The tale beyond the tail: histone core domain modifications and the regulation of chromatin structure

Histone post-translational modifications occur, not only in the N-terminal tail domains, but also in the core domains. While modifications in the N-terminal tail function largely through the regulation of the binding of non-histone proteins to chromatin, based on their location in the nucleosome, core domain modifications may also function through distinct mechanisms involving structural alterations to the nucleosome. This article reviews the recent developments in regards to these novel histone modifications and discusses their important role in the regulation of chromatin structure.     

Structures and interactions of the core histone tail domains

The core histone tail domains are "master control switches" that help define the structural and functional characteristics of chromatin at many levels. The tails modulate DNA accessibility within the nucleosome, are essential for stable folding of oligonucleosome arrays into condensed chromatin fibers, and are important for fiber-fiber interactions involved in higher order structures. Many nuclear signaling pathways impinge upon the tail domains, resulting in posttranslational modifications that are likely to alter the charge, structure, and/or interactions of the core histone tails or to serve as targets for the binding of ancillary proteins or other enzymatic functions. However, currently we have only a marginal understanding of the molecular details of core histone tail conformations and contacts. Here we review data related to the structures and interactions of the core histone tail domains and how these domains and posttranslational modifications therein may define the structure and function of chromatin.     

Delineation of the protein module that anchors HMGN proteins to nucleosomes in the chromatin of living cells

Numerous nuclear proteins bind to chromatin by targeting unique DNA sequences or specific histone modifications. In contrast, HMGN proteins recognize the generic structure of the 147-bp nucleosome core particle. HMGNs alter the structure and activity of chromatin by binding to nucleosomes; however, the determinants of the specific interaction of HMGNs with chromatin are not known. Here we use systematic mutagenesis, quantitative fluorescence recovery after photobleaching, fluorescence imaging, and mobility shift assays to identify the determinants important for the specific binding of these proteins to both the chromatin of living cells and to purified nucleosomes. We find that several regions of the protein affect the affinity of HMGNs to chromatin; however, the conserved sequence RRSARLSA, is the sole determinant of the specific interaction of HMGNs with nucleosomes. Within this sequence, each of the 4 amino acids in the R-S-RL motif are the only residues absolutely essential for anchoring HMGN protein to nucleosomes, both in vivo and in vitro. Our studies identify a new chromatin-binding module that specifically recognizes nucleosome cores independently of DNA sequence or histone tail modifications.     

Influence of combinatorial histone modifications on antibody and effector protein recognition

Increasing evidence suggests that histone posttranslational modifications (PTMs) function in a combinatorial fashion to regulate the diverse activities associated with chromatin. Yet how these patterns of histone PTMs influence the adapter proteins known to bind them is poorly understood. In addition, how histone-specific antibodies are influenced by these same patterns of PTMs is largely unknown. Here we examine the binding properties of histone-specific antibodies and histone-interacting proteins using peptide arrays containing a library of combinatorially modified histone peptides. We find that modification-specific antibodies are more promiscuous in their PTM recognition than expected and are highly influenced by neighboring PTMs. Furthermore, we find that the binding of histone-interaction domains from BPTF, CHD1, and RAG2 to H3 lysine 4 trimethylation is also influenced by combinatorial PTMs. These results provide further support for the histone code hypothesis and raise specific concerns with the quality of the currently available modification-specific histone antibodies.     

The effect of epigenetic modifications on the secondary structures and possible binding positions of the N-terminal tail of histone H3 in the nucleosome: a computational study

The roles of histone tails as substrates for reversible chemical modifications and dynamic cognate surfaces for the binding of regulatory proteins are well established. Despite these crucial roles, experimentally derived knowledge of the structure and possible binding sites of histone tails in chromatin is limited. In this study, we utilized molecular dynamics of isolated histone H3 N-terminal peptides to investigate its structure as a function of post-translational modifications that are known to be associated with defined chromatin states. We observed a structural preference for α-helices in isoforms associated with an inactive chromatin state, while isoforms associated with active chromatin states lacked α-helical content. The physicochemical effect of the post-translational modifications was highlighted by the interaction of arginine side-chains with the phosphorylated serine residues in the inactive isoform. We also showed that the isoforms exhibit different tail lengths, and, using molecular docking of the first 15 N-terminal residues of an H3 isoform, identified potential binding sites between the superhelical gyres on the octamer surface, close to the site of DNA entry/exit in the nucleosome. We discuss the possible functional role of the binding of the H3 tail within the nucleosome on both nucleosome and chromatin structure and stability.     

Role of the conserved Sir3-BAH domain in nucleosome binding and silent chromatin assembly

Silent chromatin domains in Saccharomyces cerevisiae represent examples of epigenetically heritable chromatin. The formation of these domains involves the recruitment of the SIR complex, composed of Sir2, Sir3, and Sir4, followed by iterative cycles of NAD-dependent histone deacetylation and spreading of SIR complexes over adjacent chromatin domains. We show here that the conserved bromo-adjacent homology (BAH) domain of Sir3 is a nucleosome- and histone-tail-binding domain and that its binding to nucleosomes is regulated by residues in the N terminus of histone H4 and the globular domain of histone H3 on the exposed surface of the nucleosome. Furthermore, using a partially purified system containing nucleosomes, the three Sir proteins, and NAD, we observe the formation of SIR-nucleosome filaments with a diameter of less than 20 nm. Together, these observations suggest that the SIR complex associates with an extended chromatin fiber through interactions with two different regions in the nucleosome.     

The core histone tail domains contribute to sequence-dependent nucleosome positioning

The precise positioning of nucleosomes plays a critical role in the regulation of gene expression by modulating the DNA binding activity of trans-acting factors. However, molecular determinants responsible for positioning are not well understood. We examined whether the removal of the core histone tail domains from nucleosomes reconstituted with specific DNA fragments led to alteration of translational positions. Remarkably, we find that removal of tail domains from a nucleosome assembled on a DNA fragment containing a Xenopus borealis somatic-type 5S RNA gene results in repositioning of nucleosomes along the DNA, including two related major translational positions that move about 20 bp further upstream with respect to the 5S gene. In a nucleosome reconstituted with a DNA fragment containing the promoter of a Drosophila alcohol dehydrogenase gene, several translational positions shifted by about 10 bp along the DNA upon tail removal. However, the positions of nucleosomes assembled with a DNA fragment known to have one of the highest binding affinities for core histone proteins in the mouse genome were not altered by removal of core histone tail domains. Our data support the notion that the basic tail domains bind to nucleosomal DNA and influence the selection of the translational position of nucleosomes and that once tails are removed movement between translational positions occurs in a facile manner on some sequences. However, the effect of the N-terminal tails on the positioning and movement of a nucleosome appears to be dependent on the DNA sequence such that the contribution of the tails can be masked by very high affinity DNA sequences. Our results suggest a mechanism whereby sequence-dependent nucleosome positioning can be specifically altered by regulated changes in histone tail-DNA interactions in chromatin.     

Histone proteomics and the epigenetic regulation of nucleosome mobility

Chromatin structure plays a vital role in the transmission of heritable gene expression patterns. The recent application of mass spectrometry to histone biology provides several striking insights into chromatin regulation. The continuing identification of new histone post-translational modifications is revolutionizing the ways in which we think about how access to genomic DNA is controlled. While post-translational modifications of the flexible histone tails continue to be an active area of investigation, the recent discovery of multiple modifications in the structured globular domains of histones provides new insights into how the nucleosome works. Recent experiments underscore the importance of a subgroup of these modifications: those that regulate histone-DNA interactions on the lateral surface of the nucleosome. This information highlights an emerging new paradigm in chromatin control, that of the epigenetic regulation of nucleosome mobility.     

The nucleosome: a little variation goes a long way.

Changes in the overall structure of chromatin are essential for the proper regulation of cellular processes, including gene activation and silencing, DNA repair, chromosome segregation during mitosis and meiosis, X chromosome inactivation in female mammals, and chromatin compaction during apoptosis. Such alterations of the chromatin template occur through at least 3 interrelated mechanisms: post-translational modifications of histones, ATP-dependent chromatin remodeling, and the incorporation (or replacement) of specialized histone variants into chromatin. Of these mechanisms, the exchange of variants into and out of chromatin is the least well understood. However, the exchange of conventional histones for variant histones has distinct and profound consequences within the cell. This review focuses on the growing number of mammalian histone variants, their particular biological functions and unique features, and how they may affect the structure of the nucleosome. We propose that a given nucleosome might not consist of heterotypic variants, but rather, that only specific histone variants come together to form a homotypic nucleosome, a hypothesis that we refer to as the nucleosome code. Such nucleosomes might in turn participate in marking specific chromatin domains that may contribute to epigenetic inheritance.