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MNase Digestion Protection Patterns of the Linker DNA in Chromatosomes.

ABSTRACT: The compact nucleosomal structure limits DNA accessibility and regulates DNA-dependent cellular activities. Linker histones bind to nucleosomes and compact nucleosomal arrays into a higher-order chromatin structure. Recent developments in high throughput technologies and structural computational studies provide nucleosome positioning at a high resolution and contribute to the information of linker histone location within a chromatosome. However, the precise linker histone location within the chromatin fibre remains unclear. Using monomer extension, we mapped core particle and chromatosomal positions over a core histone-reconstituted, 1.5 kb stretch of DNA from the chicken adult β-globin gene, after titration with linker histones and linker histone globular domains. Our results show that, although linker histone globular domains and linker histones display a wide variation in their binding affinity for different positioned nucleosomes, they do not alter nucleosome positions or generate new nucleosome positions. Furthermore, the extra ~20 bp of DNA protected in a chromatosome is usually symmetrically distributed at each end of the core particle, suggesting linker histones or linker histone globular domains are located close to the nucleosomal dyad axis.

SUBMITTER: Shen CH

PROVIDER: S-EPMC8469290 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The chromatin in eukaryotic cells plays a fundamental role in all processes during a cell's life cycle. This nucleoprotein is normally tightly packed but needs to be unpacked for expression and division. The linker histones are critical for such packaging processes and while most experimental and simulation works recognize their crucial importance, the focus is nearly always set on the nucleosome as the basic chromatin building block. Linker histones can undergo several modifications, but only few studies on their ubiquitylation have been conducted. Mono-ubiquitylated linker histones (HUb), while poorly understood, are expected to influence DNA compaction. The size of ubiquitin and the globular domain of the linker histone are comparable and one would expect an increased disorder upon ubiquitylation of the linker histone. However, the formation of higher order chromatin is not hindered and ubiquitylation of the linker histone may even promote gene expression. Structural data on chromatosomes is rare and HUb has never been modeled in a chromatosome so far. Descriptions of the chromatin complex with HUb would greatly benefit from computational structural data. In this study we generate molecular dynamics simulation data for six differently linked HUb variants with the help of a sampling scheme tailored to drive the exploration of phase space. We identify conformational sub-states of the six HUb variants using the sketch-map algorithm for dimensionality reduction and iterative HDBSCAN for clustering on the excessively sampled, shallow free energy landscapes. We present a highly efficient geometric scoring method to identify sub-states of HUb that fit into the nucleosome. We predict HUb conformations inside a nucleosome using on-dyad and off-dyad chromatosome structures as reference and show that unbiased simulations of HUb produce significantly more fitting than non-fitting HUb conformations. A tetranucleosome array is used to show that ubiquitylation can even occur in chromatin without too much steric clashes.

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MNase Digestion Protection Patterns of the Linker DNA in Chromatosomes.

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