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Chromatin, a dynamic protein-DNA complex that regulates eukaryotic genome accessibility and essential functions, is composed of nucleosomes connected by linker DNA with each nucleosome consisting of DNA wrapped around an octamer of histones H2A, H2B, H3 and H4. Magic angle spinning solid-state nuclear magnetic resonance (NMR) spectroscopy can yield unique insights into histone structure and dynamics in condensed nucleosomes and nucleosome arrays representative of chromatin at physiological concentrations. Recently we used J-coupling-based solid-state NMR methods to investigate with residue-specific resolution the conformational dynamics of histone H3 N-terminal tails in 16-mer nucleosome arrays containing 15, 30 or 60 bp DNA linkers. Here, we probe the H3 core domain in the 16-mer arrays as a function of DNA linker lengthviadipolar coupling-based1H-detected solid-state NMR techniques. Specifically, we established nearly complete assignments of backbone chemical shifts for H3 core residues in arrays with 15–60 bp DNA linkers reconstituted with2H,13C,15N-labeled H3. Overall, these chemical shifts were similar irrespective of the DNA linker length indicating no major changes in H3 core conformation. Notably, however, multiple residues at the H3-nucleosomal DNA interface in arrays with 15 bp DNA linkers exhibited relatively pronounced differences in chemical shifts and line broadening compared to arrays with 30 and 60 bp linkers. These findings are consistent with increased heterogeneity in nucleosome packing and structural strain within arrays containing short DNA linkers that likely leads to side-chains of these interfacial residues experiencing alternate conformations or shifts in their rotamer populations relative to arrays with the longer DNA linkers.
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Compromised two-start zigzag chromatin folding in immature mouse retina cells driven by irregularly spaced nucleosomes with short DNA linkers
Abstract The formation of condensed heterochromatin is critical for establishing cell-specific transcriptional programs. To reveal structural transitions underlying heterochromatin formation in maturing mouse rod photoreceptors, we apply cryo-electron microscopy (cryo-EM) tomography, AI-assisted denoising, and molecular modeling. We find that chromatin isolated from immature retina cells contains many closely apposed nucleosomes with extremely short or absent nucleosome linkers, which are inconsistent with the typical two-start zigzag chromatin folding. In mature retina cells, the fraction of short-linker nucleosomes is much lower, supporting stronger chromatin compaction. By cryo-EM-assisted nucleosome interaction capture, we observe that chromatin in immature retina is enriched with i ± 1 interactions, while chromatin in mature retina contains predominantly i ± 2 interactions typical of the two-start zigzag. By mesoscale modeling and computational simulation, we clarify that the unusually short linkers typical of immature retina are sufficient to inhibit the two-start zigzag and chromatin compaction by the interference of very short linkers with linker DNA stems. We propose that this short linker composition renders nucleosome arrays more open in immature retina and that, as the linker DNA length increases in mature retina, chromatin becomes globally condensed via tight zigzag folding. This mechanism may be broadly utilized to introduce higher chromatin folding entropy for epigenomic plasticity.
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- Award ID(s):
- 1911940
- PAR ID:
- 10597922
- Publisher / Repository:
- Oxford University Press
- Date Published:
- Journal Name:
- Nucleic Acids Research
- Volume:
- 53
- Issue:
- 10
- ISSN:
- 0305-1048
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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