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PreprintCited by 3Genomics and Chromatin DynamicsProtein Structure and DynamicsNuclear Structure and Function

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Mechanistic basis for the opposing effects of H2A and H2B ubiquitination on nucleosome stability and dynamics

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Baweja, Lokesh, Wereszczynski, Jeff·2025·Epigenetics & Chromatin
DOI 10.1101/2025.02.13.638112PMC PMC12628893

Preprint — published as 10.1186/s13072-025-00635-0 · Paper page

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Abstract

Nucleosome ubiquitination at lysine 119 of histone H2A (H2AK119ub) and lysine 120 of histone H2B (H2BK120ub) are prominent post-translational modifications with opposing roles in chromatin regulation. Although H2AK119ub is associated with transcriptional repression and H2BK120ub with activation, the molecular basis for these contrasting effects has remained unclear. Here, we use microsecond all-atom and millisecond coarse-grained molecular dynamics simulations to reveal how the position of ubiquitin reshapes nucleosome structure and assembly. H2AK119ub rigidifies the histone core by indirectly reinforcing the L1–L1 interface between H2A histones, strengthening both tetramer–dimer and dimer–dimer interactions, and slowing complete nucleosome assembly. In contrast, H2BK120ub disrupts these interfaces, weakens the histone core, and favors partially assembled hexasome and tetrasome states. Both modifications cause dramatic slowdowns in nucleosome folding, with H2BK120ub producing an order-of-magnitude greater effect. These simulations establish clear molecular mechanisms by which site-specific ubiquitination alters nucleosome stability and assembly kinetics. Our findings quantitatively explain how H2A and H2B ubiquitination exert opposing effects on chromatin regulation. This mechanism is directly relevant to the opposing roles of these marks in transcriptional activation and repression, and may represent one way that combinations of histone modifications modulate chromatin function in vivo .

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Fig. 1 A, B Initial conformations of the H2AK119ub and H2BK120ub nucleosome systems generated with Chimera and the Widom 601 nucleotide sequence in 5’ to 3’ orientation. The DNA exit end is AT-rich. C Super helical locations (SHLs) are sites marked on the nucleosomal DNA
Fig. 1 A, B Initial conformations of the H2AK119ub and H2BK120ub nucleosome systems generated with Chimera and the Widom 601 nucleotide sequence in 5’ to 3’ orientation. The DNA exit end is AT-rich. C Super helical locations (SHLs) are sites marked on the nucleosomal DNACC BY-NC-ND · PubMed Central
Fig. 2 A–D Root mean squared deviations (RMSD) of ubiquitin in H2A119ub and H2BK120ub nucleosome systems in all-atom MD simulations, which quantify its motion relative to the histone core. Ubiquitin has more restricted dynamics in the H2BK120ub system. E Representative configurations from the five most populated structural clusters, based on RMSD in the equilibrated portions of the trajectories
Fig. 2 A–D Root mean squared deviations (RMSD) of ubiquitin in H2A119ub and H2BK120ub nucleosome systems in all-atom MD simulations, which quantify its motion relative to the histone core. Ubiquitin has more restricted dynamics in the H2BK120ub system. E Representative configurations from the five most populated structural clusters, based on RMSD in the equilibrated portions of the trajectoriesCC BY-NC-ND · PubMed Central
Fig. 3 A Schematic of the dyad axis and the H2A axis, illustrating the angle measured to quantify dimer motion within the nucleosome core. B, C Distribution and mean values of the H2A-dyad angle in all-atom MD simulations. Both H2AK119ub (grey) and H2BK120ub (blue) systems exhibit increased outward tilting of the H2A-H2B dimers relative to the canonical nucleosome (red)
Fig. 3 A Schematic of the dyad axis and the H2A axis, illustrating the angle measured to quantify dimer motion within the nucleosome core. B, C Distribution and mean values of the H2A-dyad angle in all-atom MD simulations. Both H2AK119ub (grey) and H2BK120ub (blue) systems exhibit increased outward tilting of the H2A-H2B dimers relative to the canonical nucleosome (red)CC BY-NC-ND · PubMed Central
Fig. 4 A Structural view of the H2A-H2B dimer-dimer interface in the nucleosome, highlighting the roles of the H2A L1-L1 loops and H2B \upalpha 2 helices. B–D Per-residue contact maps showing inter-dimer interactions in the canonical, H2AK119ub, and H2BK120ub in all-atom MD simulations. The H2A L1-L1 region in H2AK119ub shows a pronounced increase in contacts. The color bar indicates the total number of contacts per residue pair
Fig. 4 A Structural view of the H2A-H2B dimer-dimer interface in the nucleosome, highlighting the roles of the H2A L1-L1 loops and H2B \upalpha 2 helices. B–D Per-residue contact maps showing inter-dimer interactions in the canonical, H2AK119ub, and H2BK120ub in all-atom MD simulations. The H2A L1-L1 region in H2AK119ub shows a pronounced increase in contacts. The color bar indicates the total number of contacts per residue pairCC BY-NC-ND · PubMed Central
Fig. 5 DNA unwrapping (“breathing”) at the nucleosome entry and exit sites, quantified by the number of base pairs unwrapped from the superhelical axis over time in all-atom MD simulations. Both the canonical and H2BK120ub nucleosomes show greater DNA unwrapping than the H2AK119ub system
Fig. 5 DNA unwrapping (“breathing”) at the nucleosome entry and exit sites, quantified by the number of base pairs unwrapped from the superhelical axis over time in all-atom MD simulations. Both the canonical and H2BK120ub nucleosomes show greater DNA unwrapping than the H2AK119ub systemCC BY-NC-ND · PubMed Central
Fig. 6 A, B Differences in atomic mutual information (dynamical correlations) between H2AK119ub and H2BK120ub nucleosomes, relative to the canonical system, in all-atom MD simulations. Positive values reflect increased coupling; negative values reflect reduced coupling. Diagonal blocks represent intra-molecular correlations; off-diagonal blocks indicate inter-molecular correlations between histones and DNA. These maps highlight how H2AK119ub enhances coupling at the DNA entry and exit regions, whereas H2BK120ub reduces histone-DNA correlations within the nucleosome core
Fig. 6 A, B Differences in atomic mutual information (dynamical correlations) between H2AK119ub and H2BK120ub nucleosomes, relative to the canonical system, in all-atom MD simulations. Positive values reflect increased coupling; negative values reflect reduced coupling. Diagonal blocks represent intra-molecular correlations; off-diagonal blocks indicate inter-molecular correlations between histones and DNA. These maps highlight how H2AK119ub enhances coupling at the DNA entry and exit regions, whereas H2BK120ub reduces histone-DNA correlations within the nucleosome coreCC BY-NC-ND · PubMed Central
Fig. 7 Top six assembly pathways for the canonical nucleosome, as identified by transition path theory from the coarse-grain Markov State Model. Each node represents a macrostate, with size proportional to equilibrium free energy. Arrow thickness indicates pathway dominance. The pathways highlight transitions between tetrasomal (T, T1, T2), hexasomal (HR, HR1, HR2, HR3, HL), and nucleosomal (N) states
Fig. 7 Top six assembly pathways for the canonical nucleosome, as identified by transition path theory from the coarse-grain Markov State Model. Each node represents a macrostate, with size proportional to equilibrium free energy. Arrow thickness indicates pathway dominance. The pathways highlight transitions between tetrasomal (T, T1, T2), hexasomal (HR, HR1, HR2, HR3, HL), and nucleosomal (N) statesCC BY-NC-ND · PubMed Central
Fig. 8 Top six assembly pathways for the H2AK119ub-modified nucleosome, as identified by transition path theory from Markov State Model analysis of the coarse-grain MD simulations. Each node represents a macrostate, with its size proportional to the equilibrium free energy. The arrow thickness indicates the pathway’s dominance. Compared to the canonical system, the nucleosomal state (N) is destabilized, while extended (T, T1) and compact (T2, Part_T) tetrasomal states remain low in free energy. The HL1 hexasomal state also appears as a relatively stable intermediate, contributing to the dominant folding routes
Fig. 8 Top six assembly pathways for the H2AK119ub-modified nucleosome, as identified by transition path theory from Markov State Model analysis of the coarse-grain MD simulations. Each node represents a macrostate, with its size proportional to the equilibrium free energy. The arrow thickness indicates the pathway’s dominance. Compared to the canonical system, the nucleosomal state (N) is destabilized, while extended (T, T1) and compact (T2, Part_T) tetrasomal states remain low in free energy. The HL1 hexasomal state also appears as a relatively stable intermediate, contributing to the dominant folding routesCC BY-NC-ND · PubMed Central
Fig. 9 Top six assembly pathways for the H2BK120ub-modified nucleosome, as identified by transition path theory from Markov State Model analysis of the coarse-grain MD simulations. Each node represents a macrostate, with its size proportional to the equilibrium free energy. The arrow thickness indicates the pathway’s dominance. Compared to both the canonical and H2AK119ub systems, the nucleosomal state (N) is sparsely populated and high in free energy, while multiple tetrasomal states are strongly stabilized. Folding pathways are highly diffuse, with many weakly populated routes, consistent with slower and less efficient nucleosome assembly
Fig. 9 Top six assembly pathways for the H2BK120ub-modified nucleosome, as identified by transition path theory from Markov State Model analysis of the coarse-grain MD simulations. Each node represents a macrostate, with its size proportional to the equilibrium free energy. The arrow thickness indicates the pathway’s dominance. Compared to both the canonical and H2AK119ub systems, the nucleosomal state (N) is sparsely populated and high in free energy, while multiple tetrasomal states are strongly stabilized. Folding pathways are highly diffuse, with many weakly populated routes, consistent with slower and less efficient nucleosome assemblyCC BY-NC-ND · PubMed Central
Fig. 10 Illustration depicting the distinct mechanisms by which H2AK119ub and H2BK120ub alter nucleosome stability and assembly. H2AK119ub indirectly strengthens the L1-L1 interface between H2A histones, stabilizing the nucleosome core and slowing assembly. In contrast, H2BK120ub disrupts these contacts and weakens the histone core, promoting partially assembled hexasome and tetrasome states. Arrows and annotations indicate the sites of ubiquitination, key affected interfaces, and the resulting structural and kinetic effects on nucleosome folding
Fig. 10 Illustration depicting the distinct mechanisms by which H2AK119ub and H2BK120ub alter nucleosome stability and assembly. H2AK119ub indirectly strengthens the L1-L1 interface between H2A histones, stabilizing the nucleosome core and slowing assembly. In contrast, H2BK120ub disrupts these contacts and weakens the histone core, promoting partially assembled hexasome and tetrasome states. Arrows and annotations indicate the sites of ubiquitination, key affected interfaces, and the resulting structural and kinetic effects on nucleosome foldingCC BY-NC-ND · PubMed Central

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Metadata

Authors
Baweja, L., & Wereszczynski, J.
Affiliations
Illinois Institute of Technology (US)
Corresponding author
Wereszczynski, Jeff
ORCID
Venue
Epigenetics & Chromatin
Publisher
openRxiv
Published
2025-02-14
Volume
18
Issue
1
Pages
74-74
ISSN
1756-8935 (ISSN-L)
DOI
10.1101/2025.02.13.638112
PMC
PMC12628893
OpenAlex
W4416392468
Cited by
3
References
1
Citations by year
2026: 2 · 2025: 1
FWCI
0.90
Type
journal article
Language
en
Open access
Licence
Locations
  • Epigenetics & Chromatin (publishedVersion, cc-by-nc-nd)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion, cc-by-nc)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
National Institute of General Medical Sciences (R35GM119647)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Nuclear Structure and Function
Keywords
histone H2A ubiquitination, histone H2B ubiquitination, H2AK119ub, nucleosome stability, hexasome, chromatin regulation, transcriptional repression, transcriptional activation, histone modifications

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Baweja, L., & Wereszczynski, J. (2025). Mechanistic basis for the opposing effects of H2A and H2B ubiquitination on nucleosome stability and dynamics. Epigenetics & Chromatin. https://doi.org/10.1101/2025.02.13.638112