ChIP-Seq)

Project description:ATP-dependent chromatin remodeling is an essential process required for the dynamic organization of chromatin structure. Here we describe the genome-wide location and activity of three remodeler proteins with diverse physiological functions in the mouse genome: Brg1, Chd4, and Snf2h. The localization patterns of all three proteins significantly overlap with one another and with regions of accessible chromatin. Furthermore, using inducible mutant variants, we demonstrate that the catalytic activity of these proteins contributes to the remodeling of chromatin genome-wide, and that each of these remodelers can independently regulate chromatin reorganization at distinct sites. Most regions require the activity of more than one remodeler to regulate accessibility. These findings provide a dynamic view of chromatin organization, and highlight the differential contributions of remodelers to chromatin maintenance in higher eukaryotes. We examine the genome-wide location and activity of three remodeler proteins (Brg1, CHD4 and Snf2h)

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Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions (Dnase-Seq)

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Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions (Dnase-Seq)

2013-12-24 | GSE53584 | GEO

Multiple Overlapping Mammalian Chromatin Remodeling Systems Collaborate Genome-wide at Dynamic Chromatin Transitions (ChIP-Seq)

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Remodelers organize cellular chromatin by counteracting intrinsic histone-DNA sequence preferences in a class-specific manner

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Project description:ATP-dependent chromatin remodelers control the accessibility of genomic DNA through nucleosome mobilization. However, the dynamics of genome exploration by remodelers, and the role of ATP hydrolysis in this process remain unclear. We used live-cell imaging of Drosophila polytene nuclei to monitor Brahma (BRM) remodeler interactions with its chromosomal targets. In parallel, we measured local chromatin condensation and its effect on BRM association. Surprisingly, only a small portion of BRM is bound to chromatin at any given time. BRM binds decondensed chromatin but is excluded from condensed chromatin, limiting its genomic search space. BRM-chromatin interactions are highly dynamic, whereas histone-exchange is limited and much slower. Intriguingly, loss of ATP hydrolysis enhanced chromatin retention and clustering of BRM, which was associated with reduced histone turnover. Thus, ATP hydrolysis couples nucleosome remodeling to remodeler release, driving a continuous transient probing of the genome.

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Genome information processing by the INO80 chromatin remodeler positions nucleosomes

Project description:The mechanistics by which ATP-dependent chromatin remodelers process (epi)genetic information to generate hallmark features of chromatin are fundamental for genome regulation. Here, we advance whole-genome reconstitutions into a fully definable, recombinant approach for probing how remodelers determine nucleosome positioning. Using wild type and structure-guided mutant versions of the Saccharomyces cerevisiae 15-subunit INO80 remodeler, we show that INO80-intrinsic nucleosome positioning relies on direct DNA shape read-out. This requires especially the interplay between the INO80 nuclear actin and core modules, to a lesser extent the HMG-box Nhp10 module and not histone modifications/variants or the Rvb1/2 AAA+-ATPase activity. Extrinsically, nucleosome positioning is guided by the barrier Reb1 via Ino80 ATPase activity modulation or by mere DNA double strand breaks. Taken together, we present a defined and unbiased approach to understand remodeler-mediated nucleosome positioning and delineate how INO80 processes genomic information into nucleosome positioning.

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Mammalian SWI/SNF continuously restores local accessibility to chromatin [MNase-Seq]

Project description:Open chromatin is a hallmark of regulatory regions and serves as a feature for their identification. Accessible genomic sites are mediated by the activity of transcription factors (TFs) that can engage with nucleosomes to enable stable gene expression changes that underlie development. While the variability of open chromatin between tissues has been studied in large detail, we have limited knowledge on the dynamics of accessible chromatin in any given cell. Answering this question is critical to better understand cellular memory but also to discern actual kinetics of TF binding and its consequence on local nucleosomal organization. Here we use small molecule inhibition of the catalytic subunit of the mouse SWI/SNF remodeler complex to show that accessibility at sites of TF binding critically relies on persistent activity of nucleosome remodelers. Within minutes of remodeler inhibition, accessibility decreases excluding that remodeler activity is only necessary in defined cell cycle transitions. Moreover, this surprisingly fast response is irrespective of TF function as it is similar for the pluripotency factor and activating TF Oct4 and the repressive TF Rest. Importantly, accessibility is rapidly restored upon inhibitor removal demonstrating that open chromatin is regenerated continuously and in a cell-autonomous fashion. Combined, this establishes a model where TF binding to chromatin and remodeler-mediated nucleosomal removal do not represent an initial opening event leading to a stable situation but instead open chromatin reflects an average of a highly dynamic situation that requires continued reestablishment. This argues against a model where open chromatin can persist in absence of continuous TF binding and remodeler activity.

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