Modulation of nucleosomal DNA accessibility via charge-altering post-translational modifications in histone core

Abstract

Background: Controlled modulation of nucleosomal DNA accessibility via post-translational modifications (PTM) is a critical component to many cellular functions. Charge-altering PTMs in the globular histone core-including acetylation, phosphorylation, crotonylation, propionylation, butyrylation, formylation, and citrullination-can alter the strong electrostatic interactions between the oppositely charged nucleosomal DNA and the histone proteins and thus modulate accessibility of the nucleosomal DNA, affecting processes that depend on access to the genetic information, such as transcription. However, direct experimental investigation of the effects of these PTMs is very difficult. Theoretical models can rationalize existing observations, suggest working hypotheses for future experiments, and provide a unifying framework for connecting PTMs with the observed effects.

Results: A physics-based framework is proposed that predicts the effect of charge-altering PTMs in the histone core, quantitatively for several types of lysine charge-neutralizing PTMs including acetylation, and qualitatively for all phosphorylations, on the nucleosome stability and subsequent changes in DNA accessibility, making a connection to resulting biological phenotypes. The framework takes into account multiple partially assembled states of the nucleosome at the atomic resolution. The framework is validated against experimentally known nucleosome stability changes due to the acetylation of specific lysines, and their effect on transcription. The predicted effect of charge-altering PTMs on DNA accessibility can vary dramatically, from virtually none to a strong, region-dependent increase in accessibility of the nucleosomal DNA; in some cases, e.g., H4K44, H2AK75, and H2BK57, the effect is significantly stronger than that of the extensively studied acetylation sites such H3K56, H3K115 or H3K122. Proximity to the DNA is suggestive of the strength of the PTM effect, but there are many exceptions. For the vast majority of charge-altering PTMs, the predicted increase in the DNA accessibility should be large enough to result in a measurable modulation of transcription. However, a few possible PTMs, such as acetylation of H4K77, counterintuitively decrease the DNA accessibility, suggestive of the repressed chromatin. A structural explanation for the phenomenon is provided. For the majority of charge-altering PTMs, the effect on DNA accessibility is simply additive (noncooperative), but there are exceptions, e.g., simultaneous acetylation of H4K79 and H3K122, where the combined effect is amplified. The amplification is a direct consequence of the nucleosome-DNA complex having more than two structural states. The effect of individual PTMs is classified based on changes in the accessibility of various regions throughout the nucleosomal DNA. The PTM's resulting imprint on the DNA accessibility, "PTMprint," is used to predict effects of many yet unexplored PTMs. For example, acetylation of H4K44 yields a PTMprint similar to the PTMprint of H3K56, and thus acetylation of H4K44 is predicted to lead to a wide range of strong biological effects.

Conclusion: Charge-altering post-translational modifications in the relatively unexplored globular histone core may provide a precision mechanism for controlling accessibility to the nucleosomal DNA.

Fig. 1.

The six conformational states used in the thermodynamic model of DNA accessibility in the nucleosome. The states differ by the degree of DNA accessibility and histone core composition. The DNA is completely inaccessible in the canonical nucleosome (W, wrapped). In each of the three partially wrapped conformations (P1, P3, P3) 20 bp are accessible. In the tetrasome (T), a total of 69 bp are accessible [62]. Each conformational state can also carry a post-translational modification (PTM) resulting in a total of twelve distinct energy states considered

Fig. 2

Nucleosomal DNA accessibility PTMprints for acetylated lysine residues within the globular histone core. a Schematic of the different DNA regions. b Accessibility of the different nucleosomal DNA regions without a PTM. The very low probability of the global region accessibility corresponds to the simultaneously unwrapping of 78 bp of the DNA. c and d show change in accessibility, (P*/P), upon lysine acetylation, by region. c Entry/Exit, and d global region. The horizontal dashed lines indicate the conservative threshold value used to define a functionally significant change in accessibility. The threshold value corresponds to the accessibility change of H4K31 for the entry/exit region, and to H4K79 for global accessibility. Note that the entry/exit nucleosomal DNA is accessible in multiple states (U, T, P1, P2, P3); the global region is only accessible in the T and U states, Fig. 1

Fig. 3

The spatial distribution of residues susceptible to acetylation throughout the globular core for each histone protein. Each PTM for a given histone is color coded according to its PTMprint classification: its predicted change on accessibility of different regions of the DNA

Fig. 4

Additivity of the effect of multiple PTMs. Shown are the relative changes in free energy for different combinations of PTMs (∆∆G¹⁺²) compared to the sum of individual PTMs (∆∆G¹ + ∆∆G²). In rare cases (two outlier blue crosses), the change in ∆∆G¹⁺² due to a pair of PTMs can be significantly different from the sum of the changes in binding affinity for each of the PTMs separately (∆∆G¹ + ∆∆G²). The green dot identifies the PTM pair for which experimental data are available, [43], and the red star shows the corresponding predicted value

Fig. 5

Histone sequences with all lysine residues annotated with experimentally known PTMs and color coded by our predicted effect on the DNA accessibility as in Figs. 4 and 5. Regions within the bold square brackets [] are considered part of the globular core, which is the focus of this work. Underlined residues denote mutations relative to the human variant of the histone. The ‘+’ in H2A refers to a missing residue relative to the human variant of H2A. Note that in the nucleosome crystal structure used here (PDB ID 1KX5) H4 is the only human histone variant; H2A and H2B are from xenopus and H3 is a bovine variant

Fig. 6

Explanation for the counterintuitive net stabilizing effect of H4K77^Ac on the nucleosome, which makes the global DNA less accessible. In the intact nucleosome, the positively charged H4K77 (blue spheres) is in close proximity of H2BR92 (green spheres): their repulsion destabilizes the nucleosome, favoring more open states with higher DNA accessibility. This destabilizing interaction is eliminated when an acetylation of H4K77 neutralizes its positive charge

Fig. 7

Thermodynamic cycles of the states used in the model