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ISWI and CHD chromatin remodelers bind promoters but act in gene bodies.

Zentner GE, Tsukiyama T, Henikoff S - PLoS Genet. (2013)

Bottom Line: Although these remodelers act in gene bodies, we find that they are also highly enriched at nucleosome-depleted regions (NDRs), where they bind to extended regions of DNA adjacent to particular transcription factors.Surprisingly, catalytically inactive remodelers show similar binding patterns.We find that remodeler occupancy at NDRs and gene bodies is associated with nucleosome turnover and transcriptional elongation rate, suggesting that remodelers act on regions of transient nucleosome unwrapping or depletion within gene bodies subsequent to transcriptional elongation.

View Article: PubMed Central - PubMed

Affiliation: Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.

ABSTRACT
ATP-dependent nucleosome remodelers influence genetic processes by altering nucleosome occupancy, positioning, and composition. In vitro, Saccharomyces cerevisiae ISWI and CHD remodelers require ∼30-85 bp of extranucleosomal DNA to reposition nucleosomes, but linker DNA in S. cerevisiae averages <20 bp. To address this discrepancy between in vitro and in vivo observations, we have mapped the genomic distributions of the yeast Isw1, Isw2, and Chd1 remodelers at base-pair resolution on native chromatin. Although these remodelers act in gene bodies, we find that they are also highly enriched at nucleosome-depleted regions (NDRs), where they bind to extended regions of DNA adjacent to particular transcription factors. Surprisingly, catalytically inactive remodelers show similar binding patterns. We find that remodeler occupancy at NDRs and gene bodies is associated with nucleosome turnover and transcriptional elongation rate, suggesting that remodelers act on regions of transient nucleosome unwrapping or depletion within gene bodies subsequent to transcriptional elongation.

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Comparison of Isw2 N-ChIP and X-ChIP data.(A) Aggregate profiles of log2(Isw2 IP/input) at sites bound by catalytically inactive (K227R) Isw2 in ChIP-chip experiments (2128 sites), sites with altered nucleosome positioning in an isw2Δ strain (1399 sites), sites bound by wild-type Isw2 in ChIP-exo experiments (1251 sites), and around random nucleosomes (1399 sites).Also shown are profiles of remodeler binding at the (B) chrIII recombination enhancer and (C) chrIII centromere (marked by vertical lines).
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pgen-1003317-g002: Comparison of Isw2 N-ChIP and X-ChIP data.(A) Aggregate profiles of log2(Isw2 IP/input) at sites bound by catalytically inactive (K227R) Isw2 in ChIP-chip experiments (2128 sites), sites with altered nucleosome positioning in an isw2Δ strain (1399 sites), sites bound by wild-type Isw2 in ChIP-exo experiments (1251 sites), and around random nucleosomes (1399 sites).Also shown are profiles of remodeler binding at the (B) chrIII recombination enhancer and (C) chrIII centromere (marked by vertical lines).

Mentions: N-ChIP-seq revealed specific sites of enrichment for all three remodelers throughout the yeast genome (Figure 1B and Figure S1). We then tested if we could recapitulate known features of remodeler-genome association by N-ChIP-seq. We focused on Isw2, which has previously been characterized by genome-wide ChIP-chip [20], [31] and the recently developed ChIP-exo method [17]. ChIP-exo is a modification of the standard ChIP-seq protocol employing exonuclease digestion to improve the resolution of crosslinking ChIP-seq. Our N-ChIP-seq methodology is distinct from these techniques in that it does not rely on formaldehyde fixation, which also crosslinks primary amines to fix protein-protein interactions. We first obtained lists of Isw2 peaks determined by ChIP-chip [20] and ChIP-exo [17] as well as sites with altered nucleosome positioning in an isw2Δ strain [20]. We then determined the input-normalized N-ChIP-seq signal for each base pair in a 2-kb window centered on each ChIP or remodeling site and averaged the signal for each base pair within the window for each class of sites (Isw2 X-ChIP-chip, Isw2 ChIP-exo, and Isw2 remodeling) to generate average N-ChIP-seq profiles. We detected enrichment of Isw2 by N-ChIP-seq at sites bound by Isw2 in both X-ChIP-chip and ChIP-exo and sites of chromatin remodeling by Isw2 but not around randomly selected nucleosomes (Figure 2A). The two peaks on either side of the midpoint represent robust enrichment of Isw2 on either side of transcription factor binding sites. We note that the enrichment of Isw2 at these sites does not imply an effect on expression of nearby genes, only that our technique is able to replicate Isw2 binding sites previously determined by X-ChIP methodologies.


ISWI and CHD chromatin remodelers bind promoters but act in gene bodies.

Zentner GE, Tsukiyama T, Henikoff S - PLoS Genet. (2013)

Comparison of Isw2 N-ChIP and X-ChIP data.(A) Aggregate profiles of log2(Isw2 IP/input) at sites bound by catalytically inactive (K227R) Isw2 in ChIP-chip experiments (2128 sites), sites with altered nucleosome positioning in an isw2Δ strain (1399 sites), sites bound by wild-type Isw2 in ChIP-exo experiments (1251 sites), and around random nucleosomes (1399 sites).Also shown are profiles of remodeler binding at the (B) chrIII recombination enhancer and (C) chrIII centromere (marked by vertical lines).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3585014&req=5

pgen-1003317-g002: Comparison of Isw2 N-ChIP and X-ChIP data.(A) Aggregate profiles of log2(Isw2 IP/input) at sites bound by catalytically inactive (K227R) Isw2 in ChIP-chip experiments (2128 sites), sites with altered nucleosome positioning in an isw2Δ strain (1399 sites), sites bound by wild-type Isw2 in ChIP-exo experiments (1251 sites), and around random nucleosomes (1399 sites).Also shown are profiles of remodeler binding at the (B) chrIII recombination enhancer and (C) chrIII centromere (marked by vertical lines).
Mentions: N-ChIP-seq revealed specific sites of enrichment for all three remodelers throughout the yeast genome (Figure 1B and Figure S1). We then tested if we could recapitulate known features of remodeler-genome association by N-ChIP-seq. We focused on Isw2, which has previously been characterized by genome-wide ChIP-chip [20], [31] and the recently developed ChIP-exo method [17]. ChIP-exo is a modification of the standard ChIP-seq protocol employing exonuclease digestion to improve the resolution of crosslinking ChIP-seq. Our N-ChIP-seq methodology is distinct from these techniques in that it does not rely on formaldehyde fixation, which also crosslinks primary amines to fix protein-protein interactions. We first obtained lists of Isw2 peaks determined by ChIP-chip [20] and ChIP-exo [17] as well as sites with altered nucleosome positioning in an isw2Δ strain [20]. We then determined the input-normalized N-ChIP-seq signal for each base pair in a 2-kb window centered on each ChIP or remodeling site and averaged the signal for each base pair within the window for each class of sites (Isw2 X-ChIP-chip, Isw2 ChIP-exo, and Isw2 remodeling) to generate average N-ChIP-seq profiles. We detected enrichment of Isw2 by N-ChIP-seq at sites bound by Isw2 in both X-ChIP-chip and ChIP-exo and sites of chromatin remodeling by Isw2 but not around randomly selected nucleosomes (Figure 2A). The two peaks on either side of the midpoint represent robust enrichment of Isw2 on either side of transcription factor binding sites. We note that the enrichment of Isw2 at these sites does not imply an effect on expression of nearby genes, only that our technique is able to replicate Isw2 binding sites previously determined by X-ChIP methodologies.

Bottom Line: Although these remodelers act in gene bodies, we find that they are also highly enriched at nucleosome-depleted regions (NDRs), where they bind to extended regions of DNA adjacent to particular transcription factors.Surprisingly, catalytically inactive remodelers show similar binding patterns.We find that remodeler occupancy at NDRs and gene bodies is associated with nucleosome turnover and transcriptional elongation rate, suggesting that remodelers act on regions of transient nucleosome unwrapping or depletion within gene bodies subsequent to transcriptional elongation.

View Article: PubMed Central - PubMed

Affiliation: Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.

ABSTRACT
ATP-dependent nucleosome remodelers influence genetic processes by altering nucleosome occupancy, positioning, and composition. In vitro, Saccharomyces cerevisiae ISWI and CHD remodelers require ∼30-85 bp of extranucleosomal DNA to reposition nucleosomes, but linker DNA in S. cerevisiae averages <20 bp. To address this discrepancy between in vitro and in vivo observations, we have mapped the genomic distributions of the yeast Isw1, Isw2, and Chd1 remodelers at base-pair resolution on native chromatin. Although these remodelers act in gene bodies, we find that they are also highly enriched at nucleosome-depleted regions (NDRs), where they bind to extended regions of DNA adjacent to particular transcription factors. Surprisingly, catalytically inactive remodelers show similar binding patterns. We find that remodeler occupancy at NDRs and gene bodies is associated with nucleosome turnover and transcriptional elongation rate, suggesting that remodelers act on regions of transient nucleosome unwrapping or depletion within gene bodies subsequent to transcriptional elongation.

Show MeSH
Related in: MedlinePlus