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Using DNA mechanics to predict in vitro nucleosome positions and formation energies.

Morozov AV, Fortney K, Gaykalova DA, Studitsky VM, Widom J, Siggia ED - Nucleic Acids Res. (2009)

Bottom Line: While competition with other DNA-binding factors and action of chromatin remodeling enzymes significantly affect nucleosome formation in vivo, nucleosome positions in vitro are determined by steric exclusion and sequence alone.We have also made a first ab initio prediction of nucleosomal DNA geometries, and checked its accuracy against the nucleosome crystal structure.We have used DNABEND to design both strong and weak histone- binding sequences, and measured the corresponding free energies of nucleosome formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics & Astronomy and BioMaPS Institute for Quantitative Biology, Rutgers University, 136 Frelinghuysen Road, Piscataway, NJ 08854, USA. morozov@physics.rutgers.edu

ABSTRACT
In eukaryotic genomes, nucleosomes function to compact DNA and to regulate access to it both by simple physical occlusion and by providing the substrate for numerous covalent epigenetic tags. While competition with other DNA-binding factors and action of chromatin remodeling enzymes significantly affect nucleosome formation in vivo, nucleosome positions in vitro are determined by steric exclusion and sequence alone. We have developed a biophysical model, DNABEND, for the sequence dependence of DNA bending energies, and validated it against a collection of in vitro free energies of nucleosome formation and a set of in vitro nucleosome positions mapped at high resolution. We have also made a first ab initio prediction of nucleosomal DNA geometries, and checked its accuracy against the nucleosome crystal structure. We have used DNABEND to design both strong and weak histone- binding sequences, and measured the corresponding free energies of nucleosome formation. We find that DNABEND can successfully predict in vitro nucleosome positions and free energies, providing a physical explanation for the intrinsic sequence dependence of histone-DNA interactions.

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DNABEND predictions of in vitro nucleosome positions. Probability of a nucleosome to start at each base pair (green), nucleosome occupancy (blue) and nucleosome formation energy (violet). Vertical lines: experimentally known nucleosome starting positions, with base pair coordinates listed in parentheses below. (a) The 180 bp sequence from the sea urchin 5S rRNA gene (bps 8,26) (36). (b) The 183 bp sequence from the pGUB plasmid (bps 11,31) (37). (c) The 215 bp fragment from the sequence of the chicken β–globinA gene (bp 52) (38). (d,e,f) Synthetic high-affinity sequences (27) 601 (bp 61), 603 (bp 81) and 605 (bp 59). Nucleosomes on sequences 601, 603 and 605 were mapped by hydroxyl radical footprinting (Supplementary Figures 2 and 3). All DNA sequences used in this calculation are available on the Nucleosome Explorer web site: http://nucleosome.rockefeller.edu.
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Figure 5: DNABEND predictions of in vitro nucleosome positions. Probability of a nucleosome to start at each base pair (green), nucleosome occupancy (blue) and nucleosome formation energy (violet). Vertical lines: experimentally known nucleosome starting positions, with base pair coordinates listed in parentheses below. (a) The 180 bp sequence from the sea urchin 5S rRNA gene (bps 8,26) (36). (b) The 183 bp sequence from the pGUB plasmid (bps 11,31) (37). (c) The 215 bp fragment from the sequence of the chicken β–globinA gene (bp 52) (38). (d,e,f) Synthetic high-affinity sequences (27) 601 (bp 61), 603 (bp 81) and 605 (bp 59). Nucleosomes on sequences 601, 603 and 605 were mapped by hydroxyl radical footprinting (Supplementary Figures 2 and 3). All DNA sequences used in this calculation are available on the Nucleosome Explorer web site: http://nucleosome.rockefeller.edu.

Mentions: We find that DNABEND predicts footprinted nucleosome positions reasonably well: the measured position is always within 1–2 bp of a local minimum in our energy profiles, and that energy minimum in 5 out of 6 cases is within 0.5–1.0 kcal/mol of the global energy minimum (Figure 5; note that the total range of sequence-dependent binding energies is ∼5 kcal/mol). We expect nucleosome energies to be approximately equal for positions that are in phase with respect to the helical twist, and indeed consecutive energy minima and maxima in Figure 5 are separated by 10–11 bp. However, in several cases (e.g. for clone 601) where the experimental nucleosome position coincides with a local rather than a global minimum, the relatively small energy difference between these minima is sufficient to misplace the predicted nucleosome by tens of base pairs from its experimentally known location. Errors of similar magnitude are made if DNA geometry is taken from the ideal superhelix [(15); Figure 5], from the 1kx5 crystal structure [(14); Supplementary Figure 6], or if nucleosome positions are predicted using the latest bioinformatics model which was trained on sequences from nucleosomes reconstituted in vitro by salt dialysis on yeast genomic DNA [(9); Supplementary Figure 7]. We compare all four prediction methods in Figure 6, using the number and the height of predicted probability peaks near experimentally known positions as a metric. Our results underscore the exacting level of accuracy (≤0.5 kcal/mol) required to position nucleosomes precisely on genomic or synthetic DNA. It is also possible that suboptimal nucleosome positions are in fact produced in experiments but not detected because such sub-populations would be relatively small.Figure 5.


Using DNA mechanics to predict in vitro nucleosome positions and formation energies.

Morozov AV, Fortney K, Gaykalova DA, Studitsky VM, Widom J, Siggia ED - Nucleic Acids Res. (2009)

DNABEND predictions of in vitro nucleosome positions. Probability of a nucleosome to start at each base pair (green), nucleosome occupancy (blue) and nucleosome formation energy (violet). Vertical lines: experimentally known nucleosome starting positions, with base pair coordinates listed in parentheses below. (a) The 180 bp sequence from the sea urchin 5S rRNA gene (bps 8,26) (36). (b) The 183 bp sequence from the pGUB plasmid (bps 11,31) (37). (c) The 215 bp fragment from the sequence of the chicken β–globinA gene (bp 52) (38). (d,e,f) Synthetic high-affinity sequences (27) 601 (bp 61), 603 (bp 81) and 605 (bp 59). Nucleosomes on sequences 601, 603 and 605 were mapped by hydroxyl radical footprinting (Supplementary Figures 2 and 3). All DNA sequences used in this calculation are available on the Nucleosome Explorer web site: http://nucleosome.rockefeller.edu.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2724288&req=5

Figure 5: DNABEND predictions of in vitro nucleosome positions. Probability of a nucleosome to start at each base pair (green), nucleosome occupancy (blue) and nucleosome formation energy (violet). Vertical lines: experimentally known nucleosome starting positions, with base pair coordinates listed in parentheses below. (a) The 180 bp sequence from the sea urchin 5S rRNA gene (bps 8,26) (36). (b) The 183 bp sequence from the pGUB plasmid (bps 11,31) (37). (c) The 215 bp fragment from the sequence of the chicken β–globinA gene (bp 52) (38). (d,e,f) Synthetic high-affinity sequences (27) 601 (bp 61), 603 (bp 81) and 605 (bp 59). Nucleosomes on sequences 601, 603 and 605 were mapped by hydroxyl radical footprinting (Supplementary Figures 2 and 3). All DNA sequences used in this calculation are available on the Nucleosome Explorer web site: http://nucleosome.rockefeller.edu.
Mentions: We find that DNABEND predicts footprinted nucleosome positions reasonably well: the measured position is always within 1–2 bp of a local minimum in our energy profiles, and that energy minimum in 5 out of 6 cases is within 0.5–1.0 kcal/mol of the global energy minimum (Figure 5; note that the total range of sequence-dependent binding energies is ∼5 kcal/mol). We expect nucleosome energies to be approximately equal for positions that are in phase with respect to the helical twist, and indeed consecutive energy minima and maxima in Figure 5 are separated by 10–11 bp. However, in several cases (e.g. for clone 601) where the experimental nucleosome position coincides with a local rather than a global minimum, the relatively small energy difference between these minima is sufficient to misplace the predicted nucleosome by tens of base pairs from its experimentally known location. Errors of similar magnitude are made if DNA geometry is taken from the ideal superhelix [(15); Figure 5], from the 1kx5 crystal structure [(14); Supplementary Figure 6], or if nucleosome positions are predicted using the latest bioinformatics model which was trained on sequences from nucleosomes reconstituted in vitro by salt dialysis on yeast genomic DNA [(9); Supplementary Figure 7]. We compare all four prediction methods in Figure 6, using the number and the height of predicted probability peaks near experimentally known positions as a metric. Our results underscore the exacting level of accuracy (≤0.5 kcal/mol) required to position nucleosomes precisely on genomic or synthetic DNA. It is also possible that suboptimal nucleosome positions are in fact produced in experiments but not detected because such sub-populations would be relatively small.Figure 5.

Bottom Line: While competition with other DNA-binding factors and action of chromatin remodeling enzymes significantly affect nucleosome formation in vivo, nucleosome positions in vitro are determined by steric exclusion and sequence alone.We have also made a first ab initio prediction of nucleosomal DNA geometries, and checked its accuracy against the nucleosome crystal structure.We have used DNABEND to design both strong and weak histone- binding sequences, and measured the corresponding free energies of nucleosome formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics & Astronomy and BioMaPS Institute for Quantitative Biology, Rutgers University, 136 Frelinghuysen Road, Piscataway, NJ 08854, USA. morozov@physics.rutgers.edu

ABSTRACT
In eukaryotic genomes, nucleosomes function to compact DNA and to regulate access to it both by simple physical occlusion and by providing the substrate for numerous covalent epigenetic tags. While competition with other DNA-binding factors and action of chromatin remodeling enzymes significantly affect nucleosome formation in vivo, nucleosome positions in vitro are determined by steric exclusion and sequence alone. We have developed a biophysical model, DNABEND, for the sequence dependence of DNA bending energies, and validated it against a collection of in vitro free energies of nucleosome formation and a set of in vitro nucleosome positions mapped at high resolution. We have also made a first ab initio prediction of nucleosomal DNA geometries, and checked its accuracy against the nucleosome crystal structure. We have used DNABEND to design both strong and weak histone- binding sequences, and measured the corresponding free energies of nucleosome formation. We find that DNABEND can successfully predict in vitro nucleosome positions and free energies, providing a physical explanation for the intrinsic sequence dependence of histone-DNA interactions.

Show MeSH
Related in: MedlinePlus