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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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ISWI and CHD remodeler association with TFBSs is ATP-independent.(A) Size distributions of mapped paired-end 2.5′ MNase-digested wild-type and catalytically inactive K227R Isw1 ChIP and input fragments. Similar profiles were seen for Isw2 K215R and Chd1 K407R (Figure S4). (B) Profiles of wild-type and K227R Isw1 binding along a representative segment of the genome. (C) V-plots of wild-type and K227R Isw1 ChIP and soluble input chromatin at binding sites for the Reb1 TF, determined by ChIP-exo, after 2.5′ MNase digestion. The overall fragment size in the Isw1 K227R ChIP and input samples is slightly reduced when compared to wild-type, indicative of technical variation in MNase digestion. Similar results were seen for K227R Isw1 and catalytically inactive Isw2 (K215R) and Chd1 (K407R) at Abf1 and other TFBSs (Figures S4 and S6).
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pgen-1003317-g005: ISWI and CHD remodeler association with TFBSs is ATP-independent.(A) Size distributions of mapped paired-end 2.5′ MNase-digested wild-type and catalytically inactive K227R Isw1 ChIP and input fragments. Similar profiles were seen for Isw2 K215R and Chd1 K407R (Figure S4). (B) Profiles of wild-type and K227R Isw1 binding along a representative segment of the genome. (C) V-plots of wild-type and K227R Isw1 ChIP and soluble input chromatin at binding sites for the Reb1 TF, determined by ChIP-exo, after 2.5′ MNase digestion. The overall fragment size in the Isw1 K227R ChIP and input samples is slightly reduced when compared to wild-type, indicative of technical variation in MNase digestion. Similar results were seen for K227R Isw1 and catalytically inactive Isw2 (K215R) and Chd1 (K407R) at Abf1 and other TFBSs (Figures S4 and S6).

Mentions: We next asked whether remodeler inactivation would alter binding around TFBSs. Surprisingly, elimination of remodeler catalytic activity via a lysine-to-arginine substitution in the conserved GXGKT ATP-binding motif [21], [44] did not substantially affect the size distribution of mapped fragments (Figure 5A and Figure S4) or the genomic binding profiles of remodelers (Figure 5B). We assessed the effect of remodeler catalytic inactivation by V-plotting wild-type and catalytically inactive (K227R) ChIP and input data at sites for the Reb1 TF generated by ChIP-exo [45], which also allowed us to assess V-plot patterns using an independent set of binding sites. V-plot patterns generated with ChIP-exo Reb1 sites were nearly indistinguishable from those created using Reb1 ChIP-chip sites (Figure 5C, Figure S3, Figure S5). Catalytic inactivation of remodelers also had no noticeable effect on V-plot patterns at TFBSs (Figure 5C, Figure S3, and Figure S5), implying that remodeler association with TFBSs is ATP-independent.


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

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

ISWI and CHD remodeler association with TFBSs is ATP-independent.(A) Size distributions of mapped paired-end 2.5′ MNase-digested wild-type and catalytically inactive K227R Isw1 ChIP and input fragments. Similar profiles were seen for Isw2 K215R and Chd1 K407R (Figure S4). (B) Profiles of wild-type and K227R Isw1 binding along a representative segment of the genome. (C) V-plots of wild-type and K227R Isw1 ChIP and soluble input chromatin at binding sites for the Reb1 TF, determined by ChIP-exo, after 2.5′ MNase digestion. The overall fragment size in the Isw1 K227R ChIP and input samples is slightly reduced when compared to wild-type, indicative of technical variation in MNase digestion. Similar results were seen for K227R Isw1 and catalytically inactive Isw2 (K215R) and Chd1 (K407R) at Abf1 and other TFBSs (Figures S4 and S6).
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pgen-1003317-g005: ISWI and CHD remodeler association with TFBSs is ATP-independent.(A) Size distributions of mapped paired-end 2.5′ MNase-digested wild-type and catalytically inactive K227R Isw1 ChIP and input fragments. Similar profiles were seen for Isw2 K215R and Chd1 K407R (Figure S4). (B) Profiles of wild-type and K227R Isw1 binding along a representative segment of the genome. (C) V-plots of wild-type and K227R Isw1 ChIP and soluble input chromatin at binding sites for the Reb1 TF, determined by ChIP-exo, after 2.5′ MNase digestion. The overall fragment size in the Isw1 K227R ChIP and input samples is slightly reduced when compared to wild-type, indicative of technical variation in MNase digestion. Similar results were seen for K227R Isw1 and catalytically inactive Isw2 (K215R) and Chd1 (K407R) at Abf1 and other TFBSs (Figures S4 and S6).
Mentions: We next asked whether remodeler inactivation would alter binding around TFBSs. Surprisingly, elimination of remodeler catalytic activity via a lysine-to-arginine substitution in the conserved GXGKT ATP-binding motif [21], [44] did not substantially affect the size distribution of mapped fragments (Figure 5A and Figure S4) or the genomic binding profiles of remodelers (Figure 5B). We assessed the effect of remodeler catalytic inactivation by V-plotting wild-type and catalytically inactive (K227R) ChIP and input data at sites for the Reb1 TF generated by ChIP-exo [45], which also allowed us to assess V-plot patterns using an independent set of binding sites. V-plot patterns generated with ChIP-exo Reb1 sites were nearly indistinguishable from those created using Reb1 ChIP-chip sites (Figure 5C, Figure S3, Figure S5). Catalytic inactivation of remodelers also had no noticeable effect on V-plot patterns at TFBSs (Figure 5C, Figure S3, and Figure S5), implying that remodeler association with TFBSs is ATP-independent.

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