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Chromatin remodelers clear nucleosomes from intrinsically unfavorable sites to establish nucleosome-depleted regions at promoters.

Tolkunov D, Zawadzki KA, Singer C, Elfving N, Morozov AV, Broach JR - Mol. Biol. Cell (2011)

Bottom Line: In snf2 mutants, excess promoter nucleosomes correlate with reduced gene expression.Cells lacking SNF2 or ASF1 still accomplish the changes in promoter nucleosome structure associated with large-scale transcriptional reprogramming.However, chromatin reorganization in the mutants is reduced in extent compared to wild-type cells, even though transcriptional changes proceed normally.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy and BioMaPS Institute for Quantitative Biology, Rutgers University, Piscataway, NJ 08854, USA.

ABSTRACT
Most promoters in yeast contain a nucleosome-depleted region (NDR), but the mechanisms by which NDRs are established and maintained in vivo are currently unclear. We have examined how genome-wide nucleosome placement is altered in the absence of two distinct types of nucleosome remodeling activity. In mutants of both SNF2, which encodes the ATPase component of the Swi/Snf remodeling complex, and ASF1, which encodes a histone chaperone, distinct sets of gene promoters carry excess nucleosomes in their NDRs relative to wild-type. In snf2 mutants, excess promoter nucleosomes correlate with reduced gene expression. In both mutants, the excess nucleosomes occupy DNA sequences that are energetically less favorable for nucleosome formation, indicating that intrinsic histone-DNA interactions are not sufficient for nucleosome positioning in vivo, and that Snf2 and Asf1 promote thermodynamic equilibration of nucleosomal arrays. Cells lacking SNF2 or ASF1 still accomplish the changes in promoter nucleosome structure associated with large-scale transcriptional reprogramming. However, chromatin reorganization in the mutants is reduced in extent compared to wild-type cells, even though transcriptional changes proceed normally. In summary, active remodeling is required for distributing nucleosomes to energetically favorable positions in vivo and for reorganizing chromatin in response to changes in transcriptional activity.

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Energetics of excess promoter nucleosomes. The difference in nucleosome occupancy between mutant and wild-type is computed for all gene promoters as in Figure 2, and the genes are sorted by the average free energy of nucleosome formation in promoters. (A) Steady-state snf2 mutant and wild-type in glucose. (B) Steady-state asf1 mutant and wild-type in glycerol. (C) snf2 mutant and wild-type 20 min after glucose-to-glycerol downshift. (D) asf1 mutant and wild-type 20 min after glycerol-to-glucose upshift. The average free energy is computed as the mean of the nucleosome formation energies in the [−400 base pairs, −100 base pairs] window upstream of the TSS (each position in the window is taken as a starting base pair of a 147–base pair nucleosome core particle). The free energy at each position is given by the model defined in Eq. (3) (see Materials and Methods). Excess promoter nucleosomes in the deletion strains tend to reside on energetically less (A, B, and D) or more (C) favorable sequences. On the right, the red lines show the mean nucleosome energy over the promoter (more positive energy values indicate that nucleosomes reside on relatively less favorable DNA sequences). The black lines are the net differences in nucleosome occupancy between the mutant and the wild-type across each promoter, smoothed with a 100-gene moving average; larger values indicate more net excess nucleosomes in mutants. In each panel, the Pearson correlation coefficient r is shown for the smoothed occupancy differences. The p values (computed using a two-tailed Student's t test) for the unsmoothed occupancy differences are < 10−10 in all cases; the Pearson correlation coefficients are 0.28, 0.15, −0.27, and 0.25 for A, B, C, and D, respectively.
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Figure 3: Energetics of excess promoter nucleosomes. The difference in nucleosome occupancy between mutant and wild-type is computed for all gene promoters as in Figure 2, and the genes are sorted by the average free energy of nucleosome formation in promoters. (A) Steady-state snf2 mutant and wild-type in glucose. (B) Steady-state asf1 mutant and wild-type in glycerol. (C) snf2 mutant and wild-type 20 min after glucose-to-glycerol downshift. (D) asf1 mutant and wild-type 20 min after glycerol-to-glucose upshift. The average free energy is computed as the mean of the nucleosome formation energies in the [−400 base pairs, −100 base pairs] window upstream of the TSS (each position in the window is taken as a starting base pair of a 147–base pair nucleosome core particle). The free energy at each position is given by the model defined in Eq. (3) (see Materials and Methods). Excess promoter nucleosomes in the deletion strains tend to reside on energetically less (A, B, and D) or more (C) favorable sequences. On the right, the red lines show the mean nucleosome energy over the promoter (more positive energy values indicate that nucleosomes reside on relatively less favorable DNA sequences). The black lines are the net differences in nucleosome occupancy between the mutant and the wild-type across each promoter, smoothed with a 100-gene moving average; larger values indicate more net excess nucleosomes in mutants. In each panel, the Pearson correlation coefficient r is shown for the smoothed occupancy differences. The p values (computed using a two-tailed Student's t test) for the unsmoothed occupancy differences are < 10−10 in all cases; the Pearson correlation coefficients are 0.28, 0.15, −0.27, and 0.25 for A, B, C, and D, respectively.

Mentions: Mutation of either SNF2 or ASF1 leads to excess promoter nucleosomes at distinct locations and consequently at distinct sets of sequences. As Snf2 is an ATPase that expends energy when remodeling or removing nucleosomes, but Asf1 is not, we wondered if those sequences that gained nucleosomes in the snf2 mutant were intrinsically more favorable for nucleosome assembly than those in the asf1 mutant. Accordingly, we compared the energy cost associated with excess nucleosomes present at steady state in both mutants. Our biophysical model of nucleosome energetics accounts for intrinsic histone–DNA interactions and for steric exclusion between neighboring nucleosomes, and thus captures nucleosome positioning preferences in the absence of external factors (Locke et al., 2010). We calculated the average sequence-dependent free energy of nucleosome formation in vitro for each promoter in the yeast genome, and compared that to the extent of excess nucleosome occupancy in that promoter (Figures 3 and S2B). Surprisingly, excess nucleosomes in snf2 cells tend to occur at promoters with a high energy cost of nucleosome assembly (Figure 3A; p < 10−10). A similar correlation holds for asf1, both in glucose (Figure S2B; p < 10−10) and glycerol (Figure 3B; p < 10−10). Thus Snf2 and Asf1 function to remove nucleosomes from those promoters on which it is most difficult to assemble nucleosomes in vitro. We conclude that under steady-state conditions both activities move nucleosomes to more thermodynamically stable positions on the genome, which suggests that excess nucleosomes are kinetically trapped at their inappropriate sites. It appears that Snf2 and Asf1 mediate transition toward thermodynamic equilibrium, lowering the total free energy of nucleosomal arrays.


Chromatin remodelers clear nucleosomes from intrinsically unfavorable sites to establish nucleosome-depleted regions at promoters.

Tolkunov D, Zawadzki KA, Singer C, Elfving N, Morozov AV, Broach JR - Mol. Biol. Cell (2011)

Energetics of excess promoter nucleosomes. The difference in nucleosome occupancy between mutant and wild-type is computed for all gene promoters as in Figure 2, and the genes are sorted by the average free energy of nucleosome formation in promoters. (A) Steady-state snf2 mutant and wild-type in glucose. (B) Steady-state asf1 mutant and wild-type in glycerol. (C) snf2 mutant and wild-type 20 min after glucose-to-glycerol downshift. (D) asf1 mutant and wild-type 20 min after glycerol-to-glucose upshift. The average free energy is computed as the mean of the nucleosome formation energies in the [−400 base pairs, −100 base pairs] window upstream of the TSS (each position in the window is taken as a starting base pair of a 147–base pair nucleosome core particle). The free energy at each position is given by the model defined in Eq. (3) (see Materials and Methods). Excess promoter nucleosomes in the deletion strains tend to reside on energetically less (A, B, and D) or more (C) favorable sequences. On the right, the red lines show the mean nucleosome energy over the promoter (more positive energy values indicate that nucleosomes reside on relatively less favorable DNA sequences). The black lines are the net differences in nucleosome occupancy between the mutant and the wild-type across each promoter, smoothed with a 100-gene moving average; larger values indicate more net excess nucleosomes in mutants. In each panel, the Pearson correlation coefficient r is shown for the smoothed occupancy differences. The p values (computed using a two-tailed Student's t test) for the unsmoothed occupancy differences are < 10−10 in all cases; the Pearson correlation coefficients are 0.28, 0.15, −0.27, and 0.25 for A, B, C, and D, respectively.
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Related In: Results  -  Collection

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Figure 3: Energetics of excess promoter nucleosomes. The difference in nucleosome occupancy between mutant and wild-type is computed for all gene promoters as in Figure 2, and the genes are sorted by the average free energy of nucleosome formation in promoters. (A) Steady-state snf2 mutant and wild-type in glucose. (B) Steady-state asf1 mutant and wild-type in glycerol. (C) snf2 mutant and wild-type 20 min after glucose-to-glycerol downshift. (D) asf1 mutant and wild-type 20 min after glycerol-to-glucose upshift. The average free energy is computed as the mean of the nucleosome formation energies in the [−400 base pairs, −100 base pairs] window upstream of the TSS (each position in the window is taken as a starting base pair of a 147–base pair nucleosome core particle). The free energy at each position is given by the model defined in Eq. (3) (see Materials and Methods). Excess promoter nucleosomes in the deletion strains tend to reside on energetically less (A, B, and D) or more (C) favorable sequences. On the right, the red lines show the mean nucleosome energy over the promoter (more positive energy values indicate that nucleosomes reside on relatively less favorable DNA sequences). The black lines are the net differences in nucleosome occupancy between the mutant and the wild-type across each promoter, smoothed with a 100-gene moving average; larger values indicate more net excess nucleosomes in mutants. In each panel, the Pearson correlation coefficient r is shown for the smoothed occupancy differences. The p values (computed using a two-tailed Student's t test) for the unsmoothed occupancy differences are < 10−10 in all cases; the Pearson correlation coefficients are 0.28, 0.15, −0.27, and 0.25 for A, B, C, and D, respectively.
Mentions: Mutation of either SNF2 or ASF1 leads to excess promoter nucleosomes at distinct locations and consequently at distinct sets of sequences. As Snf2 is an ATPase that expends energy when remodeling or removing nucleosomes, but Asf1 is not, we wondered if those sequences that gained nucleosomes in the snf2 mutant were intrinsically more favorable for nucleosome assembly than those in the asf1 mutant. Accordingly, we compared the energy cost associated with excess nucleosomes present at steady state in both mutants. Our biophysical model of nucleosome energetics accounts for intrinsic histone–DNA interactions and for steric exclusion between neighboring nucleosomes, and thus captures nucleosome positioning preferences in the absence of external factors (Locke et al., 2010). We calculated the average sequence-dependent free energy of nucleosome formation in vitro for each promoter in the yeast genome, and compared that to the extent of excess nucleosome occupancy in that promoter (Figures 3 and S2B). Surprisingly, excess nucleosomes in snf2 cells tend to occur at promoters with a high energy cost of nucleosome assembly (Figure 3A; p < 10−10). A similar correlation holds for asf1, both in glucose (Figure S2B; p < 10−10) and glycerol (Figure 3B; p < 10−10). Thus Snf2 and Asf1 function to remove nucleosomes from those promoters on which it is most difficult to assemble nucleosomes in vitro. We conclude that under steady-state conditions both activities move nucleosomes to more thermodynamically stable positions on the genome, which suggests that excess nucleosomes are kinetically trapped at their inappropriate sites. It appears that Snf2 and Asf1 mediate transition toward thermodynamic equilibrium, lowering the total free energy of nucleosomal arrays.

Bottom Line: In snf2 mutants, excess promoter nucleosomes correlate with reduced gene expression.Cells lacking SNF2 or ASF1 still accomplish the changes in promoter nucleosome structure associated with large-scale transcriptional reprogramming.However, chromatin reorganization in the mutants is reduced in extent compared to wild-type cells, even though transcriptional changes proceed normally.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy and BioMaPS Institute for Quantitative Biology, Rutgers University, Piscataway, NJ 08854, USA.

ABSTRACT
Most promoters in yeast contain a nucleosome-depleted region (NDR), but the mechanisms by which NDRs are established and maintained in vivo are currently unclear. We have examined how genome-wide nucleosome placement is altered in the absence of two distinct types of nucleosome remodeling activity. In mutants of both SNF2, which encodes the ATPase component of the Swi/Snf remodeling complex, and ASF1, which encodes a histone chaperone, distinct sets of gene promoters carry excess nucleosomes in their NDRs relative to wild-type. In snf2 mutants, excess promoter nucleosomes correlate with reduced gene expression. In both mutants, the excess nucleosomes occupy DNA sequences that are energetically less favorable for nucleosome formation, indicating that intrinsic histone-DNA interactions are not sufficient for nucleosome positioning in vivo, and that Snf2 and Asf1 promote thermodynamic equilibration of nucleosomal arrays. Cells lacking SNF2 or ASF1 still accomplish the changes in promoter nucleosome structure associated with large-scale transcriptional reprogramming. However, chromatin reorganization in the mutants is reduced in extent compared to wild-type cells, even though transcriptional changes proceed normally. In summary, active remodeling is required for distributing nucleosomes to energetically favorable positions in vivo and for reorganizing chromatin in response to changes in transcriptional activity.

Show MeSH
Related in: MedlinePlus