Incorporation Of Histone Variants Research Articles

Histone variants: emerging players in cancer biology

Histone variants are key players in shaping chromatin structure, and, thus, in regulating fundamental cellular processes such as chromosome segregation and gene expression. Emerging evidence points towards a role for histone variants in contributing to tumor progression, and, recently, the first cancer-associated mutation in a histone variant-encoding gene was reported. In addition, genetic alterations of the histone chaperones that specifically regulate chromatin incorporation of histone variants are rapidly being uncovered in numerous cancers. Collectively, these findings implicate histone variants as potential drivers of cancer initiation and/or progression, and, therefore, targeting histone deposition or the chromatin remodeling machinery may be of therapeutic value. Here, we review the mammalian histone variants of the H2A and H3 families in their respective cellular functions, and their involvement in tumor biology.

MacroH2A – An epigenetic regulator of cancer

Epigenetic regulation is one of the most promising and expanding areas of cancer research. One of the emerging, but least understood aspects of epigenetics is the facultative and locus-specific incorporation of histone variants and their function in chromatin. With the characterization of the first loss of function phenotypes of the macroH2A histone variants, previously unrecognized epigenetic mechanisms have now moved into the spotlight of cancer research. Here, we summarize data supporting different molecular mechanisms that could mediate the primarily tumor suppressive function of macroH2A. We further discuss context-dependent and isoform-specific functions. The aim of this review is to provide guidance for those assessing macroH2A’s potential as biomarker or therapeutic intervention point.

Redox Homeostasis and Epigenetics in Non-alcoholic Fatty Liver Disease (NAFLD)

Non-alcoholic fatty liver disease (NAFLD), an accumulation of intra-hepatic triglycerides that is often considered the hepatic manifestation of insulin resistance, is the most common cause of chronic liver disease in the Western countries with up to one third of the population affected. NAFLD is a spectrum of disturbances that encompasses various degrees of liver damage ranging from simple steatosis to non-alcoholic steatohepatitis (NASH). NASH is characterized by hepatocellular injury/inflammation with or without fibrosis. The individuals with NAFLD develop NASH in 10% of the cases, and are also at risk of developing hepatocellular carcinoma (HCC). Epigenetic mechanisms of nuclear chromatin remodeling, such as DNA methylation, post-translational modifications of histones, and incorporation of histone variants into the chromatin are increasingly recognized as crucial factors in the pathophysiology of NAFLD. NAFLD is often accompanied by oxidative stress: reactive oxygen species (ROS) are implicated in altered reduction/oxidation (redox) reactions that attack cellular macromolecules and are detected in the liver of patients and animal models of NAFLD. In this review, we summarize recent knowledge advancements in the hepatic epigenetic and redox mechanisms, and their possible links, involved in the pathogenesis and treatment of NAFLD.

Epigenetics and Novel Therapeutic Approaches

Epigenetic changes are targeted to the DNA, as well as chromatin core histones and unlike genetic mutations these changes are reversible, making them attractive targets for therapeutic purposes [1]. Multiple levels of epigenetic regulatory mechanisms provide orchestration of chromatin structure for gene expression in healthy cells; however, result inappropriate silencing or activation of gene expressions, finally leading to various disease states such as cancer [2]. Epigenetic modifications can be a determinant factor to characterize the disease states, and detailed profiling of these modifications may allow predictions of disease outcome or therapeutic strategies. Epigenetic therapy can offer promising tools to restore/reverse changes. Interestingly, several epigenetic inhibitors such as DNA methyltransferase inhibitors (DNMTis), and histone deacetylase inhibitors (HDACis) approved by the US Food and Drug Administration (FDA) are in clinics for the treatment of various hematological malignancies. Potential epigenetic modifications, such as DNA methylation, covalent histone modifications, such as histone acetylation/deacetylation, histone methylation/demethylation, histone phosphorylation, non-covalent mechanisms, such as incorporation of histone variants, nucleosome remodeling and positioning, microRNAs (miRNAs) expression are important targets for therapeutic purposes [2,3]. How these effectors interact to each other to affect gene expression and cause disease is the active field of study in epigenetics.

Introduction: The Use of Animals Models to Advance Epigenetic Science

Introduction: The Use of Animals Models to Advance Epigenetic Science

Deposition of Histone Variant H2A.Z within Gene Bodies Regulates Responsive Genes

The regulation of eukaryotic chromatin relies on interactions between many epigenetic factors, including histone modifications, DNA methylation, and the incorporation of histone variants. H2A.Z, one of the most conserved but enigmatic histone variants that is enriched at the transcriptional start sites of genes, has been implicated in a variety of chromosomal processes. Recently, we reported a genome-wide anticorrelation between H2A.Z and DNA methylation, an epigenetic hallmark of heterochromatin that has also been found in the bodies of active genes in plants and animals. Here, we investigate the basis of this anticorrelation using a novel h2a.z loss-of-function line in Arabidopsis thaliana. Through genome-wide bisulfite sequencing, we demonstrate that loss of H2A.Z in Arabidopsis has only a minor effect on the level or profile of DNA methylation in genes, and we propose that the global anticorrelation between DNA methylation and H2A.Z is primarily caused by the exclusion of H2A.Z from methylated DNA. RNA sequencing and genomic mapping of H2A.Z show that H2A.Z enrichment across gene bodies, rather than at the TSS, is correlated with lower transcription levels and higher measures of gene responsiveness. Loss of H2A.Z causes misregulation of many genes that are disproportionately associated with response to environmental and developmental stimuli. We propose that H2A.Z deposition in gene bodies promotes variability in levels and patterns of gene expression, and that a major function of genic DNA methylation is to exclude H2A.Z from constitutively expressed genes.

Histone H2A variants in nucleosomes and chromatin: more or less stable?

In eukaryotes, DNA is organized together with histones and non-histone proteins into a highly complex nucleoprotein structure called chromatin, with the nucleosome as its monomeric subunit. Various interconnected mechanisms regulate DNA accessibility, including replacement of canonical histones with specialized histone variants. Histone variant incorporation can lead to profound chromatin structure alterations thereby influencing a multitude of biological processes ranging from transcriptional regulation to genome stability. Among core histones, the H2A family exhibits highest sequence divergence, resulting in the largest number of variants known. Strikingly, H2A variants differ mostly in their C-terminus, including the docking domain, strategically placed at the DNA entry/exit site and implicated in interactions with the (H3–H4)2-tetramer within the nucleosome and in the L1 loop, the interaction interface of H2A–H2B dimers. Moreover, the acidic patch, important for internucleosomal contacts and higher-order chromatin structure, is altered between different H2A variants. Consequently, H2A variant incorporation has the potential to strongly regulate DNA organization on several levels resulting in meaningful biological output. Here, we review experimental evidence pinpointing towards outstanding roles of these highly variable regions of H2A family members, docking domain, L1 loop and acidic patch, and close by discussing their influence on nucleosome and higher-order chromatin structure and stability.

Placing the Plasmodium falciparum epigenome on the map

It is becoming increasingly evident that epigenetic mechanisms that act on and regulate chromatin structure play a key role in the development, adaptation, and survival of the malaria parasite within its human host. The study of epigenetics in Plasmodium falciparum started to flourish in recent years due to improvement of genomic technologies. Here we summarize the knowledge gained from genome-wide localization profiling of different epigenetic features, and discuss hypotheses emerging from the analysis of these 'descriptive' epigenetic maps. Furthermore, we highlight key questions to be answered, and provide a glimpse of developments required to gain true mechanistic understanding and to lift this maturing field to the next level.

Inactivation or non-reactivation: what accounts better for the silence of sex chromosomes during mammalian male meiosis?

During the first meiotic prophase in male mammals, sex chromosomes undergo a program of transcriptional silencing called meiotic sex chromosome inactivation (MSCI). MSCI is triggered by accumulation of proteins like BRCA1, ATR, and γH2AX on unsynapsed chromosomes, followed by local changes on the sex chromatin, including histone modifications, incorporation of specific histone variants, non-histone proteins, and RNAs. It is generally thought that MSCI represents the transition of unsynapsed chromatin from a transcriptionally active state to a repressed state. However, transcription is generally low in the whole nucleus during the early stages of the first meiotic prophase, when markers of MSCI first appear, and is then reactivated globally during pachytene. Thus, an alternative possibility is that MSCI represents the targeted maintenance and/or reinforcement of a prior repressed state, i.e., a failure to reactivate. Here, we present an analysis of the temporal and spatial appearance of transcriptional and MSCI markers, as well as chromatin modifications related to transcriptional regulation. We show that levels of RNA pol II and histone H3 acetylated at lysine 9 (H3K9ac) are low during leptotene, zygotene, and early pachytene, but increase strongly in mid-pachytene, indicating that reactivation occurs with some delay after synapsis. However, while transcription markers appear abundantly on the autosomes at mid-pachytene, they are not directed to the sex chromosomes. Interestingly, we found that chromatin modifications related to transcriptional silencing and/or MSCI, namely, histone H3 trimethylated at lysine 9 (H3K9me3), histone H3 monomethylated at lysine 4 (H3K4me1), γH2AX, SUMO1, and XMR, appear on the sex chromosomes before autosomes become reactivated. These results suggest that the onset of MSCI during late zygotene and early pachytene may prevent sex chromosome reactivation during mid-pachytene instead of promoting inactivation de novo. Additionally, we found temporal differences between the X and Y chromosomes in the recruitment of DNA repair and MSCI markers, indicating a differential regulation of these processes. We propose that many of the meiotic defects attributed to failure to silence sex chromosomes could be interpreted as a more general process of transcriptional misregulation that occurs under certain pathological circumstances in zygotene and early pachytene.

Radiation-induced alterations in histone modification patterns and their potential impact on short-term radiation effects

Detection and repair of radiation-induced DNA damage occur in the context of chromatin. An intricate network of mechanisms defines chromatin structure, including DNA methylation, incorporation of histone variants, histone modifications, and chromatin remodeling. In the last years it became clear that the cellular response to radiation-induced DNA damage involves all of these mechanisms. Here we focus on the current knowledge on radiation-induced alterations in post-translational histone modification patterns and their effect on the chromatin accessibility, transcriptional regulation and chromosomal stability.

Chromatin Structure and Dynamics: Histone Variants and Remodeling Complexes

The organization of DNA into nucleosome can be seen as a barrier for the transcription factors binding to their target DNA sequences and interferes with several basic cellular processes. ATP-remodeling machines and the incorporation of histone variants into chromatin are used by the cell to overcome the nucleosomal barrier and modulate DNA accessibility by the control of nucleosome dynamics. In this work, we use a single molecule technique (Atomic Force Microscopy, AFM) to visualize isolated mono- and oligo-nucleosomes and quantify their structure and dynamics at thermodynamical equilibrium and out of equilibrium.We have studied the impact of H2A.Bbd histone variant incorporation at the mono-nucleosome and oligo-nucleosome level, and we have shown that this variant modifies both the structure and the dynamics of the nucleosome altering the ability to form a higher structure organization of the chromatin. Using a polymer physics model, we demonstrated that the behavior of variant chromatin can be quantitatively explained by the mono-nucleosome flexibility [1].We are also interested in the mechanism of nucleosome remodeling by SWI/SNF and RSC on mono- and di-nucleosomes. Focusing on the di-nucleosomes, AFM imaging showed only a limited set of distinct configurational states for the remodeling products. No stepwise or preferred directionality of the nucleosome motion was observed [2]. Analysis of the corresponding reaction pathways allows deciphering the mechanistic features of RSC-induced nucleosome relocation. The final outcome of RSC remodeling of oligosomal templates is the packing of the nucleosomes at the edge of the template, providing large stretches of nucleosome depleted DNA.[1] F. Montel, H. Menoni, M. Castelnovo, J. Bednar, S. Dimitrov, D. Angelov & C. Faivre-Moskalenko; Biophys. J. 97, 544-553 (2009).[2] F. Montel, M. Castelnovo, H. Menoni, D. Angelov, S. Dimitrov & C. Faivre-Moskalenko; Nucleic Acids Res. 39, 2571-9 (2011).

The Histone Database: an integrated resource for histones and histone fold-containing proteins

Eukaryotic chromatin is composed of DNA and protein components—core histones—that act to compactly pack the DNA into nucleosomes, the fundamental building blocks of chromatin. These nucleosomes are connected to adjacent nucleosomes by linker histones. Nucleosomes are highly dynamic and, through various core histone post-translational modifications and incorporation of diverse histone variants, can serve as epigenetic marks to control processes such as gene expression and recombination. The Histone Sequence Database is a curated collection of sequences and structures of histones and non-histone proteins containing histone folds, assembled from major public databases. Here, we report a substantial increase in the number of sequences and taxonomic coverage for histone and histone fold-containing proteins available in the database. Additionally, the database now contains an expanded dataset that includes archaeal histone sequences. The database also provides comprehensive multiple sequence alignments for each of the four core histones (H2A, H2B, H3 and H4), the linker histones (H1/H5) and the archaeal histones. The database also includes current information on solved histone fold-containing structures. The Histone Sequence Database is an inclusive resource for the analysis of chromatin structure and function focused on histones and histone fold-containing proteins.Database URL: The Histone Sequence Database is freely available and can be accessed at http://research.nhgri.nih.gov/histones/.

Key Functional Regions in the Histone Variant H2A.Z C-Terminal Docking Domain

The incorporation of histone variants into nucleosomes represents one way of altering the chromatin structure to accommodate diverse functions. Histone variant H2A.Z has specific roles in gene regulation, heterochromatin boundary formation, and genomic integrity. The precise features required for H2A.Z to function and specify an identity different from canonical H2A remain to be fully explored. Analysis of the C-terminal docking domain of H2A.Z in Saccharomyces cerevisiae using epistatic miniarray profile (E-MAP) uncovered nuanced requirements of the H2A.Z C-terminal region for cell growth when additional genes were compromised. Moreover, the H2A.Z(1-114) truncation, lacking the last 20 amino acids of the protein, did not support regular H2A.Z functions, such as resistance to genotoxic stress, restriction of heterochromatin in its native context, GAL1 gene activation, and chromatin anchoring. The corresponding region of H2A could fully rescue the strong defects caused by loss of this functionally essential region in the C terminus of H2A.Z. Despite the dramatic reduction in function, the H2A.Z(1-114) truncation still bound the H2A.Z deposition complex SWR1-C, the histone chaperone Chz1, and histone H2B. These data are consistent with a model in which retaining the variant in chromatin after its deposition by SWR1-C is a crucial determinant of its function.

Epigenetics in C. elegans: Facts and challenges

Epigenetics is defined as the study of heritable changes in gene expression that are not accompanied by changes in the DNA sequence. Epigenetic mechanisms include histone post-translational modifications, histone variant incorporation, non-coding RNAs, and nucleosome remodeling and exchange. In addition, the functional compartmentalization of the nucleus also contributes to epigenetic regulation of gene expression. Studies on the molecular mechanisms underlying epigenetic phenomena and their biological function have relied on various model systems, including yeast, plants, flies, and cultured mammalian cells. Here we will expose the reader to the current understanding of epigenetic regulation in the roundworm C. elegans. We will review recent models of nuclear organization and its impact on gene expression, the biological role of enzymes modifying core histones, and the function of chromatin-associated factors, with special emphasis on Polycomb (PcG) and Trithorax (Trx-G) group proteins. We will discuss how the C. elegans model has provided novel insight into mechanisms of epigenetic regulation as well as suggest directions for future research.

Phosphorylation of Histone H2B Serine 32 Is Linked to Cell Transformation

Various types of post-translational modifications of the histone tails have been revealed, but a few modifications have been found within the histone core sequences. Histone core post-translational modifications have the potential to modulate nucleosome structure and DNA accessibility. Here, we studied the histone H2B core domain and found that phosphorylation of H2B serine 32 occurs in normal cycling and mitogen-stimulated cells. Notably, this phosphorylation is elevated in skin cancer cell lines and tissues compared with normal counterparts. The JB6 Cl41 mouse skin epidermal cell line is a well established model for tumor promoter-induced cell transformation and was used to study the function of H2B during EGF-induced carcinogenesis. Remarkably, cells overexpressing a nonphosphorylatable H2BS32A mutant exhibited suppressed growth and EGF-induced cell transformation, possibly because of decreased activation of activator protein-1, compared with control cells overexpressing wild type H2B. We identified ribosomal S6 kinase 2 (RSK2) as the kinase responsible for H2BS32 phosphorylation. Serum-starved JB6 cells contain very little endogenous H2BS32 phosphorylation, and EGF treatment induced this phosphorylation. The phosphorylation was attenuated in RSK2 knock-out MEFs and RSK2 knockdown JB6 cells. Taken together, our results demonstrate a novel role for H2B phosphorylation in cell transformation and show that H2BS32 phosphorylation is critical for controlling activator protein-1 activity, which is a major driver in cell transformation.

Chromatin Response to DNA Double-Strand Break Damage

Manipulation of chromatin, in which genomic DNA is packaged, is a fundamental requirement for all DNA-based metabolic processes in eukayotic cells. Histone variant incorporation, histone post-translational modifications, and ATP-dependent chromatin remodeling are three major strategies for chromatin manipulation, and are relatively well characterized in transcriptional regulation. Emerging lines of evidence indicate that histone variants (H2AX and H2A.Z), histone post-translational modifications (acetylation, phosphorylation, methylation and ubiquitination) and chromatin-remodeling complexes (INO80, SWR1, SWI/SNF, RSC and NuRD) are important and direct players in the DNA double-strand break (DSB) response as well. New studies also reveal that incorporation of histone variants into nucleosomes, histone modifications and ATP-dependent chromatin remodeling are specifically and intimately connected during the DSB damage response. This article summarizes the recent advances in our understanding of the relationship between chromatin modifications and the DSB damage response.

Special issue on epigenetic inheritance by histone modifications, histone variants and non-coding RNAs

Keeping in view the ever-growing importance of understanding the epigenetic phenomena shaping the behavior of life, our team decided to embark on the idea to organize this special issue of Frontiers in Biology on Epigenetics. Epigenetics refers to the study of heritable changes in gene expression without changes in DNA sequence, which is accomplished by DNA methylation, histone modifications, histone variants, chromatin remodeling, and non-coding RNAs. With the course of time the mechanisms underlying epigenetic regulation are getting inter-connected to each other. This issue comprises eight exciting invited review articles providing a mechanistic and comprehensive insight into different levels of epigenetic inheritance. Majority of the articles highlight the epigenetic regulation by histone modifications, histone variants and non-coding RNAs. This issue of the journal also features a selection of two regular articles, not related to Epigenetics, which proves our devotion to publish all quality articles on a timely basis. The long-sought-after debates about how histone modification patterns faithfully propagate into daughter cells is reviewed. Hui Wu and Bing Zhu elegantly overviewed two historic models for H3-H4 tetramer spilt or non-spilt decision that govern epigenetic inheritance of histone modifications. The deposition of histone variants to nucleosome adds an additional layer of complexity to the epigenetic regulation. The review article by Leilei Shi and Yuda Fang summarizes the diverse roles played by the incorporation of histone variants into the nucleosome in various biological processes. Wenchao Yin et al. outlined that repeat sequences contribute to centromere formation but are not sufficient to establish and maintain it without epigenetic components. Ying Cao and Ligeng Ma discussed the mechanism of dynamic H2B ubiquitination; the biological functions as well as the molecular mechanisms of H2Bub1. Dosage compensation refers to the equal expression from chromosomes at different copy numbers. James A. Birchler et al. in this special issue described the implications of the gene balance hypothesis for dosage compensation. Non-coding RNAs are functional RNA molecules that are not translated into proteins including microRNAs, siRNAs and piRNAs and the long ncRNAs etc. Yun Ju Kim and Xuemei Chen reviewed the roles of Mediator, a conserved multiprotein cofactor of RNA Polymerase II in epigenetic regulation via non-coding RNA production. The review by Xiaoyun Jia et al. highlights the key features of newly emerging miRNA-directed DNA methylation in plants. Dacheng Liang et al. presented a model, based on the current knowledge, of the systemic gene silencing in plants mediated by non-coding RNAs. The current issue does not cover all the aspects of highly diverse field of Epigenetics; however, it does present some of the interesting areas of this rapidly growing field of science.We are convinced that this effort will surely further our current understanding of epigenetic mechanisms involved in countless biological phenomena and will pave the path for the future work to understand many known as well as unknown biological enigmas. We would like to cordially thank all of the authors who generously contributed their knowledge and wisdom to the fulfillment of this effort.

Histone variants: making structurally and functionally divergent nucleosomes and linkers in chromatin

In addition to the post-translational modifications of histone proteins, emerging literature suggests that the mosaic nucleosomes formed by incorporation of various histone variants provide another mechanism for modifying chromatin structure and function. The locally defined chromatin by histone variants is involved in transcriptional regulation, DNA repair, centromere packaging, maintenance of pericentromeric heterochromatin, stress responses, temperature sensing, development, and many other biological processes. Here, we review the universal histone variants in H2A, H3 and H1 families and their roles in epigenetics.

Recent progress in the epigenetics and chromatin field

Recent progress in the epigenetics and chromatin field

Histone H2A.Z acid patch residues required for deposition and function

The incorporation of histone variants is one mechanism used by the eukaryotic cell to alter the generally repressive chromatin template. However, the exact molecular mechanisms that direct this incorporation are not well understood. The SWR1 chromatin remodeling complex that binds to and directs incorporation of histone variant H2A.Z into chromatin has been characterized, but significantly less information is available concerning the requirements on the H2A.Z target molecule. We performed an unbiased mutagenic screen designed to elucidate the function of H2A.Z in Saccharomyces cerevisiae. The screen identified residues within the conserved acidic patch of H2A.Z as being important for the function of the variant. We characterized single point mutations in the patch that are phenotypically sensitive to a variety of growth conditions and are expressed at lower protein levels, but are functionally defective (htz1-D99A, htz1-D99K, and htz1-E101K). The mutants were significantly less detectable by chromatin immunoprecipitation at PHO5, a gene previously described to be enriched for H2A.Z. These results identify acidic patch residues of H2A.Z that are critical for mediating deposition and function in chromatin, and represent potential candidates for the interaction of H2A.Z with its deposition and/or targeting machinery.