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Chd1 co-localizes with early transcription elongation factors independently of H3K36 methylation and releases stalled RNA polymerase II at introns.

Park D, Shivram H, Iyer VR - Epigenetics Chromatin (2014)

Bottom Line: Using genome-wide approaches, we found that the loss of Chd1 significantly disrupted nucleosome arrays within the gene bodies of highly transcribed genes.We also found that Chd1 is physically recruited to gene bodies, and that its occupancy specifically corresponds to that of the early elongating form of RNA polymerase, RNAPII Ser 5-P.We also found that deletion of the histone methyltransferase for H3K36 (SET2) did not affect either Chd1 occupancy or nucleosome organization genome-wide.

View Article: PubMed Central - PubMed

Affiliation: Center for Systems and Synthetic Biology, Institute for Cellular and Molecular Biology, Department of Molecular Biosciences, University of Texas, 2500 Speedway, Austin, TX 78712 USA.

ABSTRACT

Background: Chromatin consists of ordered nucleosomal arrays that are controlled by highly conserved adenosine triphosphate (ATP)-dependent chromatin remodeling complexes. One such remodeler, chromodomain helicase DNA binding protein 1 (Chd1), is believed to play an integral role in nucleosomal organization, as the loss of Chd1 is known to disrupt chromatin. However, the specificity and basis for the functional and physical localization of Chd1 on chromatin remains largely unknown.

Results: Using genome-wide approaches, we found that the loss of Chd1 significantly disrupted nucleosome arrays within the gene bodies of highly transcribed genes. We also found that Chd1 is physically recruited to gene bodies, and that its occupancy specifically corresponds to that of the early elongating form of RNA polymerase, RNAPII Ser 5-P. Conversely, RNAPII Ser 5-P occupancy was affected by the loss of Chd1, suggesting that Chd1 is associated with early transcription elongation. Surprisingly, the occupancy of RNAPII Ser 5-P was affected by the loss of Chd1 specifically at intron-containing genes. Nucleosome turnover was also affected at these sites in the absence of Chd1. We also found that deletion of the histone methyltransferase for H3K36 (SET2) did not affect either Chd1 occupancy or nucleosome organization genome-wide.

Conclusions: Chd1 is specifically recruited onto the gene bodies of highly transcribed genes in an elongation-dependent but H3K36me3-independent manner. Chd1 co-localizes with the early elongating form of RNA polymerase, and affects the occupancy of RNAPII only at genes containing introns, suggesting a role in relieving splicing-related pausing of RNAPII.

No MeSH data available.


Related in: MedlinePlus

Nucleosome occupancy inchd1Δ.(A) Average nucleosome profile for all genes (n =5,207) shows that nucleosome occupancy is reduced at gene bodies in chd1Δ. The X-axis shows genomic position relative to the transcription start site (TSS). The Y-axis represents average read counts per million reads (M). (B) Example of smoothing MNase-seq read data using a spline function. Smoothing removes the noise caused by fluctuations in read coverage that can give erroneous correlation coefficients. Blue line is the raw, binned read count data and the red line is smoothed (C) Smoothing followed by Pearson correlation, called shapeDiff analysis, measures the nucleosome occupancy similarity as 0.31 between wild-type (WT) and chd1Δ over the transcribed region of RPL37B. MET14 has a high correlation coefficient between WT and chd1Δ as determined by shapeDiff due to high similarity of nucleosome occupancy profiles. (D) Average nucleosome profile for RP genes (n =136) shows strong nucleosome disruption upon deletion of CHD1. The shaded bands represent the 95% confidence interval of the data. (E) Quantitation of nucleosome disruption at ribosomal protein (RP) genes after heat shock or upon deletion of CHD1 (chd1Δ). The Y axis shows the correlation coefficient as measured by shapeDiff for genes in each class as indicated on the X-axis.
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Fig1: Nucleosome occupancy inchd1Δ.(A) Average nucleosome profile for all genes (n =5,207) shows that nucleosome occupancy is reduced at gene bodies in chd1Δ. The X-axis shows genomic position relative to the transcription start site (TSS). The Y-axis represents average read counts per million reads (M). (B) Example of smoothing MNase-seq read data using a spline function. Smoothing removes the noise caused by fluctuations in read coverage that can give erroneous correlation coefficients. Blue line is the raw, binned read count data and the red line is smoothed (C) Smoothing followed by Pearson correlation, called shapeDiff analysis, measures the nucleosome occupancy similarity as 0.31 between wild-type (WT) and chd1Δ over the transcribed region of RPL37B. MET14 has a high correlation coefficient between WT and chd1Δ as determined by shapeDiff due to high similarity of nucleosome occupancy profiles. (D) Average nucleosome profile for RP genes (n =136) shows strong nucleosome disruption upon deletion of CHD1. The shaded bands represent the 95% confidence interval of the data. (E) Quantitation of nucleosome disruption at ribosomal protein (RP) genes after heat shock or upon deletion of CHD1 (chd1Δ). The Y axis shows the correlation coefficient as measured by shapeDiff for genes in each class as indicated on the X-axis.

Mentions: We used MNase-seq to map nucleosome positions in wild-type (WT) and chd1Δ strains of budding yeast[4]. We found that the loss of Chd1 disrupted nucleosome organization within gene bodies, consistent with previous observations in both budding and fission yeast[10, 12, 13] (Figure 1A). We further confirmed this phenotype in the chd1Δ strain with a different resistance marker (Methods, [see Additional file1: Figure S1A]). Although these genome-wide profiles indicate that nucleosome occupancy is generally affected by Chd1, they do not reveal which subsets of genes are specifically dependent on Chd1 function, and what the molecular basis of this dependency might be. To gain insights into these questions, we developed an approach based on quantitatively scoring all genes by the extent of nucleosome disruption.In this approach, which we called ‘shapeDiff analysis,’ we first smoothed nucleosome occupancy signals based on read counts using a spline function, then measured the correlation coefficient between the WT and mutant nucleosome profiles for every gene in the genome (Methods). This approach has the advantage that the correlation measurement is relatively insensitive to noisy fluctuations in the nucleosome occupancy signal caused by low read counts (Figure 1B).Figure 1


Chd1 co-localizes with early transcription elongation factors independently of H3K36 methylation and releases stalled RNA polymerase II at introns.

Park D, Shivram H, Iyer VR - Epigenetics Chromatin (2014)

Nucleosome occupancy inchd1Δ.(A) Average nucleosome profile for all genes (n =5,207) shows that nucleosome occupancy is reduced at gene bodies in chd1Δ. The X-axis shows genomic position relative to the transcription start site (TSS). The Y-axis represents average read counts per million reads (M). (B) Example of smoothing MNase-seq read data using a spline function. Smoothing removes the noise caused by fluctuations in read coverage that can give erroneous correlation coefficients. Blue line is the raw, binned read count data and the red line is smoothed (C) Smoothing followed by Pearson correlation, called shapeDiff analysis, measures the nucleosome occupancy similarity as 0.31 between wild-type (WT) and chd1Δ over the transcribed region of RPL37B. MET14 has a high correlation coefficient between WT and chd1Δ as determined by shapeDiff due to high similarity of nucleosome occupancy profiles. (D) Average nucleosome profile for RP genes (n =136) shows strong nucleosome disruption upon deletion of CHD1. The shaded bands represent the 95% confidence interval of the data. (E) Quantitation of nucleosome disruption at ribosomal protein (RP) genes after heat shock or upon deletion of CHD1 (chd1Δ). The Y axis shows the correlation coefficient as measured by shapeDiff for genes in each class as indicated on the X-axis.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig1: Nucleosome occupancy inchd1Δ.(A) Average nucleosome profile for all genes (n =5,207) shows that nucleosome occupancy is reduced at gene bodies in chd1Δ. The X-axis shows genomic position relative to the transcription start site (TSS). The Y-axis represents average read counts per million reads (M). (B) Example of smoothing MNase-seq read data using a spline function. Smoothing removes the noise caused by fluctuations in read coverage that can give erroneous correlation coefficients. Blue line is the raw, binned read count data and the red line is smoothed (C) Smoothing followed by Pearson correlation, called shapeDiff analysis, measures the nucleosome occupancy similarity as 0.31 between wild-type (WT) and chd1Δ over the transcribed region of RPL37B. MET14 has a high correlation coefficient between WT and chd1Δ as determined by shapeDiff due to high similarity of nucleosome occupancy profiles. (D) Average nucleosome profile for RP genes (n =136) shows strong nucleosome disruption upon deletion of CHD1. The shaded bands represent the 95% confidence interval of the data. (E) Quantitation of nucleosome disruption at ribosomal protein (RP) genes after heat shock or upon deletion of CHD1 (chd1Δ). The Y axis shows the correlation coefficient as measured by shapeDiff for genes in each class as indicated on the X-axis.
Mentions: We used MNase-seq to map nucleosome positions in wild-type (WT) and chd1Δ strains of budding yeast[4]. We found that the loss of Chd1 disrupted nucleosome organization within gene bodies, consistent with previous observations in both budding and fission yeast[10, 12, 13] (Figure 1A). We further confirmed this phenotype in the chd1Δ strain with a different resistance marker (Methods, [see Additional file1: Figure S1A]). Although these genome-wide profiles indicate that nucleosome occupancy is generally affected by Chd1, they do not reveal which subsets of genes are specifically dependent on Chd1 function, and what the molecular basis of this dependency might be. To gain insights into these questions, we developed an approach based on quantitatively scoring all genes by the extent of nucleosome disruption.In this approach, which we called ‘shapeDiff analysis,’ we first smoothed nucleosome occupancy signals based on read counts using a spline function, then measured the correlation coefficient between the WT and mutant nucleosome profiles for every gene in the genome (Methods). This approach has the advantage that the correlation measurement is relatively insensitive to noisy fluctuations in the nucleosome occupancy signal caused by low read counts (Figure 1B).Figure 1

Bottom Line: Using genome-wide approaches, we found that the loss of Chd1 significantly disrupted nucleosome arrays within the gene bodies of highly transcribed genes.We also found that Chd1 is physically recruited to gene bodies, and that its occupancy specifically corresponds to that of the early elongating form of RNA polymerase, RNAPII Ser 5-P.We also found that deletion of the histone methyltransferase for H3K36 (SET2) did not affect either Chd1 occupancy or nucleosome organization genome-wide.

View Article: PubMed Central - PubMed

Affiliation: Center for Systems and Synthetic Biology, Institute for Cellular and Molecular Biology, Department of Molecular Biosciences, University of Texas, 2500 Speedway, Austin, TX 78712 USA.

ABSTRACT

Background: Chromatin consists of ordered nucleosomal arrays that are controlled by highly conserved adenosine triphosphate (ATP)-dependent chromatin remodeling complexes. One such remodeler, chromodomain helicase DNA binding protein 1 (Chd1), is believed to play an integral role in nucleosomal organization, as the loss of Chd1 is known to disrupt chromatin. However, the specificity and basis for the functional and physical localization of Chd1 on chromatin remains largely unknown.

Results: Using genome-wide approaches, we found that the loss of Chd1 significantly disrupted nucleosome arrays within the gene bodies of highly transcribed genes. We also found that Chd1 is physically recruited to gene bodies, and that its occupancy specifically corresponds to that of the early elongating form of RNA polymerase, RNAPII Ser 5-P. Conversely, RNAPII Ser 5-P occupancy was affected by the loss of Chd1, suggesting that Chd1 is associated with early transcription elongation. Surprisingly, the occupancy of RNAPII Ser 5-P was affected by the loss of Chd1 specifically at intron-containing genes. Nucleosome turnover was also affected at these sites in the absence of Chd1. We also found that deletion of the histone methyltransferase for H3K36 (SET2) did not affect either Chd1 occupancy or nucleosome organization genome-wide.

Conclusions: Chd1 is specifically recruited onto the gene bodies of highly transcribed genes in an elongation-dependent but H3K36me3-independent manner. Chd1 co-localizes with the early elongating form of RNA polymerase, and affects the occupancy of RNAPII only at genes containing introns, suggesting a role in relieving splicing-related pausing of RNAPII.

No MeSH data available.


Related in: MedlinePlus