Changing Chromatin Fiber Conformation by Nucleosome Repositioning

Chromatin conformation is dynamic and heterogeneous with respect to nucleosome positions, which can be changed by chromatin remodeling complexes in the cell. These molecular machines hydrolyze ATP to translocate or evict nucleosomes, and establish loci with regularly and more irregularly spaced nucleosomes as well as nucleosome-depleted regions. The impact of nucleosome repositioning on the three-dimensional chromatin structure is only poorly understood. Here, we address this issue by using a coarse-grained computer model of arrays of 101 nucleosomes considering several chromatin fiber models with and without linker histones, respectively. We investigated the folding of the chain in dependence of the position of the central nucleosome by changing the length of the adjacent linker DNA in basepair steps. We found in our simulations that these translocations had a strong effect on the shape and properties of chromatin fibers: i), Fiber curvature and flexibility at the center were largely increased and long-range contacts between distant nucleosomes on the chain were promoted. ii), The highest destabilization of the fiber conformation occurred for a nucleosome shifted by two basepairs from regular spacing, whereas effects of linker DNA changes of ∼10 bp in phase with the helical twist of DNA were minimal. iii), A fiber conformation can stabilize a regular spacing of nucleosomes inasmuch as favorable stacking interactions between nucleosomes are facilitated. This can oppose nucleosome translocations and increase the energetic costs for chromatin remodeling. Our computational modeling framework makes it possible to describe the conformational heterogeneity of chromatin in terms of nucleosome positions, and thus advances theoretical models toward a better understanding of how genome compaction and access are regulated within the cell.     

Chromatin Fiber Invasion and Nucleosome Displacement by the Rap1 Transcription Factor

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Kinase-Mediated Changes in Nucleosome Conformation Trigger Chromatin Decondensation via Poly(ADP-Ribosyl)ation

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核小体及染色质修饰的基因组分布模式和染色质状态

染色质是真核DNA的存在方式,可以通过影响DNA的可及性调节基因转录,其基本单元为核小体,系由约147 bp的DNA缠绕在组蛋白八联体上形成的结构,核小体之间以连接DNA相连．核小体组蛋白上能发生甲基化和乙酰化等化学修饰．核小体位置、DNA的甲基化和组蛋白的修饰等对染色质状态(常染色质或异染色质)及基因组之间的长程相互作用有重要影响．近年,基于高通量测序技术,核小体位置和染色质修饰在多种细胞中的基因组分布已被测定．结果显示,这些标记的分布模式具有位点特异、动态变化、相互偶联和高度复杂的特征．本文详细回顾并评述了核小体位置和染色质修饰的分布模式、对应生物学功能、修饰之间的关联、实验测定技术、染色质状态的计算分析等内容．该工作对于深入认识和理解染色质的表观遗传调节机制有重要意义．     

Conserved Substitution Patterns around Nucleosome Footprints in Eukaryotes and Archaea Derive from Frequent Nucleosome Repositioning through Evolution

Nucleosomes, the basic repeat units of eukaryotic chromatin, have been suggested to influence the evolution of eukaryotic genomes, both by altering the propensity of DNA to mutate and by selection acting to maintain or exclude nucleosomes in particular locations. Contrary to the popular idea that nucleosomes are unique to eukaryotes, histone proteins have also been discovered in some archaeal genomes. Archaeal nucleosomes, however, are quite unlike their eukaryotic counterparts in many respects, including their assembly into tetramers (rather than octamers) from histone proteins that lack N- and C-terminal tails. Here, we show that despite these fundamental differences the association between nucleosome footprints and sequence evolution is strikingly conserved between humans and the model archaeon Haloferax volcanii. In light of this finding we examine whether selection or mutation can explain concordant substitution patterns in the two kingdoms. Unexpectedly, we find that neither the mutation nor the selection model are sufficient to explain the observed association between nucleosomes and sequence divergence. Instead, we demonstrate that nucleosome-associated substitution patterns are more consistent with a third model where sequence divergence results in frequent repositioning of nucleosomes during evolution. Indeed, we show that nucleosome repositioning is both necessary and largely sufficient to explain the association between current nucleosome positions and biased substitution patterns. This finding highlights the importance of considering the direction of causality between genetic and epigenetic change.     

Chromatin Polymorphism and the Nucleosome Superfamily: A Genealogy

Nucleosomes were discovered more than thirty years ago as the basic repeating units of chromatin. Since then, nucleosomes have progressively revealed their taste to come in many appearances, upon either adjunction of other proteins (e.g. a fifth histone or a non-histone protein, HMG-N), histone substitution for isoforms (histone variants), depletion of one or the two H2A-H2B dimers (sub-nucleosomes), intimate two-particle association, or isomeric structural alterations. The resulting entities, some of them are only transient, acquire new properties useful for their specific roles in chromatin function. These structures are presented here in the chronological order of their identification, from the chromatosome to the sub-nucleosomal hexasome and tetrasome, and from the dinucleosomal altosome and nucleodisome to the nucleosome variants and altered forms: the old lexosome and the most recent reversome.     

Salt-Induced Conformation and Interaction Changes of Nucleosome Core Particles

Small angle x-ray scattering was used to follow changes in the conformation and interactions of nucleosome core particles (NCP) as a function of the monovalent salt concentration C_s. The maximal extension (D_max) of the NCP (145 ± 3-bp DNA) increases from 137 ± 5 Å to 165 ± 5 Å when C_s rises from 10 to 50 mM and remains constant with further increases of C_s up to 200 mM. In view of the very weak increase of the R_g value in the same C_s range, we attribute this D_max variation to tail extension, a proposal confirmed by simulations of the entire I(q) curves, considering an ideal solution of particles with tails either condensed or extended. This tail extension is observed at higher salt values when particles contain longer DNA fragments (165 ± 10 bp). The maximal extension of the tails always coincides with the screening of repulsive interactions between particles. The second virial coefficient becomes smaller than the hard sphere virial coefficient and eventually becomes negative (net attractive interactions) for NCP₁₄₅. Addition of salt simultaneously screens Coulombic repulsive interactions between NCP and Coulombic attractive interactions between tails and DNA inside the NCP. We discuss how the coupling of these two phenomena may be of biological relevance.     

Beyond the Nucleosome: Nucleosome-Protein Interactions and Higher Order Chromatin Structure

The regulation of chromatin biology ultimately depends on the manipulation of its smallest subunit, the nucleosome. The proteins that bind and operate on the nucleosome do so, while their substrate is part of a polymer embedded in the dense nuclear environment. Their molecular interactions must in some way be tuned to deal with this complexity. Due to the rapid increase in the number of high-resolution structures of nucleosome-protein complexes and the increasing understanding of the cellular chromatin structure, it is starting to become clearer how chromatin factors operate in this complex environment. In this review, we analyze the current literature on the interplay between nucleosome-protein interactions and higher-order chromatin structure. We examine in what way nucleosomes-protein interactions can affect and can be affected by chromatin organization at the oligonucleosomal level. In addition, we review the characteristics of nucleosome-protein interactions that can cause phase separation of chromatin. Throughout, we hope to illustrate the exciting challenges in characterizing nucleosome-protein interactions beyond the nucleosome.     

CENP-A Nucleosome is a Sensitive Allosteric Scaffold for DNA and Chromatin Factors

Centromeric loci of chromosomes are defined by nucleosomes containing the histone H3 variant CENP-A, which bind their DNA termini more permissively than their canonical counterpart, a feature that is critical for the mitotic fidelity. A recent cryo-EM study demonstrated that the DNA termini of CENP-A nucleosomes, reconstituted with the Widom 601 DNA sequence, are asymmetrically flexible, meaning one terminus is more clearly resolved than the other. However, an earlier work claimed that both ends could be resolved in the presence of two stabilizing single chain variable fragment (scFv) antibodies per nucleosome, and thus are likely permanently bound to the histone octamer. This suggests that the binding of scFv antibodies to the histone octamer surface would be associated with CENP-A nucleosome conformational changes, including stable binding of the DNA termini. Here, we present computational evidence that allows to explain at atomistic level the structural rearrangements of CENP-A nucleosomes resulting from the antibody binding. The antibodies, while they only bind the octamer façades, are capable of altering the dynamics of the nucleosomal core, and indirectly also the surrounding DNA. This effect has more drastic implications for the structure and the dynamics of the CENP-A nucleosome in comparison to its canonical counterpart. Furthermore, we find evidence that the antibodies bind the left and the right octamer façades at different affinities, another manifestation of the DNA sequence. We speculate that the cells could use induction of similar allosteric effects to control centromere function.     

The Accidental Ally: Nucleosome Barriers Can Accelerate Cohesin-Mediated Loop Formation in Chromatin

An important question in the context of the three-dimensional organization of chromosomes is the mechanism of formation of large loops between distant basepairs. Recent experiments suggest that the formation of loops might be mediated by loop extrusion factor proteins such as cohesin. Experiments on cohesin have shown that cohesins walk diffusively on the DNA and that nucleosomes act as obstacles to the diffusion, lowering the permeability and hence reducing the effective diffusion constant. An estimation of the times required to form the loops of typical sizes seen in Hi-C experiments using these low-effective-diffusion constants leads to times that are unphysically large. The puzzle then is the following: how does a cohesin molecule diffusing on the DNA backbone achieve speeds necessary to form the large loops seen in experiments? We propose a simple answer to this puzzle and show that although at low densities, nucleosomes act as barriers to cohesin diffusion, beyond a certain concentration they can reduce loop formation times because of a subtle interplay between the nucleosome size and the mean linker length. This effect is further enhanced on considering stochastic binding kinetics of nucleosomes on the DNA backbone and leads to predictions of lower loop formation times than might be expected from a naive obstacle picture of nucleosomes.     

The Effect of DNA CpG Methylation on the Dynamic Conformation of a Nucleosome

DNA methylation is an important epigenetic mark that is known to induce chromatin condensation and gene silencing. We used a time-domain fluorescence lifetime measurement to quantify the effects of DNA hypermethylation on the conformation and dynamics of a nucleosome. Nucleosomes reconstituted on an unmethylated and a methylated DNA both exhibit dynamic conformations under physiological conditions. The DNA end breathing motion and the H2A-H2B dimer destabilization dominate the dynamic behavior of nucleosomes at low to medium ionic strength. Extensive DNA CpG methylation, surprisingly, does not help to restrain the DNA breathing motion, but facilitates the formation of a more open nucleosome conformation. The presence of the divalent cation, Mg²⁺, essential for chromatin compaction, and the methyl donor molecule SAM, required for DNA methyltransferase reaction, facilitate the compaction of both types of nucleosomes. The difference between the unmethylated and the methylated nucleosome persists within a broad range of salt concentrations, but vanishes under high magnesium concentrations. Reduced DNA backbone rigidity due to the presence of methyl groups is believed to contribute to the observed structural and dynamic differences. The observation of this study suggests that DNA methylation alone does not compact chromatin at the nucleosomal level and provides molecular details to understand the regulatory role of DNA methylation in gene expression.