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Sample records for core histone variant

  1. Histones in functional diversification. Core histone variants.

    Science.gov (United States)

    Pusarla, Rama-Haritha; Bhargava, Purnima

    2005-10-01

    Recent research suggests that minor changes in the primary sequence of the conserved histones may become major determinants for the chromatin structure regulating gene expression and other DNA-related processes. An analysis of the involvement of different core histone variants in different nuclear processes and the structure of different variant nucleosome cores shows that this may indeed be so. Histone variants may also be involved in demarcating functional regions of the chromatin. We discuss in this review why two of the four core histones show higher variation. A comparison of the status of variants in yeast with those from higher eukaryotes suggests that histone variants have evolved in synchrony with functional requirement of the cell.

  2. Identification and characterization of the genes encoding the core histones and histone variants of Neurospora crassa.

    OpenAIRE

    Hays, Shan M.; Swanson, Johanna; Selker, Eric U.

    2002-01-01

    We have identified and characterized the complete complement of genes encoding the core histones of Neurospora crassa. In addition to the previously identified pair of genes that encode histones H3 and H4 (hH3 and hH4-1), we identified a second histone H4 gene (hH4-2), a divergently transcribed pair of genes that encode H2A and H2B (hH2A and hH2B), a homolog of the F/Z family of H2A variants (hH2Az), a homolog of the H3 variant CSE4 from Saccharomyces cerevisiae (hH3v), and a highly diverged ...

  3. Identification and characterization of the genes encoding the core histones and histone variants of Neurospora crassa.

    OpenAIRE

    Hays, Shan M.; Swanson, Johanna; Selker, Eric U.

    2002-01-01

    We have identified and characterized the complete complement of genes encoding the core histones of Neurospora crassa. In addition to the previously identified pair of genes that encode histones H3 and H4 (hH3 and hH4-1), we identified a second histone H4 gene (hH4-2), a divergently transcribed pair of genes that encode H2A and H2B (hH2A and hH2B), a homolog of the F/Z family of H2A variants (hH2Az), a homolog of the H3 variant CSE4 from Saccharomyces cerevisiae (hH3v), and a highly diverged ...

  4. Histone Variants and Epigenetics

    Science.gov (United States)

    Henikoff, Steven; Smith, M. Mitchell

    2015-01-01

    Histones package and compact DNA by assembling into nucleosome core particles. Most histones are synthesized at S phase for rapid deposition behind replication forks. In addition, the replacement of histones deposited during S phase by variants that can be deposited independently of replication provide the most fundamental level of chromatin differentiation. Alternative mechanisms for depositing different variants can potentially establish and maintain epigenetic states. Variants have also evolved crucial roles in chromosome segregation, transcriptional regulation, DNA repair, and other processes. Investigations into the evolution, structure, and metabolism of histone variants provide a foundation for understanding the participation of chromatin in important cellular processes and in epigenetic memory. PMID:25561719

  5. Distinct features of the histone core structure in nucleosomes containing the histone H2A.B variant

    National Research Council Canada - National Science Library

    Sugiyama, Masaaki; Arimura, Yasuhiro; Shirayama, Kazuyoshi; Fujita, Risa; Oba, Yojiro; Sato, Nobuhiro; Inoue, Rintaro; Oda, Takashi; Sato, Mamoru; Heenan, Richard K; Kurumizaka, Hitoshi

    2014-01-01

    .... Comparisons with the canonical H2A nucleosome structure revealed that the DNA termini of the H2A.B nucleosome are detached from the histone core surface, and flexibly expanded toward the solvent...

  6. Histone variants and lipid metabolism

    NARCIS (Netherlands)

    Borghesan, Michela; Mazzoccoli, Gianluigi; Sheedfar, Fareeba; Oben, Jude; Pazienza, Valerio; Vinciguerra, Manlio

    2014-01-01

    Within nucleosomes, canonical histones package the genome, but they can be opportunely replaced with histone variants. The incorporation of histone variants into the nucleosome is a chief cellular strategy to regulate transcription and cellular metabolism. In pathological terms, cellular steatosis

  7. Histone variants: key players of chromatin.

    Science.gov (United States)

    Biterge, Burcu; Schneider, Robert

    2014-06-01

    Histones are fundamental structural components of chromatin. Eukaryotic DNA is wound around an octamer of the core histones H2A, H2B, H3, and H4. Binding of linker histone H1 promotes higher order chromatin organization. In addition to their structural role, histones impact chromatin function and dynamics by, e.g., post-translational histone modifications or the presence of specific histone variants. Histone variants exhibit differential expression timings (DNA replication-independent) and mRNA characteristics compared to canonical histones. Replacement of canonical histones with histone variants can affect nucleosome stability and help to create functionally distinct chromatin domains. In line with this, several histone variants have been implicated in the regulation of cellular processes such as DNA repair and transcriptional activity. In this review, we focus on recent progress in the study of core histone variants H2A.X, H2A.Z, macroH2A, H3.3, and CENP-A, as well as linker histone H1 variants, their functions and their links to development and disease.

  8. Histone variants and lipid metabolism

    NARCIS (Netherlands)

    Borghesan, Michela; Mazzoccoli, Gianluigi; Sheedfar, Fareeba; Oben, Jude; Pazienza, Valerio; Vinciguerra, Manlio

    2014-01-01

    Within nucleosomes, canonical histones package the genome, but they can be opportunely replaced with histone variants. The incorporation of histone variants into the nucleosome is a chief cellular strategy to regulate transcription and cellular metabolism. In pathological terms, cellular steatosis i

  9. The Structural Determinants behind the Epigenetic Role of Histone Variants

    Directory of Open Access Journals (Sweden)

    Manjinder S. Cheema

    2015-07-01

    Full Text Available Histone variants are an important part of the histone contribution to chromatin epigenetics. In this review, we describe how the known structural differences of these variants from their canonical histone counterparts impart a chromatin signature ultimately responsible for their epigenetic contribution. In terms of the core histones, H2A histone variants are major players while H3 variant CenH3, with a controversial role in the nucleosome conformation, remains the genuine epigenetic histone variant. Linker histone variants (histone H1 family haven’t often been studied for their role in epigenetics. However, the micro-heterogeneity of the somatic canonical forms of linker histones appears to play an important role in maintaining the cell-differentiated states, while the cell cycle independent linker histone variants are involved in development. A picture starts to emerge in which histone H2A variants, in addition to their individual specific contributions to the nucleosome structure and dynamics, globally impair the accessibility of linker histones to defined chromatin locations and may have important consequences for determining different states of chromatin metabolism.

  10. HistoneDB 2.0: a histone database with variants--an integrated resource to explore histones and their variants.

    Science.gov (United States)

    Draizen, Eli J; Shaytan, Alexey K; Mariño-Ramírez, Leonardo; Talbert, Paul B; Landsman, David; Panchenko, Anna R

    2016-01-01

    Compaction of DNA into chromatin is a characteristic feature of eukaryotic organisms. The core (H2A, H2B, H3, H4) and linker (H1) histone proteins are responsible for this compaction through the formation of nucleosomes and higher order chromatin aggregates. Moreover, histones are intricately involved in chromatin functioning and provide a means for genome dynamic regulation through specific histone variants and histone post-translational modifications. 'HistoneDB 2.0--with variants' is a comprehensive database of histone protein sequences, classified by histone types and variants. All entries in the database are supplemented by rich sequence and structural annotations with many interactive tools to explore and compare sequences of different variants from various organisms. The core of the database is a manually curated set of histone sequences grouped into 30 different variant subsets with variant-specific annotations. The curated set is supplemented by an automatically extracted set of histone sequences from the non-redundant protein database using algorithms trained on the curated set. The interactive web site supports various searching strategies in both datasets: browsing of phylogenetic trees; on-demand generation of multiple sequence alignments with feature annotations; classification of histone-like sequences and browsing of the taxonomic diversity for every histone variant. HistoneDB 2.0 is a resource for the interactive comparative analysis of histone protein sequences and their implications for chromatin function. Database URL: http://www.ncbi.nlm.nih.gov/projects/HistoneDB2.0.

  11. Analysis of histones and histone variants in plants.

    Science.gov (United States)

    Trivedi, Ila; Rai, Krishan Mohan; Singh, Sunil Kumar; Kumar, Verandra; Singh, Mala; Ranjan, Amol; Lodhi, Niraj; Sawant, Samir V

    2012-01-01

    Histone proteins are the major protein components of chromatin - the physiologically relevant form of the genome (or epigenome) in all eukaryotic cells. For many years, histones were considered passive structural components of eukaryotic chromatin. In recent years, it has been demonstrated that dynamic association of histones and their variants to the genome plays a very important role in gene regulation. Histones are extensively modified during posttranslation viz. acetylation, methylation, phosphorylation, ubiquitylation, etc., and the identification of these covalent marks on canonical and variant histones is crucial for the understanding of their biological significance. Different biochemical techniques have been developed to purify and separate histone proteins; here, we describe techniques for analysis of histones from plant tissues.

  12. Histone variants in plant transcriptional regulation.

    Science.gov (United States)

    Jiang, Danhua; Berger, Frédéric

    2017-01-01

    Chromatin based organization of eukaryotic genome plays a profound role in regulating gene transcription. Nucleosomes form the basic subunits of chromatin by packaging DNA with histone proteins, impeding the access of DNA to transcription factors and RNA polymerases. Exchange of histone variants in nucleosomes alters the properties of nucleosomes and thus modulates DNA exposure during transcriptional regulation. Growing evidence indicates the important function of histone variants in programming transcription during developmental transitions and stress response. Here we review how histone variants and their deposition machineries regulate the nucleosome stability and dynamics, and discuss the link between histone variants and transcriptional regulation in plants. This article is part of a Special Issue entitled: Plant Gene Regulatory Mechanisms and Networks, edited by Dr. Erich Grotewold and Dr. Nathan Springer.

  13. Histone variant innovation in a rapidly evolving chordate lineage

    Directory of Open Access Journals (Sweden)

    Jansen Pascal WTC

    2011-07-01

    Full Text Available Abstract Background Histone variants alter the composition of nucleosomes and play crucial roles in transcription, chromosome segregation, DNA repair, and sperm compaction. Modification of metazoan histone variant lineages occurs on a background of genome architecture that shows global similarities from sponges to vertebrates, but the urochordate, Oikopleura dioica, a member of the sister group to vertebrates, exhibits profound modification of this ancestral architecture. Results We show that a histone complement of 47 gene loci encodes 31 histone variants, grouped in distinct sets of developmental expression profiles throughout the life cycle. A particularly diverse array of 15 male-specific histone variants was uncovered, including a testes-specific H4t, the first metazoan H4 sequence variant reported. Universal histone variants H3.3, CenH3, and H2A.Z are present but O. dioica lacks homologs of macroH2A and H2AX. The genome encodes many H2A and H2B variants and the repertoire of H2A.Z isoforms is expanded through alternative splicing, incrementally regulating the number of acetylatable lysine residues in the functionally important N-terminal "charge patch". Mass spectrometry identified 40 acetylation, methylation and ubiquitylation posttranslational modifications (PTMs and showed that hallmark PTMs of "active" and "repressive" chromatin were present in O. dioica. No obvious reduction in silent heterochromatic marks was observed despite high gene density in this extraordinarily compacted chordate genome. Conclusions These results show that histone gene complements and their organization differ considerably even over modest phylogenetic distances. Substantial innovation among all core and linker histone variants has evolved in concert with adaptation of specific life history traits in this rapidly evolving chordate lineage.

  14. Canonical and variant histones of protozoan parasites.

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    Dalmasso, Maria Carolina; Sullivan, William Joseph; Angel, Sergio Oscar

    2011-06-01

    Protozoan parasites have tremendously diverse lifestyles that require adaptation to a remarkable assortment of different environmental conditions. In order to complete their life cycles, protozoan parasites rely on fine-tuning gene expression. In general, protozoa use novel regulatory elements, transcription factors, and epigenetic mechanisms to regulate their transcriptomes. One of the most surprising findings includes the nature of their histones--these primitive eukaryotes lack some histones yet harbor novel histone variants of unknown function. In this review, we describe the histone components of different protozoan parasites based on literature and database searching. We summarize the key discoveries regarding histones and histone variants and their impact on chromatin regulation in protozoan parasites. In addition, we list histone genes IDs, sequences, and genomic localization of several protozoan parasites and Microsporidia histones, obtained from a thorough search of genome databases. We then compare these findings with those observed in higher eukaryotes, allowing us to highlight some novel aspects of epigenetic regulation in protists and to propose questions to be addressed in the upcoming years.

  15. Histone variants and melanoma: facts and hypotheses.

    Science.gov (United States)

    Konstantinov, Nikifor K; Ulff-Møller, Constance J; Dimitrov, Stefan

    2016-07-01

    Melanoma is the most aggressive form of skin cancer with rising incidence and morbidity. Despite advances in treatment, the 10-yr survival for patients with metastatic disease is less than 10%. During the past few years, ongoing research on different epigenomic aberrations in melanoma has catalyzed better understanding of its pathogenesis and identification of new therapeutics. In our review, we will focus on the role of histone variants, key epigenetic players in melanoma initiation and progression. Specifically, incorporation of histone variants enables additional layers of chromatin structure, and here, we will describe how alterations in this epigenetic behavior impact melanoma.

  16. The human histone chaperone sNASP interacts with linker and core histones through distinct mechanisms.

    Science.gov (United States)

    Wang, Huanyu; Ge, Zhongqi; Walsh, Scott T R; Parthun, Mark R

    2012-01-01

    Somatic nuclear autoantigenic sperm protein (sNASP) is a human homolog of the N1/N2 family of histone chaperones. sNASP contains the domain structure characteristic of this family, which includes a large acidic patch flanked by several tetratricopeptide repeat (TPR) motifs. sNASP possesses a unique binding specificity in that it forms specific complexes with both histone H1 and histones H3/H4. Based on the binding affinities of sNASP variants to histones H1, H3.3, H4 and H3.3/H4 complexes, sNASP uses distinct structural domains to interact with linker and core histones. For example, one of the acidic patches of sNASP was essential for linker histone binding but not for core histone interactions. The fourth TPR of sNASP played a critical role in interactions with histone H3/H4 complexes, but did not influence histone H1 binding. Finally, analysis of cellular proteins demonstrated that sNASP existed in distinct complexes that contained either linker or core histones.

  17. Histone Variants in Development and Diseases

    Institute of Scientific and Technical Information of China (English)

    Ping Chen; Jicheng Zhao; Guohong Li

    2013-01-01

    Eukaryotic genomic DNA is highly packaged into chromatin by histones to fit inside the nucleus.Other than the bulk packaging role of canonical histones with an expression peak at S phase and replication-coupled deposition,different histone variants have evolved distinct regulatory mechanisms for their expression,deposition and functional implications.The diversity of histone variants results in structural plasticity of chromatin and highlights functionally distinct chromosomal domain,which plays critical roles in development from a fertilized egg into a complex organism,as well as in aging and diseases.However,the mechanisms of this fundamental process are poorly understood so far.It is of particular interest to investigate how the variants are incorporated into chromatin and mark specific chromatin states to regulate gene expression,and how they are involved in development and diseases.In this review,we focus on recent progress in studies of epigenetic regulation of three extensively investigated variants including H2A.Z,macroH2A and H3.3,and their functional implications in development and diseases.

  18. Global turnover of histone post-translational modifications and variants in human cells

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    Zee Barry M

    2010-12-01

    Full Text Available Abstract Background Post-translational modifications (PTMs on the N-terminal tails of histones and histone variants regulate distinct transcriptional states and nuclear events. Whereas the functional effects of specific PTMs are the current subject of intense investigation, most studies characterize histone PTMs/variants in a non-temporal fashion and very few studies have reported kinetic information about these histone forms. Previous studies have used radiolabeling, fluorescence microscopy and chromatin immunoprecipitation to determine rates of histone turnover, and have found interesting correlations between increased turnover and increased gene expression. Therefore, histone turnover is an understudied yet potentially important parameter that may contribute to epigenetic regulation. Understanding turnover in the context of histone modifications and sequence variants could provide valuable additional insight into the function of histone replacement. Results In this study, we measured the metabolic rate of labeled isotope incorporation into the histone proteins of HeLa cells by combining stable isotope labeling of amino acids in cell culture (SILAC pulse experiments with quantitative mass spectrometry-based proteomics. In general, we found that most core histones have similar turnover rates, with the exception of the H2A variants, which exhibit a wider range of rates, potentially consistent with their epigenetic function. In addition, acetylated histones have a significantly faster turnover compared with general histone protein and methylated histones, although these rates vary considerably, depending on the site and overall degree of methylation. Histones containing transcriptionally active marks have been consistently found to have faster turnover rates than histones containing silent marks. Interestingly, the presence of both active and silent marks on the same peptide resulted in a slower turnover rate than either mark alone on that same

  19. Phylogenomics of unusual histone H2A Variants in Bdelloid rotifers.

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    Karine Van Doninck

    2009-03-01

    Full Text Available Rotifers of Class Bdelloidea are remarkable in having evolved for millions of years, apparently without males and meiosis. In addition, they are unusually resistant to desiccation and ionizing radiation and are able to repair hundreds of radiation-induced DNA double-strand breaks per genome with little effect on viability or reproduction. Because specific histone H2A variants are involved in DSB repair and certain meiotic processes in other eukaryotes, we investigated the histone H2A genes and proteins of two bdelloid species. Genomic libraries were built and probed to identify histone H2A genes in Adineta vaga and Philodina roseola, species representing two different bdelloid families. The expressed H2A proteins were visualized on SDS-PAGE gels and identified by tandem mass spectrometry. We find that neither the core histone H2A, present in nearly all other eukaryotes, nor the H2AX variant, a ubiquitous component of the eukaryotic DSB repair machinery, are present in bdelloid rotifers. Instead, they are replaced by unusual histone H2A variants of higher mass. In contrast, a species of rotifer belonging to the facultatively sexual, desiccation- and radiation-intolerant sister class of bdelloid rotifers, the monogononts, contains a canonical core histone H2A and appears to lack the bdelloid H2A variant genes. Applying phylogenetic tools, we demonstrate that the bdelloid-specific H2A variants arose as distinct lineages from canonical H2A separate from those leading to the H2AX and H2AZ variants. The replacement of core H2A and H2AX in bdelloid rotifers by previously uncharacterized H2A variants with extended carboxy-terminal tails is further evidence for evolutionary diversity within this class of histone H2A genes and may represent adaptation to unusual features specific to bdelloid rotifers.

  20. The Histone Deacetylase Complex 1 Protein of Arabidopsis Has the Capacity to Interact with Multiple Proteins Including Histone 3-Binding Proteins and Histone 1 Variants.

    Science.gov (United States)

    Perrella, Giorgio; Carr, Craig; Asensi-Fabado, Maria A; Donald, Naomi A; Páldi, Katalin; Hannah, Matthew A; Amtmann, Anna

    2016-05-01

    Intrinsically disordered proteins can adopt multiple conformations, thereby enabling interaction with a wide variety of partners. They often serve as hubs in protein interaction networks. We have previously shown that the Histone Deacetylase Complex 1 (HDC1) protein from Arabidopsis (Arabidopsis thaliana) interacts with histone deacetylases and quantitatively determines histone acetylation levels, transcriptional activity, and several phenotypes, including abscisic acid sensitivity during germination, vegetative growth rate, and flowering time. HDC1-type proteins are ubiquitous in plants, but they contain no known structural or functional domains. Here, we explored the protein interaction spectrum of HDC1 using a quantitative bimolecular fluorescence complementation assay in tobacco (Nicotiana benthamiana) epidermal cells. In addition to binding histone deacetylases, HDC1 directly interacted with histone H3-binding proteins and corepressor-associated proteins but not with H3 or the corepressors themselves. Surprisingly, HDC1 also was able to interact with variants of the linker histone H1. Truncation of HDC1 to the ancestral core sequence narrowed the spectrum of interactions and of phenotypic outputs but maintained binding to a H3-binding protein and to H1. Thus, HDC1 provides a potential link between H1 and histone-modifying complexes.

  1. Dynamic deposition of histone variant H3.3 accompanies developmental remodeling of the Arabidopsis transcriptome.

    Directory of Open Access Journals (Sweden)

    Heike Wollmann

    Full Text Available In animals, replication-coupled histone H3.1 can be distinguished from replication-independent histone H3.3. H3.3 variants are enriched at active genes and their promoters. Furthermore, H3.3 is specifically incorporated upon gene activation. Histone H3 variants evolved independently in plants and animals, and it is unclear whether different replication-independent H3.3 variants developed similar properties in both phyla. We studied Arabidopsis H3 variants in order to find core properties of this class of histones. Here we present genome-wide maps of H3.3 and H3.1 enrichment and the dynamic changes of their profiles upon cell division arrest. We find H3.3 enrichment to positively correlate with gene expression and to be biased towards the transcription termination site. In contrast with H3.1, heterochromatic regions are mostly depleted of H3.3. We report that, in planta, dynamic changes in H3.3 profiles are associated with the extensive remodeling of the transcriptome that occurs during cell differentiation. We propose that H3.3 dynamics are linked to transcription and are involved in resetting covalent histone marks at a genomic scale during plant development. Our study suggests that H3 variants properties likely result from functionally convergent evolution.

  2. A unified phylogeny-based nomenclature for histone variants

    Directory of Open Access Journals (Sweden)

    Talbert Paul B

    2012-06-01

    Full Text Available Abstract Histone variants are non-allelic protein isoforms that play key roles in diversifying chromatin structure. The known number of such variants has greatly increased in recent years, but the lack of naming conventions for them has led to a variety of naming styles, multiple synonyms and misleading homographs that obscure variant relationships and complicate database searches. We propose here a unified nomenclature for variants of all five classes of histones that uses consistent but flexible naming conventions to produce names that are informative and readily searchable. The nomenclature builds on historical usage and incorporates phylogenetic relationships, which are strong predictors of structure and function. A key feature is the consistent use of punctuation to represent phylogenetic divergence, making explicit the relationships among variant subtypes that have previously been implicit or unclear. We recommend that by default new histone variants be named with organism-specific paralog-number suffixes that lack phylogenetic implication, while letter suffixes be reserved for structurally distinct clades of variants. For clarity and searchability, we encourage the use of descriptors that are separate from the phylogeny-based variant name to indicate developmental and other properties of variants that may be independent of structure.

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

    Institute of Scientific and Technical Information of China (English)

    Xiaofeng CAO

    2011-01-01

    @@ 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.

  4. Global dynamics of newly constructed oligonucleosomes of conventional and variant H2A.Z histone

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    Ioshikhes Ilya

    2007-11-01

    Full Text Available Abstract Background Complexes of nucleosomes, which often occur in the gene promoter areas, are one of the fundamental levels of chromatin organization and thus are important for transcription regulation. Investigating the dynamic structure of a single nucleosome as well as nucleosome complexes is important for understanding transcription within chromatin. In a previous work, we highlighted the influence of histone variants on the functional dynamics of a single nucleosome using normal mode analysis developed by Bahar et al. The present work further analyzes the dynamics of nucleosome complexes (nucleosome oligomers or oligonucleosomes such as dimer, trimer and tetramer (beads on a string model with conventional core histones as well as with the H2A.Z histone variant using normal mode analysis. Results The global dynamics of oligonucleosomes reveal larger amplitude of motion within the nucleosomes that contain the H2A.Z variant with in-planar and out-of-planar fluctuations as the common mode of relaxation. The docking region of H2A.Z and the L1:L1 interactions between H2A.Z monomers of nucleosome (that are responsible for the highly stable nucleosome containing variant H2A.Z-histone are highly dynamic throughout the first two dynamic modes. Conclusion Dissection of the dynamics of oligonucleosomes discloses in-plane as well as out-of-plane fluctuations as the common mode of relaxation throughout the global motions. The dynamics of individual nucleosomes and the combination of the relaxation mechanisms expressed by the individual nucleosome are quite interesting and highly dependent on the number of nucleosome fragments present in the complexes. Distortions generated by the non-planar dynamics influence the DNA conformation, and hence the histone-DNA interactions significantly alter the dynamics of the DNA. The variant H2A.Z histone is a major source of weaker intra- and inter-molecular correlations resulting in more disordered motions.

  5. CTCF induces histone variant incorporation, erases the H3K27me3 histone mark and opens chromatin

    NARCIS (Netherlands)

    O. Weth (Oliver); C. Paprotka (Christine); K. Günther (Katharina); A. Schulte (Astrid); M. Baierl (Manuel); J. Leers (Joerg); N.J. Galjart (Niels); R. Renkawitz (Rainer)

    2014-01-01

    textabstractInsulators functionally separate active chromatin domains frominactive ones. The insulator factor, CTCF, has been found to bind to boundaries and to mediate insulator function. CTCF binding sites are depleted for the histone modification H3K27me3 and are enriched for the histone variant

  6. Post-translational modifications of linker histone H1 variants in mammals

    Science.gov (United States)

    Starkova, T. Yu; Polyanichko, A. M.; Artamonova, T. O.; Khodorkovskii, M. A.; Kostyleva, E. I.; Chikhirzhina, E. V.; Tomilin, A. N.

    2017-02-01

    The covalent modifications of the linker histone H1 and the core histones are thought to play an important role in the control of chromatin functioning. Histone H1 variants from K562 cell line (hH1), mouse (mH1) and calf (cH1) thymi were studied by matrix-activated laser desorption/ionization fourier transform ion cyclotron resonance mass-spectroscopy (MALDI-FT-ICR-MS). The proteomics analysis revealed novel post-translational modifications of the histone H1, such as meK34-mH1.4, meK35-cH1.1, meK35-mH1.1, meK75-hH1.2, meK75-hH1.3, acK26-hH1.4, acK26-hH1.3 and acK17-hH1.1. The comparison of the hH1, mH1 and cH1 proteins has demonstrated that the types and positions of the post-translational modifications of the globular domains of the H1.2–H1.4 variants are very conservative. However, the post-translational modifications of the N- and C-terminal tails of H1.2, H1.3 and H1.4 are different. The differences of post-translational modifications in the N- and C-terminal tails of H1.2, H1.3 and H1.4 likely lead to the differences in DNA-H1 and H1-protein interactions.

  7. Macro histone variants are critical for the differentiation of human pluripotent cells.

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    Barrero, María J; Sese, Borja; Martí, Mercè; Izpisua Belmonte, Juan Carlos

    2013-05-31

    We have previously shown that macro histone variants (macroH2A) are expressed at low levels in stem cells and are up-regulated during differentiation. Here we show that the knockdown of macro histone variants impaired the in vitro and in vivo differentiation of human pluripotent cells, likely through defects in the silencing of pluripotency-related genes. ChIP experiments showed that during differentiation macro histone variants are recruited to the regulatory regions of pluripotency and developmental genes marked with H3K27me3 contributing to the silencing of these genes.

  8. Structural insights into yeast histone chaperone Hif1: a scaffold protein recruiting protein complexes to core histones.

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    Liu, Hejun; Zhang, Mengying; He, Wei; Zhu, Zhongliang; Teng, Maikun; Gao, Yongxiang; Niu, Liwen

    2014-09-15

    Yeast Hif1 [Hat1 (histone acetyltransferase 1)-interacting factor], a homologue of human NASP (nuclear autoantigenic sperm protein), is a histone chaperone that is involved in various protein complexes which modify histones during telomeric silencing and chromatin reassembly. For elucidating the structural basis of Hif1, in the present paper we demonstrate the crystal structure of Hif1 consisting of a superhelixed TPR (tetratricopeptide repeat) domain and an extended acid loop covering the rear of TPR domain, which represent typical characteristics of SHNi-TPR [Sim3 (start independent of mitosis 3)-Hif1-NASP interrupted TPR] proteins. Our binding assay indicates that Hif1 could bind to the histone octamer via histones H3 and H4. The acid loop is shown to be crucial for the binding of histones and may also change the conformation of the TPR groove. By binding to the core histone complex Hif1 may recruit functional protein complexes to modify histones during chromatin reassembly.

  9. Novel nucleosomal particles containing core histones and linker DNA but no histone H1.

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    Cole, Hope A; Cui, Feng; Ocampo, Josefina; Burke, Tara L; Nikitina, Tatiana; Nagarajavel, V; Kotomura, Naoe; Zhurkin, Victor B; Clark, David J

    2016-01-29

    Eukaryotic chromosomal DNA is assembled into regularly spaced nucleosomes, which play a central role in gene regulation by determining accessibility of control regions. The nucleosome contains ∼147 bp of DNA wrapped ∼1.7 times around a central core histone octamer. The linker histone, H1, binds both to the nucleosome, sealing the DNA coils, and to the linker DNA between nucleosomes, directing chromatin folding. Micrococcal nuclease (MNase) digests the linker to yield the chromatosome, containing H1 and ∼160 bp, and then converts it to a core particle, containing ∼147 bp and no H1. Sequencing of nucleosomal DNA obtained after MNase digestion (MNase-seq) generates genome-wide nucleosome maps that are important for understanding gene regulation. We present an improved MNase-seq method involving simultaneous digestion with exonuclease III, which removes linker DNA. Remarkably, we discovered two novel intermediate particles containing 154 or 161 bp, corresponding to 7 bp protruding from one or both sides of the nucleosome core. These particles are detected in yeast lacking H1 and in H1-depleted mouse chromatin. They can be reconstituted in vitro using purified core histones and DNA. We propose that these 'proto-chromatosomes' are fundamental chromatin subunits, which include the H1 binding site and influence nucleosome spacing independently of H1.

  10. Dynamic expression of combinatorial replication-dependent histone variant genes during mouse spermatogenesis.

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    Sun, Rongfang; Qi, Huayu

    2014-01-01

    Nucleosomes are basic chromatin structural units that are formed by DNA sequences wrapping around histones. Global chromatin states in different cell types are specified by combinatorial effects of post-translational modifications of histones and the expression of histone variants. During mouse spermatogenesis, spermatogonial stem cells (SSCs) self-renew while undergo differentiation, events that occur in the company of constant re-modeling of chromatin structures. Previous studies have shown that testes contain highly expressed or specific histone variants to facilitate these epigenetic modifications. However, mechanisms of regulating the epigenetic changes and the specific histone compositions of spermatogenic cells are not fully understood. Using real time quantitative RT-PCR, we examined the dynamic expression of replication-dependent histone genes in post-natal mouse testes. It was found that distinct sets of histone genes are expressed in various spermatogenic cells at different stages during spermatogenesis. While gonocyte-enriched testes from mice at 2-dpp (days post partum) express pre-dominantly thirteen histone variant genes, SSC-stage testes at 9-dpp highly express a different set of eight histone genes. During differentiation stage when testes are occupied mostly by spermatocytes and spermatids, another twenty-two histone genes are expressed much higher than the rest, including previously known testis-specific hist1h1t, hist1h2ba and hist1h4c. In addition, histone genes that are pre-dominantly expressed in gonocytes and SSCs are also highly expressed in embryonic stem cells. Several of them were changed when embryoid bodies were formed from ES cells, suggesting their roles in regulating pluripotency of the cells. Further more, differentially expressed histone genes are specifically localized in either SSCs or spermatocytes and spermatids, as demonstrated by in situ hybridization using gene specific probes. Taken together, results presented here

  11. Genome-Wide Identification, Evolutionary, and Expression Analyses of Histone H3 Variants in Plants

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    Jinteng Cui

    2015-01-01

    Full Text Available Histone variants alter the nucleosome structure and play important roles in chromosome segregation, transcription, DNA repair, and sperm compaction. Histone H3 is encoded by many genes in most eukaryotic species and is the histone that contains the largest variety of posttranslational modifications. Compared with the metazoan H3 variants, little is known about the complex evolutionary history of H3 variants proteins in plants. Here, we study the identification, evolutionary, and expression analyses of histone H3 variants from genomes in major branches in the plant tree of life. Firstly we identified all the histone three related (HTR genes from the examined genomes, then we classified the four groups variants: centromeric H3, H3.1, H3.3 and H3-like, by phylogenetic analysis, intron information, and alignment. We further demonstrated that the H3 variants have evolved under strong purifying selection, indicating the conservation of HTR proteins. Expression analysis revealed that the HTR has a wide expression profile in maize and rice development and plays important roles in development.

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

    Science.gov (United States)

    Mariño-Ramírez, Leonardo; Levine, Kevin M; Morales, Mario; Zhang, Suiyuan; Moreland, R Travis; Baxevanis, Andreas D; Landsman, David

    2011-01-01

    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.

  13. Protein Profiling and Histone Deacetylation Activities in Somaclonal Variants of Oil Palm (Elaeis guineensis Jacq.

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    Jamilah Syafawati Yaacob

    2013-01-01

    Full Text Available Mantled fruits as a result of somaclonal variation are often observed from the oil palm plantlets regenerated via tissue culture. The mantling of fruits with finger-like and thick outer coating phenotypes significantly reduces the seed size and oil content, posing a threat to oil palm planters, and may jeopardize the economic growth of countries that depend particularly on oil palm plantation. The molecular aspects of the occurrence of somaclonal variations are yet to be known, possibly due to gene repression such as DNA methylation, histone methylation and histone deacetylation. Histone deacetylases (HDACs, involved in eukaryotic gene regulation by catalyzing the acetyl groups are removal from lysine residues on histone, hence transcriptionally repress gene expression. This paper described the total protein polymorphism profiles of somaclonal variants of oil palm and the effects of histone deacetylation on this phenomenon. Parallel to the different phenotypes, the protein polymorphism profiles of the mantled samples (leaves, fruits, and florets and the phenotypically normal samples were proven to be different. Higher HDAC activity was found in mantled leaf samples than in the phenotypically normal leaf samples, leading to a preliminary conclusion that histone deacetylation suppressed gene expression and contributed to the development of somaclonal variants.

  14. Protein profiling and histone deacetylation activities in somaclonal variants of oil palm (Elaeis guineensis Jacq.).

    Science.gov (United States)

    Yaacob, Jamilah Syafawati; Loh, Hwei-San; Mat Taha, Rosna

    2013-01-01

    Mantled fruits as a result of somaclonal variation are often observed from the oil palm plantlets regenerated via tissue culture. The mantling of fruits with finger-like and thick outer coating phenotypes significantly reduces the seed size and oil content, posing a threat to oil palm planters, and may jeopardize the economic growth of countries that depend particularly on oil palm plantation. The molecular aspects of the occurrence of somaclonal variations are yet to be known, possibly due to gene repression such as DNA methylation, histone methylation and histone deacetylation. Histone deacetylases (HDACs), involved in eukaryotic gene regulation by catalyzing the acetyl groups are removal from lysine residues on histone, hence transcriptionally repress gene expression. This paper described the total protein polymorphism profiles of somaclonal variants of oil palm and the effects of histone deacetylation on this phenomenon. Parallel to the different phenotypes, the protein polymorphism profiles of the mantled samples (leaves, fruits, and florets) and the phenotypically normal samples were proven to be different. Higher HDAC activity was found in mantled leaf samples than in the phenotypically normal leaf samples, leading to a preliminary conclusion that histone deacetylation suppressed gene expression and contributed to the development of somaclonal variants.

  15. The role of variant histone H2AV in Drosophila melanogaster larval hematopoiesis.

    Science.gov (United States)

    Grigorian, Melina; DeBruhl, Heather; Lipsick, Joseph S

    2017-04-15

    Replication-independent histone variants can replace the canonical replication-dependent histones. Vertebrates have multiple H2A variant histones, including H2AZ and H2AX that are present in most eukaryotes. H2AZ regulates transcriptional activation as well as the maintenance of gene silencing, while H2AX is important in DNA damage repair. The fruit fly Drosophila melanogaster has only one histone H2A variant (H2AV), which is a chimera of H2AZ and H2AX. In this study we found that lack of H2AV led to the formation of black melanotic masses in Drosophila third instar larvae. The formation of these masses was found in conjunction with a loss of the majority of the primary lymph gland lobes. Interestingly, the cells of the posterior signaling center were preserved in these mutants. Reduction of H2AV levels by RNAi knockdown caused a milder phenotype that preserved the lymph gland structure but that included precocious differentiation of the prohemocytes located within the medullary zone and the secondary lobes of the lymph gland. Mutant rescue experiments suggest that the H2AZ-like rather than the H2AX-like function of H2AV is primarily required for normal hematopoiesis. © 2017. Published by The Company of Biologists Ltd.

  16. Weaver Syndrome‐Associated EZH2 Protein Variants Show Impaired Histone Methyltransferase Function In Vitro

    Science.gov (United States)

    Yap, Damian B.; Lewis, M.E. Suzanne; Chijiwa, Chieko; Ramos‐Arroyo, Maria A.; Tkachenko, Natália; Milano, Valentina; Fradin, Mélanie; McKinnon, Margaret L.; Townsend, Katelin N.; Xu, Jieqing; Van Allen, M.I.; Ross, Colin J.D.; Dobyns, William B.; Weaver, David D.; Gibson, William T.

    2016-01-01

    ABSTRACT Weaver syndrome (WS) is a rare congenital disorder characterized by generalized overgrowth, macrocephaly, specific facial features, accelerated bone age, intellectual disability, and susceptibility to cancers. De novo mutations in the enhancer of zeste homolog 2 (EZH2) have been shown to cause WS. EZH2 is a histone methyltransferase that acts as the catalytic agent of the polycomb‐repressive complex 2 (PRC2) to maintain gene repression via methylation of lysine 27 on histone H3 (H3K27). Functional studies investigating histone methyltransferase activity of mutant EZH2 from various cancers have been reported, whereas WS‐associated mutations remain poorly characterized. To investigate the role of EZH2 in WS, we performed functional studies using artificially assembled PRC2 complexes containing mutagenized human EZH2 that reflected the codon changes predicted from patients with WS. We found that WS‐associated amino acid alterations reduce the histone methyltransferase function of EZH2 in this in vitro assay. Our results support the hypothesis that WS is caused by constitutional mutations in EZH2 that alter the histone methyltransferase function of PRC2. However, histone methyltransferase activities of different EZH2 variants do not appear to correlate directly with the phenotypic variability between WS patients and individuals with a common c.553G>C (p.Asp185His) polymorphism in EZH2. PMID:26694085

  17. Classification of Promoters Based on the Combination of Core Promoter Elements Exhibits Different Histone Modification Patterns

    Science.gov (United States)

    Natsume-Kitatani, Yayoi; Mamitsuka, Hiroshi

    2016-01-01

    Four different histones (H2A, H2B, H3, and H4; two subunits each) constitute a histone octamer, around which DNA wraps to form histone-DNA complexes called nucleosomes. Amino acid residues in each histone are occasionally modified, resulting in several biological effects, including differential regulation of transcription. Core promoters that encompass the transcription start site have well-conserved DNA motifs, including the initiator (Inr), TATA box, and DPE, which are collectively called the core promoter elements (CPEs). In this study, we systematically studied the associations between the CPEs and histone modifications by integrating the Drosophila Core Promoter Database and time-series ChIP-seq data for histone modifications (H3K4me3, H3K27ac, and H3K27me3) during development in Drosophila melanogaster via the modENCODE project. We classified 96 core promoters into four groups based on the presence or absence of the TATA box or DPE, calculated the histone modification ratio at the core promoter region, and transcribed region for each core promoter. We found that the histone modifications in TATA-less groups were static during development and that the core promoters could be clearly divided into three types: i) core promoters with continuous active marks (H3K4me3 and H3K27ac), ii) core promoters with a continuous inactive mark (H3K27me3) and occasional active marks, and iii) core promoters with occasional histone modifications. Linear regression analysis and non-linear regression by random forest showed that the TATA-containing groups included core promoters without histone modifications, for which the measured RNA expression values were not predictable accurately from the histone modification status. DPE-containing groups had a higher relative frequency of H3K27me3 in both the core promoter region and transcribed region. In summary, our analysis showed that there was a systematic link between the existence of the CPEs and the dynamics, frequency and influence

  18. Classification of Promoters Based on the Combination of Core Promoter Elements Exhibits Different Histone Modification Patterns.

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    Yayoi Natsume-Kitatani

    Full Text Available Four different histones (H2A, H2B, H3, and H4; two subunits each constitute a histone octamer, around which DNA wraps to form histone-DNA complexes called nucleosomes. Amino acid residues in each histone are occasionally modified, resulting in several biological effects, including differential regulation of transcription. Core promoters that encompass the transcription start site have well-conserved DNA motifs, including the initiator (Inr, TATA box, and DPE, which are collectively called the core promoter elements (CPEs. In this study, we systematically studied the associations between the CPEs and histone modifications by integrating the Drosophila Core Promoter Database and time-series ChIP-seq data for histone modifications (H3K4me3, H3K27ac, and H3K27me3 during development in Drosophila melanogaster via the modENCODE project. We classified 96 core promoters into four groups based on the presence or absence of the TATA box or DPE, calculated the histone modification ratio at the core promoter region, and transcribed region for each core promoter. We found that the histone modifications in TATA-less groups were static during development and that the core promoters could be clearly divided into three types: i core promoters with continuous active marks (H3K4me3 and H3K27ac, ii core promoters with a continuous inactive mark (H3K27me3 and occasional active marks, and iii core promoters with occasional histone modifications. Linear regression analysis and non-linear regression by random forest showed that the TATA-containing groups included core promoters without histone modifications, for which the measured RNA expression values were not predictable accurately from the histone modification status. DPE-containing groups had a higher relative frequency of H3K27me3 in both the core promoter region and transcribed region. In summary, our analysis showed that there was a systematic link between the existence of the CPEs and the dynamics, frequency

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

    OpenAIRE

    Mariño-Ramírez, Leonardo; Levine, Kevin M.; Morales, Mario; Zhang, Suiyuan; Moreland, R. Travis; Baxevanis, Andreas D; Landsman, David

    2011-01-01

    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 Hi...

  20. Dengue Virus Capsid Protein Binds Core Histones and Inhibits Nucleosome Formation in Human Liver Cells

    Science.gov (United States)

    Colpitts, Tonya M.; Barthel, Sebastian; Wang, Penghua; Fikrig, Erol

    2011-01-01

    Dengue virus (DENV) is a member of the Flaviviridae and a globally (re)emerging pathogen that causes serious human disease. There is no specific antiviral or vaccine for dengue virus infection. Flavivirus capsid (C) is a structural protein responsible for gathering the viral RNA into a nucleocapsid that forms the core of a mature virus particle. Flaviviral replication is known to occur in the cytoplasm yet a large portion of capsid protein localizes to the nucleus during infection. The reasons for the nuclear presences of capsid are not completely understood. Here, we expressed mature DENV C in a tandem affinity purification assay to identify potential binding partners in human liver cells. DENV C targeted the four core histones, H2A, H2B, H3 and H4. DENV C bound recombinant histones in solution and colocalized with histones in the nucleus and cytoplasm of liver cells during DENV infection. We show that DENV C acts as a histone mimic, forming heterodimers with core histones, binding DNA and disrupting nucleosome formation. We also demonstrate that DENV infection increases the amounts of core histones in livers cells, which may be a cellular response to C binding away the histone proteins. Infection with DENV additionally alters levels of H2A phosphorylation in a time-dependent manner. The interactions of C and histones add an interesting new role for the presence of C in the nucleus during DENV infection. PMID:21909430

  1. Dengue virus capsid protein binds core histones and inhibits nucleosome formation in human liver cells.

    Directory of Open Access Journals (Sweden)

    Tonya M Colpitts

    Full Text Available Dengue virus (DENV is a member of the Flaviviridae and a globally (reemerging pathogen that causes serious human disease. There is no specific antiviral or vaccine for dengue virus infection. Flavivirus capsid (C is a structural protein responsible for gathering the viral RNA into a nucleocapsid that forms the core of a mature virus particle. Flaviviral replication is known to occur in the cytoplasm yet a large portion of capsid protein localizes to the nucleus during infection. The reasons for the nuclear presences of capsid are not completely understood. Here, we expressed mature DENV C in a tandem affinity purification assay to identify potential binding partners in human liver cells. DENV C targeted the four core histones, H2A, H2B, H3 and H4. DENV C bound recombinant histones in solution and colocalized with histones in the nucleus and cytoplasm of liver cells during DENV infection. We show that DENV C acts as a histone mimic, forming heterodimers with core histones, binding DNA and disrupting nucleosome formation. We also demonstrate that DENV infection increases the amounts of core histones in livers cells, which may be a cellular response to C binding away the histone proteins. Infection with DENV additionally alters levels of H2A phosphorylation in a time-dependent manner. The interactions of C and histones add an interesting new role for the presence of C in the nucleus during DENV infection.

  2. Post-translational Modifications of Trypanosoma cruzi Canonical and Variant Histones.

    Science.gov (United States)

    Picchi, Gisele F A; Zulkievicz, Vanessa; Krieger, Marco A; Zanchin, Nilson T; Goldenberg, Samuel; de Godoy, Lyris M F

    2017-03-03

    Chagas disease, caused by Trypanosoma cruzi, still affects millions of people around the world. No vaccines nor treatment for chronic Chagas disease are available, and chemotherapy for the acute phase is hindered by limited efficacy and severe side effects. The processes by which the parasite acquires infectivity and survives in different hosts involve tight regulation of gene expression, mainly post-transcriptionally. Nevertheless, chromatin structure/organization of trypanosomatids is similar to other eukaryotes, including histone variants and post-translational modifications. Emerging evidence suggests that epigenetic mechanisms also play an important role in the biology/pathogenesis of these parasites, making epigenetic targets suitable candidates to drug discovery. Here, we present the first comprehensive map of post-translational modifications of T. cruzi canonical and variant histones and show that its histone code can be as sophisticated as that of other eukaryotes. A total of 13 distinct modification types were identified, including rather novel and unusual ones such as alternative lysine acylations, serine/threonine acetylation, and N-terminal methylation. Some histone marks correlate to those described for other organisms, suggesting that similar regulatory mechanisms may be in place. Others, however, are unique to T. cruzi or to trypanosomatids as a group and might represent good candidates for the development of antiparasitic drugs.

  3. Core histone genes of Giardia intestinalis: genomic organization, promoter structure, and expression

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    Adam Rodney D

    2007-04-01

    Full Text Available Abstract Background Giardia intestinalis is a protist found in freshwaters worldwide, and is the most common cause of parasitic diarrhea in humans. The phylogenetic position of this parasite is still much debated. Histones are small, highly conserved proteins that associate tightly with DNA to form chromatin within the nucleus. There are two classes of core histone genes in higher eukaryotes: DNA replication-independent histones and DNA replication-dependent ones. Results We identified two copies each of the core histone H2a, H2b and H3 genes, and three copies of the H4 gene, at separate locations on chromosomes 3, 4 and 5 within the genome of Giardia intestinalis, but no gene encoding a H1 linker histone could be recognized. The copies of each gene share extensive DNA sequence identities throughout their coding and 5' noncoding regions, which suggests these copies have arisen from relatively recent gene duplications or gene conversions. The transcription start sites are at triplet A sequences 1–27 nucleotides upstream of the translation start codon for each gene. We determined that a 50 bp region upstream from the start of the histone H4 coding region is the minimal promoter, and a highly conserved 15 bp sequence called the histone motif (him is essential for its activity. The Giardia core histone genes are constitutively expressed at approximately equivalent levels and their mRNAs are polyadenylated. Competition gel-shift experiments suggest that a factor within the protein complex that binds him may also be a part of the protein complexes that bind other promoter elements described previously in Giardia. Conclusion In contrast to other eukaryotes, the Giardia genome has only a single class of core histone genes that encode replication-independent histones. Our inability to locate a gene encoding the linker histone H1 leads us to speculate that the H1 protein may not be required for the compaction of Giardia's small and gene-rich genome.

  4. The DNA intercalators ethidium bromide and propidium iodide also bind to core histones

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    Amrita Banerjee

    2014-01-01

    Full Text Available Eukaryotic DNA is compacted in the form of chromatin, in a complex with histones and other non-histone proteins. The intimate association of DNA and histones in chromatin raises the possibility that DNA-interactive small molecules may bind to chromatin-associated proteins such as histones. Employing biophysical and biochemical techniques we have characterized the interaction of a classical intercalator, ethidium bromide (EB and its structural analogue propidium iodide (PI with hierarchical genomic components: long chromatin, chromatosome, core octamer and chromosomal DNA. Our studies show that EB and PI affect both chromatin structure and function, inducing chromatin compaction and disruption of the integrity of the chromatosome. Calorimetric studies and fluorescence measurements of the ligands demonstrated and characterized the association of these ligands with core histones and the intact octamer in absence of DNA. The ligands affect acetylation of histone H3 at lysine 9 and acetylation of histone H4 at lysine 5 and lysine 8 ex vivo. PI alters the post-translational modifications to a greater extent than EB. This is the first report showing the dual binding (chromosomal DNA and core histones property of a classical intercalator, EB, and its longer analogue, PI, in the context of chromatin.

  5. Charge State of the Globular Histone Core Controls Stability of the Nucleosome

    Science.gov (United States)

    Fenley, Andrew T.; Adams, David A.; Onufriev, Alexey V.

    2010-01-01

    Presented here is a quantitative model of the wrapping and unwrapping of the DNA around the histone core of the nucleosome that suggests a mechanism by which this transition can be controlled: alteration of the charge state of the globular histone core. The mechanism is relevant to several classes of posttranslational modifications such as histone acetylation and phosphorylation; several specific scenarios consistent with recent in vivo experiments are considered. The model integrates a description based on an idealized geometry with one based on the atomistic structure of the nucleosome, and the model consistently accounts for both the electrostatic and nonelectrostatic contributions to the nucleosome free energy. Under physiological conditions, isolated nucleosomes are predicted to be very stable (38 ± 7 kcal/mol). However, a decrease in the charge of the globular histone core by one unit charge, for example due to acetylation of a single lysine residue, can lead to a significant decrease in the strength of association with its DNA. In contrast to the globular histone core, comparable changes in the charge state of the histone tail regions have relatively little effect on the nucleosome's stability. The combination of high stability and sensitivity explains how the nucleosome is able to satisfy the seemingly contradictory requirements for thermodynamic stability while allowing quick access to its DNA informational content when needed by specific cellular processes such as transcription. PMID:20816070

  6. Structure of human nucleosome containing the testis-specific histone variant TSH2B

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    Urahama, Takashi; Horikoshi, Naoki; Osakabe, Akihisa; Tachiwana, Hiroaki; Kurumizaka, Hitoshi, E-mail: kurumizaka@waseda.jp [Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480 (Japan)

    2014-03-25

    The crystal structure of human nucleosome containing the testis-specific TSH2B variant has been determined. The TSH2B Ser85 residue does not interact with H4 in the nucleosome, and induces a local structural difference between TSH2B and H2B in nucleosomes. The human histone H2B variant TSH2B is highly expressed in testis and may function in the chromatin transition during spermatogenesis. In the present study, the crystal structure of the human testis-specific nucleosome containing TSH2B was determined at 2.8 Å resolution. A local structural difference between TSH2B and canonical H2B in nucleosomes was detected around the TSH2B-specific amino-acid residue Ser85. The TSH2B Ser85 residue does not interact with H4 in the nucleosome, but in the canonical nucleosome the H2B Asn84 residue (corresponding to the TSH2B Ser85 residue) forms water-mediated hydrogen bonds with the H4 Arg78 residue. In contrast, the other TSH2B-specific amino-acid residues did not induce any significant local structural changes in the TSH2B nucleosome. These findings may provide important information for understanding how testis-specific histone variants form nucleosomes during spermatogenesis.

  7. Interaction of mitoxantrone, as an anticancer drug, with chromatin proteins, core histones and H1, in solution.

    Science.gov (United States)

    Hajihassan, Zahra; Rabbani-Chadegani, Azra

    2011-01-01

    In the present study, for the first time we have investigated the interaction of anticancer drug mitoxantrone with histone H1 and core histone proteins in solution using fluorescence, UV/Vis, CD spectroscopy and thermal denaturation techniques. The results showed that mitoxantrone reduced the absorbencies of H1 and core histone proteins at 210 nm (hypochromicity) and fluorescence emission intensity was decreased in a dose dependent. Binding of mitoxantrone changed secondary structures of the proteins as circular dichroism analysis confirmed it. Also, mitoxantrone increased the melting temperature of core histones at the final step of denaturation. The results suggest higher affinity of mitoxantrone to histone H1 compared to core histones providing histone proteins as a new target for mitoxantrone action at the chromatin level. Copyright © 2010 Elsevier B.V. All rights reserved.

  8. The histone variant H2A.Bbd is enriched at sites of DNA synthesis.

    Science.gov (United States)

    Sansoni, Viola; Casas-Delucchi, Corella S; Rajan, Malini; Schmidt, Andreas; Bönisch, Clemens; Thomae, Andreas W; Staege, Martin S; Hake, Sandra B; Cardoso, M Cristina; Imhof, Axel

    2014-06-01

    Histone variants play an important role in shaping the mammalian epigenome and their aberrant expression is frequently observed in several types of cancer. However, the mechanisms that mediate their function and the composition of the variant-containing chromatin are still largely unknown. A proteomic interrogation of chromatin containing the different H2A variants macroH2A.1.2, H2A.Bbd and H2A revealed a strikingly different protein composition. Gene ontology analysis reveals a strong enrichment of splicing factors as well as components of the mammalian replisome in H2A.Bbd-containing chromatin. We find H2A.Bbd localizing transiently to sites of DNA synthesis during S-phase and during DNA repair. Cells that express H2A.Bbd have a shortened S-phase and are more susceptible to DNA damage, two phenotypes that are also observed in human Hodgkin's lymphoma cells that aberrantly express this variant. Based on our experiments we conclude that H2A.Bbd is targeted to newly synthesized DNA during replication and DNA repair. The transient incorporation of H2A.Bbd may be due to the intrinsic instability of nucleosomes carrying this variant or a faster chromatin loading. This potentially leads to a disturbance of the existing chromatin structure, which may have effects on cell cycle regulation and DNA damage sensitivity.

  9. Acetylation and methylation patterns of core histones are modified after heat or arsenite treatment of Drosophila tissue culture cells

    OpenAIRE

    Arrigo, André-Patrick

    1983-01-01

    Exposure of Drosophilamelanogaster tissue culture cells to 37°C (heat shock) or to arsenite induces a severe deacetylation of core histones and blocks the methylation of histone H3 and H4. Heat shock induces the methylation of histone H2b. These results are discussed in view of chromatin structure and function.

  10. Testis-Specific Histone Variant H3t Gene Is Essential for Entry into Spermatogenesis

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    Jun Ueda

    2017-01-01

    Full Text Available Cellular differentiation is associated with dynamic chromatin remodeling in establishing a cell-type-specific epigenomic landscape. Here, we find that mouse testis-specific and replication-dependent histone H3 variant H3t is essential for very early stages of spermatogenesis. H3t gene deficiency leads to azoospermia because of the loss of haploid germ cells. When differentiating spermatogonia emerge in normal spermatogenesis, H3t appears and replaces the canonical H3 proteins. Structural and biochemical analyses reveal that H3t-containing nucleosomes are more flexible than the canonical nucleosomes. Thus, by incorporating H3t into the genome during spermatogonial differentiation, male germ cells are able to enter meiosis and beyond.

  11. The genomic distribution and function of histone variant HTZ-1 during C. elegans embryogenesis.

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    Christina M Whittle

    Full Text Available In all eukaryotes, histone variants are incorporated into a subset of nucleosomes to create functionally specialized regions of chromatin. One such variant, H2A.Z, replaces histone H2A and is required for development and viability in all animals tested to date. However, the function of H2A.Z in development remains unclear. Here, we use ChIP-chip, genetic mutation, RNAi, and immunofluorescence microscopy to interrogate the function of H2A.Z (HTZ-1 during embryogenesis in Caenorhabditis elegans, a key model of metazoan development. We find that HTZ-1 is expressed in every cell of the developing embryo and is essential for normal development. The sites of HTZ-1 incorporation during embryogenesis reveal a genome wrought by developmental processes. HTZ-1 is incorporated upstream of 23% of C. elegans genes. While these genes tend to be required for development and occupied by RNA polymerase II, HTZ-1 incorporation does not specify a stereotypic transcription program. The data also provide evidence for unexpectedly widespread independent regulation of genes within operons during development; in 37% of operons, HTZ-1 is incorporated upstream of internally encoded genes. Fewer sites of HTZ-1 incorporation occur on the X chromosome relative to autosomes, which our data suggest is due to a paucity of developmentally important genes on X, rather than a direct function for HTZ-1 in dosage compensation. Our experiments indicate that HTZ-1 functions in establishing or maintaining an essential chromatin state at promoters regulated dynamically during C. elegans embryogenesis.

  12. Characterization of centromeric histone H3 (CENH3 variants in cultivated and wild carrots (Daucus sp..

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    Frank Dunemann

    Full Text Available In eukaryotes, centromeres are the assembly sites for the kinetochore, a multi-protein complex to which spindle microtubules are attached at mitosis and meiosis, thereby ensuring segregation of chromosomes during cell division. They are specified by incorporation of CENH3, a centromere specific histone H3 variant which replaces canonical histone H3 in the nucleosomes of functional centromeres. To lay a first foundation of a putative alternative haploidization strategy based on centromere-mediated genome elimination in cultivated carrots, in the presented research we aimed at the identification and cloning of functional CENH3 genes in Daucus carota and three distantly related wild species of genus Daucus varying in basic chromosome numbers. Based on mining the carrot transcriptome followed by a subsequent PCR-based cloning, homologous coding sequences for CENH3s of the four Daucus species were identified. The ORFs of the CENH3 variants were very similar, and an amino acid sequence length of 146 aa was found in three out of the four species. Comparison of Daucus CENH3 amino acid sequences with those of other plant CENH3s as well as their phylogenetic arrangement among other dicot CENH3s suggest that the identified genes are authentic CENH3 homologs. To verify the location of the CENH3 protein in the kinetochore regions of the Daucus chromosomes, a polyclonal antibody based on a peptide corresponding to the N-terminus of DcCENH3 was developed and used for anti-CENH3 immunostaining of mitotic root cells. The chromosomal location of CENH3 proteins in the centromere regions of the chromosomes could be confirmed. For genetic localization of the CENH3 gene in the carrot genome, a previously constructed linkage map for carrot was used for mapping a CENH3-specific simple sequence repeat (SSR marker, and the CENH3 locus was mapped on the carrot chromosome 9.

  13. Linker histone variant H1T targets rDNA repeats.

    Science.gov (United States)

    Tani, Ruiko; Hayakawa, Koji; Tanaka, Satoshi; Shiota, Kunio

    2016-04-02

    H1T is a linker histone H1 variant that is highly expressed at the primary spermatocyte stage through to the early spermatid stage of spermatogenesis. While the functions of the somatic types of H1 have been extensively investigated, the intracellular role of H1T is unclear. H1 variants specifically expressed in germ cells show low amino acid sequence homology to somatic H1s, which suggests that the functions or target loci of germ cell-specific H1T differ from those of somatic H1s. Here, we describe the target loci and function of H1T. H1T was expressed not only in the testis but also in tumor cell lines, mouse embryonic stem cells (mESCs), and some normal somatic cells. To elucidate the intracellular localization and target loci of H1T, fluorescent immunostaining and ChIP-seq were performed in tumor cells and mESCs. We found that H1T accumulated in nucleoli and predominantly targeted rDNA repeats, which differ from somatic H1 targets. Furthermore, by nuclease sensitivity assay and RT-qPCR, we showed that H1T repressed rDNA transcription by condensing chromatin structure. Imaging analysis indicated that H1T expression affected nucleolar formation. We concluded that H1T plays a role in rDNA transcription, by distinctively targeting rDNA repeats.

  14. Exome-based Variant Detection in Core Promoters.

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    Kim, Yeong C; Cui, Jian; Luo, Jiangtao; Xiao, Fengxia; Downs, Bradley; Wang, San Ming

    2016-07-28

    Core promoter controls the initiation of transcription. Core promoter sequence change can disrupt transcriptional regulation, lead to impairment of gene expression and ultimately diseases. Therefore, comprehensive characterization of core promoters is essential to understand normal and abnormal gene expression in biomedical studies. Here we report the development of EVDC (Exome-based Variant Detection in Core promoters) method for genome-scale analysis of core-promoter sequence variation. This method is based on the fact that exome sequences contain the sequences not only from coding exons but also from non-coding region including core promoters generated by random fragmentation in exome sequencing process. Using exome data from three cell types of CD4+ T cells, CD19+ B cells and neutrophils of a single individual, we characterized the features of core promoter-mapped exome sequences, and analysed core-promoter variation in this individual genome. We also compared the core promoters between YRI (Yoruba in Ibadan, Nigeria) and the CEU (Utah residents of European decedent) populations using the exome data generated by the 1000 Genome project, and observed much higher variation in YRI population than in CEU population. Our study demonstrates that the EVDC method provides a simple but powerful means for genome-wile de novo characterization of core promoter sequence variation.

  15. Esperanto for histones: CENP-A, not CenH3, is the centromeric histone H3 variant.

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    Earnshaw, W C; Allshire, R C; Black, B E; Bloom, K; Brinkley, B R; Brown, W; Cheeseman, I M; Choo, K H A; Copenhaver, G P; Deluca, J G; Desai, A; Diekmann, S; Erhardt, S; Fitzgerald-Hayes, M; Foltz, D; Fukagawa, T; Gassmann, R; Gerlich, D W; Glover, D M; Gorbsky, G J; Harrison, S C; Heun, P; Hirota, T; Jansen, L E T; Karpen, G; Kops, G J P L; Lampson, M A; Lens, S M; Losada, A; Luger, K; Maiato, H; Maddox, P S; Margolis, R L; Masumoto, H; McAinsh, A D; Mellone, B G; Meraldi, P; Musacchio, A; Oegema, K; O'Neill, R J; Salmon, E D; Scott, K C; Straight, A F; Stukenberg, P T; Sullivan, B A; Sullivan, K F; Sunkel, C E; Swedlow, J R; Walczak, C E; Warburton, P E; Westermann, S; Willard, H F; Wordeman, L; Yanagida, M; Yen, T J; Yoda, K; Cleveland, D W

    2013-04-01

    The first centromeric protein identified in any species was CENP-A, a divergent member of the histone H3 family that was recognised by autoantibodies from patients with scleroderma-spectrum disease. It has recently been suggested to rename this protein CenH3. Here, we argue that the original name should be maintained both because it is the basis of a long established nomenclature for centromere proteins and because it avoids confusion due to the presence of canonical histone H3 at centromeres.

  16. The top-down, middle-down, and bottom-up mass spectrometry approaches for characterization of histone variants and their post-translational modifications.

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    Moradian, Annie; Kalli, Anastasia; Sweredoski, Michael J; Hess, Sonja

    2014-03-01

    Epigenetic regulation of gene expression is, at least in part, mediated by histone modifications. PTMs of histones change chromatin structure and regulate gene transcription, DNA damage repair, and DNA replication. Thus, studying histone variants and their modifications not only elucidates their functional mechanisms in chromatin regulation, but also provides insights into phenotypes and diseases. A challenge in this field is to determine the best approach(es) to identify histone variants and their PTMs using a robust high-throughput analysis. The large number of histone variants and the enormous diversity that can be generated through combinatorial modifications, also known as histone code, makes identification of histone PTMs a laborious task. MS has been proven to be a powerful tool in this regard. Here, we focus on bottom-up, middle-down, and top-down MS approaches, including CID and electron-capture dissociation/electron-transfer dissociation based techniques for characterization of histones and their PTMs. In addition, we discuss advances in chromatographic separation that take advantage of the chemical properties of the specific histone modifications. This review is also unique in its discussion of current bioinformatic strategies for comprehensive histone code analysis.

  17. Histone H1 variant-specific lysine methylation by G9a/KMT1C and Glp1/KMT1D

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    Weiss Thomas

    2010-03-01

    Full Text Available Abstract Background The linker histone H1 has a key role in establishing and maintaining higher order chromatin structure and in regulating gene expression. Mammals express up to 11 different H1 variants, with H1.2 and H1.4 being the predominant ones in most somatic cells. Like core histones, H1 has high levels of covalent modifications; however, the full set of modifications and their biological role are largely unknown. Results In this study, we used a candidate screen to identify enzymes that methylate H1 and to map their corresponding methylation sites. We found that the histone lysine methyltransferases G9a/KMT1C and Glp1/KMT1D methylate H1.2 in vitro and in vivo, and we mapped this novel site to lysine 187 (H1.2K187 in the C-terminus of H1. This H1.2K187 methylation is variant-specific. The main target for methylation by G9a in H1.2, H1.3, H1.5 and H1.0 is in the C-terminus, whereas H1.4 is preferentially methylated at K26 (H1.4K26me in the N-terminus. We found that the readout of these marks is different; H1.4K26me can recruit HP1, but H1.2K187me cannot. Likewise, JMJD2D/KDM4 only reverses H1.4K26 methylation, clearly distinguishing these two methylation sites. Further, in contrast to C-terminal H1 phosphorylation, H1.2K187 methylation level is steady throughout the cell cycle. Conclusions We have characterised a novel methylation site in the C-terminus of H1 that is the target of G9a/Glp1 both in vitro and in vivo. To our knowledge, this is the first demonstration of variant-specific histone methylation by the same methyltransferases, but with differing downstream readers, thereby supporting the hypothesis of H1 variants having specific functions.

  18. Solution structure of histone chaperone ANP32B: interaction with core histones H3-H4 through its acidic concave domain.

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    Tochio, Naoya; Umehara, Takashi; Munemasa, Yoshiko; Suzuki, Toru; Sato, Shin; Tsuda, Kengo; Koshiba, Seizo; Kigawa, Takanori; Nagai, Ryozo; Yokoyama, Shigeyuki

    2010-08-01

    Eukaryotic gene expression is regulated by histone deposition onto and eviction from nucleosomes, which are mediated by several chromatin-modulating factors. Among them, histone chaperones are key factors that facilitate nucleosome assembly. Acidic nuclear phosphoprotein 32B (ANP32B) belongs to the ANP32 family, which shares N-terminal leucine-rich repeats (LRRs) and a C-terminal variable anionic region. The C-terminal region functions as an inhibitor of histone acetylation, but the functional roles of the LRR domain in chromatin regulation have remained elusive. Here, we report that the LRR domain of ANP32B possesses histone chaperone activity and forms a curved structure with a parallel beta-sheet on the concave side and mostly helical elements on the convex side. Our analyses revealed that the interaction of ANP32B with the core histones H3-H4 occurs on its concave side, and both the acidic and hydrophobic residues that compose the concave surface are critical for histone binding. These results provide a structural framework for understanding the functional mechanisms of acidic histone chaperones.

  19. The histone chaperone sNASP binds a conserved peptide motif within the globular core of histone H3 through its TPR repeats.

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    Bowman, Andrew; Lercher, Lukas; Singh, Hari R; Zinne, Daria; Timinszky, Gyula; Carlomagno, Teresa; Ladurner, Andreas G

    2016-04-20

    Eukaryotic chromatin is a complex yet dynamic structure, which is regulated in part by the assembly and disassembly of nucleosomes. Key to this process is a group of proteins termed histone chaperones that guide the thermodynamic assembly of nucleosomes by interacting with soluble histones. Here we investigate the interaction between the histone chaperone sNASP and its histone H3 substrate. We find that sNASP binds with nanomolar affinity to a conserved heptapeptide motif in the globular domain of H3, close to the C-terminus. Through functional analysis of sNASP homologues we identified point mutations in surface residues within the TPR domain of sNASP that disrupt H3 peptide interaction, but do not completely disrupt binding to full length H3 in cells, suggesting that sNASP interacts with H3 through additional contacts. Furthermore, chemical shift perturbations from(1)H-(15)N HSQC experiments show that H3 peptide binding maps to the helical groove formed by the stacked TPR motifs of sNASP. Our findings reveal a new mode of interaction between a TPR repeat domain and an evolutionarily conserved peptide motif found in canonical H3 and in all histone H3 variants, including CenpA and have implications for the mechanism of histone chaperoning within the cell.

  20. Assembly of nucleosomal arrays from recombinant core histones and nucleosome positioning DNA.

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    Rogge, Ryan A; Kalashnikova, Anna A; Muthurajan, Uma M; Porter-Goff, Mary E; Luger, Karolin; Hansen, Jeffrey C

    2013-09-10

    Core histone octamers that are repetitively spaced along a DNA molecule are called nucleosomal arrays. Nucleosomal arrays are obtained in one of two ways: purification from in vivo sources, or reconstitution in vitro from recombinant core histones and tandemly repeated nucleosome positioning DNA. The latter method has the benefit of allowing for the assembly of a more compositionally uniform and precisely positioned nucleosomal array. Sedimentation velocity experiments in the analytical ultracentrifuge yield information about the size and shape of macromolecules by analyzing the rate at which they migrate through solution under centrifugal force. This technique, along with atomic force microscopy, can be used for quality control, ensuring that the majority of DNA templates are saturated with nucleosomes after reconstitution. Here we describe the protocols necessary to reconstitute milligram quantities of length and compositionally defined nucleosomal arrays suitable for biochemical and biophysical studies of chromatin structure and function.

  1. Expression of P. falciparum var genes involves exchange of the histone variant H2A.Z at the promoter.

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    Michaela Petter

    2011-02-01

    Full Text Available Plasmodium falciparum employs antigenic variation to evade the human immune response by switching the expression of different variant surface antigens encoded by the var gene family. Epigenetic mechanisms including histone modifications and sub-nuclear compartmentalization contribute to transcriptional regulation in the malaria parasite, in particular to control antigenic variation. Another mechanism of epigenetic control is the exchange of canonical histones with alternative variants to generate functionally specialized chromatin domains. Here we demonstrate that the alternative histone PfH2A.Z is associated with the epigenetic regulation of var genes. In many eukaryotic organisms the histone variant H2A.Z mediates an open chromatin structure at promoters and facilitates diverse levels of regulation, including transcriptional activation. Throughout the asexual, intraerythrocytic lifecycle of P. falciparum we found that the P. falciparum ortholog of H2A.Z (PfH2A.Z colocalizes with histone modifications that are characteristic of transcriptionally-permissive euchromatin, but not with markers of heterochromatin. Consistent with this finding, antibodies to PfH2A.Z co-precipitate the permissive modification H3K4me3. By chromatin-immunoprecipitation we show that PfH2A.Z is enriched in nucleosomes around the transcription start site (TSS in both transcriptionally active and silent stage-specific genes. In var genes, however, PfH2A.Z is enriched at the TSS only during active transcription in ring stage parasites. Thus, in contrast to other genes, temporal var gene regulation involves histone variant exchange at promoter nucleosomes. Sir2 histone deacetylases are important for var gene silencing and their yeast ortholog antagonises H2A.Z function in subtelomeric yeast genes. In immature P. falciparum parasites lacking Sir2A or Sir2B high var transcription levels correlate with enrichment of PfH2A.Z at the TSS. As Sir2A knock out parasites mature the var

  2. PARP1 orchestrates variant histone exchange in signal-mediated transcriptional activation.

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    O'Donnell, Amanda; Yang, Shen-Hsi; Sharrocks, Andrew D

    2013-12-01

    Transcriptional activation is accompanied by multiple molecular events that remodel the local chromatin environment in promoter regions. These molecular events are often orchestrated in response to the activation of signalling pathways, as exemplified by the response of immediate early genes such as FOS to ERK MAP kinase signalling. Here, we demonstrate that inducible NFI recruitment permits PARP1 binding to the FOS promoter by a mutually reinforcing loop. PARP1 and its poly(ADP-ribosyl)ation activity are required for maintaining FOS activation kinetics. We also show that the histone variant H2A.Z associates with the FOS promoter and acts in a transcription-suppressive manner. However, in response to ERK pathway signalling, H2A.Z is replaced by H2A; PARP1 activity is required to promote this exchange. Thus, our work has revealed an additional facet of PARP1 function in promoting dynamic remodelling of promoter-associated nucleosomes to allow transcriptional activation in response to cellular signalling.

  3. Diversity and Divergence of Dinoflagellate Histone Proteins.

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    Marinov, Georgi K; Lynch, Michael

    2015-12-08

    Histone proteins and the nucleosomal organization of chromatin are near-universal eukaroytic features, with the exception of dinoflagellates. Previous studies have suggested that histones do not play a major role in the packaging of dinoflagellate genomes, although several genomic and transcriptomic surveys have detected a full set of core histone genes. Here, transcriptomic and genomic sequence data from multiple dinoflagellate lineages are analyzed, and the diversity of histone proteins and their variants characterized, with particular focus on their potential post-translational modifications and the conservation of the histone code. In addition, the set of putative epigenetic mark readers and writers, chromatin remodelers and histone chaperones are examined. Dinoflagellates clearly express the most derived set of histones among all autonomous eukaryote nuclei, consistent with a combination of relaxation of sequence constraints imposed by the histone code and the presence of numerous specialized histone variants. The histone code itself appears to have diverged significantly in some of its components, yet others are conserved, implying conservation of the associated biochemical processes. Specifically, and with major implications for the function of histones in dinoflagellates, the results presented here strongly suggest that transcription through nucleosomal arrays happens in dinoflagellates. Finally, the plausible roles of histones in dinoflagellate nuclei are discussed.

  4. One-pot refolding of core histones from bacterial inclusion bodies allows rapid reconstitution of histone octamer.

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    Lee, Young-Tae; Gibbons, Garrett; Lee, Shirley Y; Nikolovska-Coleska, Zaneta; Dou, Yali

    2015-06-01

    We report an optimized method to purify and reconstitute histone octamer, which utilizes high expression of histones in inclusion bodies but eliminates the time consuming steps of individual histone purification. In the newly modified protocol, Xenopus laevis H2A, H2B, H3, and H4 are expressed individually into inclusion bodies of bacteria, which are subsequently mixed together and denatured in 8M guanidine hydrochloride. Histones are refolded and reconstituted into soluble octamer by dialysis against 2M NaCl, and metal-affinity purified through an N-terminal polyhistidine-tag added on the H2A. After cleavage of the polyhistidine-tag, histone octamer is further purified by size exclusion chromatography. We show that the nucleosomes reconstituted using the purified histone octamer above are fully functional. They serve as effective substrates for the histone methyltransferases DOT1L and MLL1. Small angle X-ray scattering further confirms that the reconstituted nucleosomes have correct structural integration of histone octamer and DNA as observed in the X-ray crystal structure. Our new protocol enables rapid reconstitution of histone octamer with an optimal yield. We expect this simplified approach to facilitate research using recombinant nucleosomes in vitro. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. TRF2 Protein Interacts with Core Histones to Stabilize Chromosome Ends.

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    Konishi, Akimitsu; Izumi, Takashi; Shimizu, Shigeomi

    2016-09-23

    Mammalian chromosome ends are protected by a specialized nucleoprotein complex called telomeres. Both shelterin, a telomere-specific multi-protein complex, and higher order telomeric chromatin structures combine to stabilize the chromosome ends. Here, we showed that TRF2, a component of shelterin, binds to core histones to protect chromosome ends from inappropriate DNA damage response and loss of telomeric DNA. The N-terminal Gly/Arg-rich domain (GAR domain) of TRF2 directly binds to the globular domain of core histones. The conserved arginine residues in the GAR domain of TRF2 are required for this interaction. A TRF2 mutant with these arginine residues substituted by alanine lost the ability to protect telomeres and induced rapid telomere shortening caused by the cleavage of a loop structure of the telomeric chromatin. These findings showed a previously unnoticed interaction between the shelterin complex and nucleosomal histones to stabilize the chromosome ends. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. The H1 linker histones: multifunctional proteins beyond the nucleosomal core particle.

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    Hergeth, Sonja P; Schneider, Robert

    2015-11-01

    The linker histone H1 family members are a key component of chromatin and bind to the nucleosomal core particle around the DNA entry and exit sites. H1 can stabilize both nucleosome structure and higher-order chromatin architecture. In general, H1 molecules consist of a central globular domain with more flexible tail regions at both their N- and C-terminal ends. The existence of multiple H1 subtypes and a large variety of posttranslational modifications brings about a considerable degree of complexity and makes studying this protein family challenging. Here, we review recent progress in understanding the function of linker histones and their subtypes beyond their role as merely structural chromatin components. We summarize current findings on the role of H1 in heterochromatin formation, transcriptional regulation and embryogenesis with a focus on H1 subtypes and their specific modifications. © 2015 The Authors.

  7. Cu2+-assisted two dimensional charge-mass double focusing gel electrophoresis and mass spectrometric analysis of histone variants.

    Science.gov (United States)

    Zhang, Wenyang; Tang, Xuemei; Ding, Mengjie; Zhong, Hongying

    2014-12-10

    Abundant isoforms and dynamic posttranslational modifications cause the separation and identification of histone variants to be experimentally challenging. To meet this need, we employ two-dimensional electrophoretic gel separation followed by mass spectrometric detection which takes advantage of the chelation of Cu(2+) with amino acid residues exposed on the surfaces of the histone proteins. Acid-extracted rat liver histones were first mixed with CuSO4 solution and then separated in one dimension with triton-acid-urea (TAU) gel electrophoresis and in a second dimension using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The separations result from both the changes in charge and mass upon Cu(2+) chelation. Identities of each separated gel bands were obtained by using matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS). It was found that the migration of H3 histone isoforms of rat liver is markedly affected by the use of Cu(2+) ions. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. RNA polymerase II pausing downstream of core histone genes is different from genes producing polyadenylated transcripts.

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    Krishanpal Anamika

    Full Text Available Recent genome-wide chromatin immunoprecipitation coupled high throughput sequencing (ChIP-seq analyses performed in various eukaryotic organisms, analysed RNA Polymerase II (Pol II pausing around the transcription start sites of genes. In this study we have further investigated genome-wide binding of Pol II downstream of the 3' end of the annotated genes (EAGs by ChIP-seq in human cells. At almost all expressed genes we observed Pol II occupancy downstream of the EAGs suggesting that Pol II pausing 3' from the transcription units is a rather common phenomenon. Downstream of EAGs Pol II transcripts can also be detected by global run-on and sequencing, suggesting the presence of functionally active Pol II. Based on Pol II occupancy downstream of EAGs we could distinguish distinct clusters of Pol II pause patterns. On core histone genes, coding for non-polyadenylated transcripts, Pol II occupancy is quickly dropping after the EAG. In contrast, on genes, whose transcripts undergo polyA tail addition [poly(A(+], Pol II occupancy downstream of the EAGs can be detected up to 4-6 kb. Inhibition of polyadenylation significantly increased Pol II occupancy downstream of EAGs at poly(A(+ genes, but not at the EAGs of core histone genes. The differential genome-wide Pol II occupancy profiles 3' of the EAGs have also been confirmed in mouse embryonic stem (mES cells, indicating that Pol II pauses genome-wide downstream of the EAGs in mammalian cells. Moreover, in mES cells the sharp drop of Pol II signal at the EAG of core histone genes seems to be independent of the phosphorylation status of the C-terminal domain of the large subunit of Pol II. Thus, our study uncovers a potential link between different mRNA 3' end processing mechanisms and consequent Pol II transcription termination processes.

  9. Different reaction of the core histones H2A and H2B to red laser irradiation

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    Brill, G. E.; Egorova, A. V.; Bugaeva, I. O.; Postnov, D. E.; Ushakova, O. V.

    2017-03-01

    Analysis of the influence of red laser irradiation on the processes of self-assembly of the core histones H2A and H2B was performed using a wedge dehydration method. Image-analysis of facies included their qualitative characteristics and calculation of quantitative parameters with subsequent statistical processing. It was established that linearly polarized red laser light (λ - 660 nm, 1 J/cm2) significantly modified the process of self-assembly of core histone H2B, whereas the structure of the facies of H2A histone changed to a lesser extent. Histones were used in the form of aqueous salt solutions. The effect of red light seems to result from the formation of singlet oxygen by direct laser excitation of molecular oxygen.

  10. The histone variant His2Av is required for adult stem cell maintenance in the Drosophila testis.

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    Jose Rafael Morillo Prado

    2013-11-01

    Full Text Available Many tissues are sustained by adult stem cells, which replace lost cells by differentiation and maintain their own population through self-renewal. The mechanisms through which adult stem cells maintain their identity are thus important for tissue homeostasis and repair throughout life. Here, we show that a histone variant, His2Av, is required cell autonomously for maintenance of germline and cyst stem cells in the Drosophila testis. The ATP-dependent chromatin-remodeling factor Domino is also required in this tissue for adult stem cell maintenance possibly by regulating the incorporation of His2Av into chromatin. Interestingly, although expression of His2Av was ubiquitous, its function was dispensable for germline and cyst cell differentiation, suggesting a specific role for this non-canonical histone in maintaining the stem cell state in these lineages.

  11. Histone variant macroH2A1 deletion in mice causes female-specific steatosis

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    Boulard Mathieu

    2010-04-01

    Full Text Available Abstract Background Vertebrate heterochromatin contains a non-allelic variant of the histone H2A called macroH2A1, which has the characteristic of being three times the size of the canonical H2A. The macroH2A1 C-terminal extension can recruit onto chromatin the poly-ADP-ribose polymerase (PARP1, which is crucial for DNA repair. This led to the speculation that macroH2A1 could be essential for genome surveillance; however, no experimental evidence supported this hypothesis. Because macroH2A1 has been found to be enriched on the inactive X-chromosome in females, it is thought to play a role in sex chromosome dosage compensation through its ability to regulate gene expression. However, more genetic data are needed to further understand the function of macroH2A1 in mammals. Results Deletion of the murine gene H2afy, which encodes for macroH2A1, resulted in lipid accumulation in liver. Hepatic steatosis caused by H2afy disruption occurred specifically in homozygous mutant females. The metabolic disorder constantly affected half of the number of homozygote females. Given the mixed genetic background of the mutants, an unreported genetic modifier is likely to influence the penetrance of the phenotype. In addition, the X-linked thyroxine-binding globulin (Tbg gene was specifically upregulated in steatotic livers. Chromatin immunoprecitation indicated that macroH2A1 is enriched at the Tbg promoter in wild-type female animals, indicating that increased Tbg expression in H2afy null mutants is likely to be a direct consequence of the absence of macroH2A1. Furthermore, male mice, which are not prone to the metabolic disorder, had a reduced level of macroH2A1 incorporated into the Tbg promoter. Conclusions Because TBG is the main carrier of the thyroid hormone T4, which regulates energy metabolism, we propose that overexpression of TBG is responsible for the fat accumulation observed in H2afy-deficient liver. Moreover, our results suggest that the sexual

  12. Anticancer drug mithramycin interacts with core histones: An additional mode of action of the DNA groove binder

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    Amrita Banerjee

    2014-01-01

    Full Text Available Mithramycin (MTR is a clinically approved DNA-binding antitumor antibiotic currently in Phase 2 clinical trials at National Institutes of Health for treatment of osteosarcoma. In view of the resurgence in the studies of this generic antibiotic as a human medicine, we have examined the binding properties of MTR with the integral component of chromatin – histone proteins – as a part of our broad objective to classify DNA-binding molecules in terms of their ability to bind chromosomal DNA alone (single binding mode or both histones and chromosomal DNA (dual binding mode. The present report shows that besides DNA, MTR also binds to core histones present in chromatin and thus possesses the property of dual binding in the chromatin context. In contrast to the MTR–DNA interaction, association of MTR with histones does not require obligatory presence of bivalent metal ion like Mg2+. As a consequence of its ability to interact with core histones, MTR inhibits histone H3 acetylation at lysine 18, an important signature of active chromatin, in vitro and ex vivo. Reanalysis of microarray data of Ewing sarcoma cell lines shows that upon MTR treatment there is a significant down regulation of genes, possibly implicating a repression of H3K18Ac-enriched genes apart from DNA-binding transcription factors. Association of MTR with core histones and its ability to alter post-translational modification of histone H3 clearly indicates an additional mode of action of this anticancer drug that could be implicated in novel therapeutic strategies.

  13. Identification and characterization of the two isoforms of the vertebrate H2A.Z histone variant.

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    Matsuda, Ryo; Hori, Tetsuya; Kitamura, Hiroshi; Takeuchi, Kozo; Fukagawa, Tatsuo; Harata, Masahiko

    2010-07-01

    Histone variants play important roles in the epigenetic regulation of genome function. The histone variant H2A.Z is evolutionarily conserved from yeast to vertebrates, and it has been reported to have multiple effects upon gene expression and insulation, and chromosome segregation. Recently two genes encoding H2A.Z were identified in the vertebrate genome. However, it is not yet clear whether the proteins transcribed from these genes are functionally distinct. To address this issue, we knocked out each gene individually in chicken DT40 cells. We found that two distinct proteins, H2A.Z-1 and H2A.Z-2, were produced from these genes, and that these proteins could be separated on a long SDS-PAGE gel. The two isoforms were deposited to a similar extent by the SRCAP chromatin-remodeling complex, suggesting redundancy to their function. However, cells lacking either one of the two isoforms exhibited distinct alterations in cell growth and gene expression, suggesting that the two isoforms have differential effects upon nucleosome stability and chromatin structure. These findings provide insight into the molecular basis of the multiple functions of the H2A.Z gene products.

  14. Purification and characterization of adenovirus core protein VII: a histone-like protein that is critical for adenovirus core formation.

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    Sharma, Gaurav; Moria, Nithesh; Williams, Martin; Krishnarjuna, Bankala; Pouton, Colin W

    2017-07-01

    Adenovirus protein VII is a highly cationic core protein that forms a nucleosome-like structure in the adenovirus core by condensing DNA in combination with protein V and mu. It has been proposed that protein VII could condense DNA in a manner analogous to mammalian histones. Due to the lack of an expression and purification protocol, the interactions between protein VII and DNA are poorly understood. In this study we describe methods for the purification of biologically active recombinant protein VII using an E. coli expression system. We expressed a cleavable fusion of protein VII with thioredoxin and established methods for purification of this fusion protein in denatured form. We describe an efficient method for resolving the cleavage products to obtain pure protein VII using hydroxyapatite column chromatography. Mass spectroscopy data confirmed its mass and purity to be 19.4 kDa and >98 %, respectively. Purified recombinant protein VII spontaneously condensed dsDNA to form particles, as shown by dye exclusion assay, electrophoretic mobility shift assay and nuclease protection assay. Additionally, an in vitro bioluminescence assay revealed that protein VII can be used to enhance the transfection of mammalian cells with lipofectamine/DNA complexes. The availability of recombinant protein VII will facilitate future studies of the structure of the adenovirus core. Improved understanding of the structure and function of protein VII will be valuable in elucidating the mechanism of adenoviral DNA condensation, defining the morphology of the adenovirus core and establishing the mechanism by which adenoviral DNA enters the nucleus.

  15. Replication-Independent Histone Variant H3.3 Controls Animal Lifespan through the Regulation of Pro-longevity Transcriptional Programs

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    Antonia Piazzesi

    2016-10-01

    Full Text Available Chromatin structure orchestrates the accessibility to the genetic material. Replication-independent histone variants control transcriptional plasticity in postmitotic cells. The life-long accumulation of these histones has been described, yet the implications on organismal aging remain elusive. Here, we study the importance of the histone variant H3.3 in Caenorhabditis elegans longevity pathways. We show that H3.3-deficient nematodes have negligible lifespan differences compared to wild-type animals. However, H3.3 is essential for the lifespan extension of C. elegans mutants in which pronounced transcriptional changes control longevity programs. Notably, H3.3 loss critically affects the expression of a very large number of genes in long-lived nematodes, resulting in transcriptional profiles similar to wild-type animals. We conclude that H3.3 positively contributes to diverse lifespan-extending signaling pathways, with potential implications on age-related processes in multicellular organisms.

  16. DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression

    NARCIS (Netherlands)

    Borghesan, Michela; Fusilli, Caterina; Rappa, Francesca; Panebianco, Concetta; Rizzo, Giovanni; Oben, Jude A.; Mazzoccoli, Gianluigi; Faulkes, Chris; Pata, Illar; Agodi, Antonella; Rezaee, Farhad; Minogue, Shane; Warren, Alessandra; Peterson, Abigail; Sedivy, John M.; Douet, Julien; Buschbeck, Marcus; Cappello, Francesco; Mazza, Tommaso; Pazienza, Valerio; Vinciguerra, Manlio

    2016-01-01

    Aging is a major risk factor for progression of liver diseases to hepatocellular carcinoma (HCC). Cellular senescence contributes to age-related tissue dysfunction, but the epigenetic basis underlying drug-induced senescence remains unclear. macroH2A1, a variant of histone H2A, is a marker of

  17. The histone acetyltransferase MOF is a key regulator of the embryonic stem cell core transcriptional network.

    Science.gov (United States)

    Li, Xiangzhi; Li, Li; Pandey, Ruchi; Byun, Jung S; Gardner, Kevin; Qin, Zhaohui; Dou, Yali

    2012-08-03

    Pluripotent embryonic stem cells (ESCs) maintain self-renewal and the potential for rapid response to differentiation cues. Both ESC features are subject to epigenetic regulation. Here we show that the histone acetyltransferase Mof plays an essential role in the maintenance of ESC self-renewal and pluripotency. ESCs with Mof deletion lose characteristic morphology, alkaline phosphatase (AP) staining, and differentiation potential. They also have aberrant expression of the core transcription factors Nanog, Oct4, and Sox2. Importantly, the phenotypes of Mof null ESCs can be partially suppressed by Nanog overexpression, supporting the idea that Mof functions as an upstream regulator of Nanog in ESCs. Genome-wide ChIP-sequencing and transcriptome analyses further demonstrate that Mof is an integral component of the ESC core transcriptional network and that Mof primes genes for diverse developmental programs. Mof is also required for Wdr5 recruitment and H3K4 methylation at key regulatory loci, highlighting the complexity and interconnectivity of various chromatin regulators in ESCs.

  18. Expressed Centromere Specific Histone 3 (CENH3 Variants in Cultivated Triploid and Wild Diploid Bananas (Musa spp.

    Directory of Open Access Journals (Sweden)

    Kariuki S. Muiruri

    2017-06-01

    Full Text Available Centromeres are specified by a centromere specific histone 3 (CENH3 protein, which exists in a complex environment, interacting with conserved proteins and rapidly evolving satellite DNA sequences. The interactions may become more challenging if multiple CENH3 versions are introduced into the zygote as this can affect post-zygotic mitosis and ultimately sexual reproduction. Here, we characterize CENH3 variant transcripts expressed in cultivated triploid and wild diploid progenitor bananas. We describe both splice- and allelic-[Single Nucleotide Polymorphisms (SNP] variants and their effects on the predicted secondary structures of protein. Expressed CENH3 transcripts from six banana genotypes were characterized and clustered into three groups (MusaCENH-1A, MusaCENH-1B, and MusaCENH-2 based on similarity. The CENH3 groups differed with SNPs as well as presence of indels resulting from retained and/or skipped exons. The CENH3 transcripts from different banana genotypes were spliced in either 7/6, 5/4 or 6/5 exons/introns. The 7/6 and the 5/4 exon/intron structures were found in both diploids and triploids, however, 7/6 was most predominant. The 6/5 exon/introns structure was a result of failure of the 7/6 to splice correctly. The various transcripts obtained were predicted to encode highly variable N-terminal tails and a relatively conserved C-terminal histone fold domain (HFD. The SNPs were predicted in some cases to affect the secondary structure of protein by lengthening or shorting the affected domains. Sequencing of banana CENH3 transcripts predicts SNP variations that affect amino acid sequences and alternatively spliced transcripts. Most of these changes affect the N-terminal tail of CENH3.

  19. Affinity of anticancer drug, daunomycin, to core histones in solution:comparison of free and cross-linked proteins

    Institute of Scientific and Technical Information of China (English)

    Azra RABBANI; Sayeh ABDOSAMADI; Naghmeh SARI-SARAF

    2007-01-01

    Aim: The interaction of anthracyclinc anticancer drugs with chromatin, nuclco-somes and historic H1 has been extensively studied. In the present study, for the first time, we have investigated the binding of anthracycline antibiotic, daunomycin,to free and cross-linked thymus core histones (CL-core) in solution and in the absence of DNA. Methods: Fluorescence, UV/Vis spectroscopy and equilibrium dialysis techniques were used. Results: The UV spectroscopy results show that daunomycin induces hypochromicity in the absorption spectra of the core histones.Fluorescence emission intensity is decreased upon daunomycin binding and the process is concentration dependent. The equilibrium dialysis shows that the bind-ing is positive cooperative with the binding sites as Scatchard plot and Hill Coef-ficient confirm it. Conclusion: The results suggest that daunomycin shows much higher affinity to core histories free in solution than to CL-core, implying that the binding is most likely due to the accessibility of these proteins to the environment.It is suggested that daunomycin binds strongly to open state of histones, such as in tumor cells, rather than to their compact structure seen in normal chromatin.

  20. Histone chaperones: assisting histone traffic and nucleosome dynamics.

    Science.gov (United States)

    Gurard-Levin, Zachary A; Quivy, Jean-Pierre; Almouzni, Geneviève

    2014-01-01

    The functional organization of eukaryotic DNA into chromatin uses histones as components of its building block, the nucleosome. Histone chaperones, which are proteins that escort histones throughout their cellular life, are key actors in all facets of histone metabolism; they regulate the supply and dynamics of histones at chromatin for its assembly and disassembly. Histone chaperones can also participate in the distribution of histone variants, thereby defining distinct chromatin landscapes of importance for genome function, stability, and cell identity. Here, we discuss our current knowledge of the known histone chaperones and their histone partners, focusing on histone H3 and its variants. We then place them into an escort network that distributes these histones in various deposition pathways. Through their distinct interfaces, we show how they affect dynamics during DNA replication, DNA damage, and transcription, and how they maintain genome integrity. Finally, we discuss the importance of histone chaperones during development and describe how misregulation of the histone flow can link to disease.

  1. Rapid divergence of histones in Hydrozoa (Cnidaria) and evolution of a novel histone involved in DNA damage response in hydra.

    Science.gov (United States)

    Reddy, Puli Chandramouli; Ubhe, Suyog; Sirwani, Neha; Lohokare, Rasika; Galande, Sanjeev

    2017-08-01

    Histones are fundamental components of chromatin in all eukaryotes. Hydra, an emerging model system belonging to the basal metazoan phylum Cnidaria, provides an ideal platform to understand the evolution of core histone components at the base of eumetazoan phyla. Hydra exhibits peculiar properties such as tremendous regenerative capacity, lack of organismal senescence and rarity of malignancy. In light of the role of histone modifications and histone variants in these processes it is important to understand the nature of histones themselves and their variants in hydra. Here, we report identification of the complete repertoire of histone-coding genes in the Hydra magnipapillata genome. Hydra histones were classified based on their copy numbers, gene structure and other characteristic features. Genomic organization of canonical histone genes revealed the presence of H2A-H2B and H3-H4 paired clusters in high frequency and also a cluster with all core histones along with H1. Phylogenetic analysis of identified members of H2A and H2B histones suggested rapid expansion of these groups in Hydrozoa resulting in the appearance of unique subtypes. Amino acid sequence level comparisons of H2A and H2B forms with bilaterian counterparts suggest the possibility of a highly mobile nature of nucleosomes in hydra. Absolute quantitation of transcripts confirmed the high copy number of histones and supported the canonical nature of H2A. Furthermore, functional characterization of H2A.X.1 and a unique variant H2A.X.2 in the gastric region suggest their role in the maintenance of genome integrity and differentiation processes. These findings provide insights into the evolution of histones and their variants in hydra. Copyright © 2017 Elsevier GmbH. All rights reserved.

  2. Three-Dimensional Architecture of the Human BRCA1-A Histone Deubiquitinase Core Complex

    Directory of Open Access Journals (Sweden)

    Otto J.P. Kyrieleis

    2016-12-01

    Full Text Available BRCA1 is a tumor suppressor found to be mutated in hereditary breast and ovarian cancer and plays key roles in the maintenance of genomic stability by homologous recombination repair. It is recruited to damaged chromatin as a component of the BRCA1-A deubiquitinase, which cleaves K63-linked ubiquitin chains attached to histone H2A and H2AX. BRCA1-A contributes to checkpoint regulation, repair pathway choice, and HR repair efficiency through molecular mechanisms that remain largely obscure. The structure of an active core complex comprising two Abraxas/BRCC36/BRCC45/MERIT40 tetramers determined by negative-stain electron microscopy (EM reveals a distorted V-shape architecture in which a dimer of Abraxas/BRCC36 heterodimers sits at the base, with BRCC45/Merit40 pairs occupying each arm. The location and ubiquitin-binding activity of BRCC45 suggest that it may provide accessory interactions with nucleosome-linked ubiquitin chains that contribute to their efficient processing. Our data also suggest how ataxia telangiectasia mutated (ATM-dependent BRCA1 dimerization may stabilize self-association of the entire BRCA1-A complex.

  3. The histone variant macroH2A is an epigenetic regulator of key developmental genes

    DEFF Research Database (Denmark)

    Buschbeck, Marcus; Uribesalgo, Iris; Wibowo, Indra;

    2009-01-01

    variants at many genes encoding key regulators of development and cell fate decisions. On these genes, the presence of macroH2A1+2 is a repressive mark that overlaps locally and functionally with Polycomb repressive complex 2. We demonstrate that macroH2A1+2 contribute to the fine-tuning of temporal...... activation of HOXA cluster genes during neuronal differentiation. Furthermore, elimination of macroH2A2 function in zebrafish embryos produced severe but specific phenotypes. Taken together, our data demonstrate that macroH2A variants constitute an important epigenetic mark involved in the concerted...... regulation of gene expression programs during cellular differentiation and vertebrate development....

  4. Analysis of the Genes Encoding the Histones of Microsporidia Nosema bombycis

    Directory of Open Access Journals (Sweden)

    Liu Yang

    2013-02-01

    Full Text Available Histone proteins are essential components of eukaryotic chromosomes, the objective of the study is to provide some new insights into its evolution through analysis of N. bombycis Histone genes at genomic level. In the study, genes encoding core Histone H2A, H2B, H3 and H4 from Nosema bombycis were analyzed by multiple sequence alignments. Analysis showed that: each type of the core Histone genes, sharing high similarity with each other in both coding and non-coding regions, has low copy number. Multiple sequence alignments showed N. bombycis core Histones diverge obviously, relative-rate test revealed Histone proteins have accelerated in the evolutionary rate of amino acid substitution. The distance between the stop codon and consensus poly (A signal is compacted, no conserved hair-pin element was found in 3'-untranslated regions of Histone mRNAs and overlapping gene transcription was observed in the downstream region of Histone variant H3_3, that implies there maybe have only single class of core Histone genes encoding replication-independent Histones in N. bombycis. Surveying the upstream of the coding region of all core Histone genes, there were no canonical TATA or CAAT boxes except that a common Histone motif (TTTCCCTCC was discovered. Moreover, no similar Histone motif mentioned above existed in Encephalitozoon cuniculi, the closely related organisms. That means that similar Histone motif maybe exists in microsporidian last common ancestor, N. bombycis retained Histone motif, while E. cuniculi have lost Histone motif after the differentiation from the common ancestor with the change of the host. Therefore the analysis of the genes encoding the Histones ofN. bombycis revealed that there maybe have two evolution directions in microsporidia, that is, genome extreme compact and mild compact, during the course of evolution. It contributes us to have the knowledge of that there have different genome size in microsporidia and provide useful

  5. Computer modeling reveals that modifications of the histone tail charges define salt-dependent interaction of the nucleosome core particles.

    Science.gov (United States)

    Yang, Ye; Lyubartsev, Alexander P; Korolev, Nikolay; Nordenskiöld, Lars

    2009-03-18

    Coarse-grained Langevin molecular dynamics computer simulations were conducted for systems that mimic solutions of nucleosome core particles (NCPs). The NCP was modeled as a negatively charged spherical particle representing the complex of DNA and the globular part of the histones combined with attached strings of connected charged beads modeling the histone tails. The size, charge, and distribution of the tails relative to the core were built to match real NCPs. Three models of NCPs were constructed to represent different extents of covalent modification on the histone tails: (nonmodified) recombinant (rNCP), acetylated (aNCP), and acetylated and phosphorylated (paNCP). The simulation cell contained 10 NCPs in a dielectric continuum with explicit mobile counterions and added salt. The NCP-NCP interaction is decisively dependent on the modification state of the histone tails and on salt conditions. Increasing the monovalent salt concentration (KCl) from salt-free to physiological concentration leads to NCP aggregation in solution for rNCP, whereas NCP associates are observed only occasionally in the system of aNCPs. In the presence of divalent salt (Mg(2+)), rNCPs form dense stable aggregates, whereas aNCPs form aggregates less frequently. Aggregates are formed via histone-tail bridging and accumulation of counterions in the regions of NCP-NCP contacts. The paNCPs do not show NCP-NCP interaction upon addition of KCl or in the presence of Mg(2+). Simulations for systems with a gradual substitution of K(+) for Mg(2+), to mimic the Mg(2+) titration of an NCP solution, were performed. The rNCP system showed stronger aggregation that occurred at lower concentrations of added Mg(2+), compared to the aNCP system. Additional molecular dynamics simulations performed with a single NCP in the simulation cell showed that detachment of the tails from the NCP core was modest under a wide range of salt concentrations. This implies that salt-induced tail dissociation of the

  6. Core Histone Tail Domains Mediate Oligonucleosome Folding and Nucleosomal DNA Organization through Distinct Molecular Mechanisms

    National Research Council Canada - National Science Library

    Terace M. Fletcher; Jeffrey C. Hansen

    1995-01-01

    .... wrapping of DNA around the histone octamer. Mg ions can substitute for the tail domains to yield a trypsinized oligonucleosome structure that is indistinguishable from that of an intact nucleosomal array in low salt...

  7. Chromatin remodeling defects in pediatric and young adult glioblastoma: a tale of a variant histone 3 tail.

    Science.gov (United States)

    Fontebasso, Adam M; Liu, Xiao-Yang; Sturm, Dominik; Jabado, Nada

    2013-03-01

    Primary brain tumors occur in 8 out of 100 000 people and are the leading cause of cancer-related death in children. Among brain tumors, high-grade astrocytomas (HGAs) including glioblastoma multiforme (GBM) are aggressive and are lethal human cancers. Despite decades of concerted therapeutic efforts, HGAs remain essentially incurable in adults and children. Recent discoveries have revolutionized our understanding of these tumors in children and young adults. Recurrent somatic driver mutations in the tail of histone 3 variant 3 (H3.3), leading to amino acid substitutions at key residues, namely lysine (K) 27 (K27M) and glycine 34 (G34R/G34V), were identified as a new molecular mechanism in pediatric GBM. These mutations represent the pediatric counterpart of the recurrent mutations in isocitrate dehydrogenases (IDH) identified in young adult gliomas and provide a much-needed new pathway that can be targeted for therapeutic development. This review will provide an overview of the potential role of these mutations in altering chromatin structure and affecting specific molecular pathways ultimately leading to gliomagenesis. The distinct changes in chromatin structure and the specific downstream events induced by each mutation need characterizing independently if progress is to be made in tackling this devastating cancer. © 2013 The Authors; Brain Pathology © 2013 International Society of Neuropathology.

  8. Distinct distribution of ectopically expressed histone variants H2A.Bbd and MacroH2A in open and closed chromatin domains.

    Directory of Open Access Journals (Sweden)

    Elena S Ioudinkova

    Full Text Available BACKGROUND: It becomes increasingly evident that nuclesomes are far from being identical to each other. This nucleosome diversity is due partially to the existence of histone variants encoded by separate genes. Among the known histone variants the less characterized are H2A.Bbd and different forms of macroH2A. This is especially true in the case of H2A.Bbd as there are still no commercially available antibodies specific to H2A.Bbd that can be used for chromatin immunoprecipitation (ChIP. METHODS: We have generated HeLa S3 cell lines stably expressing epitope-tagged versions of macroH2A1.1, H2A.Bbd or canonical H2A and analyzed genomic distribution of the tagged histones using ChIP-on-chip technique. RESULTS: The presence of histone H2A variants macroH2A1.1 and H2A.Bbd has been analyzed in the chromatin of several segments of human chromosomes 11, 16 and X that have been chosen for their different gene densities and chromatin status. Chromatin immunoprecipitation (ChIP followed by hybridization with custom NimbleGene genomic microarrays demonstrated that in open chromatin domains containing tissue-specific along with housekeeping genes, the H2A.Bbd variant was preferentially associated with the body of a subset of transcribed genes. The macroH2A1.1 variant was virtually absent from some genes and underrepresented in others. In contrast, in closed chromatin domains which contain only tissue-specific genes inactive in HeLa S3 cells, both macroH2A1.1 and H2A.Bbd histone variants were present and often colocalized. CONCLUSIONS: Genomic distribution of macro H2A and H2A.Bbd does not follow any simple rule and is drastically different in open and closed genomic domains.

  9. Onset of grain filling is associated with a change in properties of linker histone variants in maize kernels

    DEFF Research Database (Denmark)

    Kalamajka, R.; Finnie, Christine; Grasser, K.D.

    2010-01-01

    to more basic protein forms suggests a reduction in the level of post-translational modifications of linker histones during kernel development. Analysis of their DNA-binding affinity revealed that the different linker histone gene products bind double-stranded DNA with similar affinity. Interestingly......) initiation of storage synthesis. Six linker histone gene products were identified by MALDI-TOF mass spectrometry. A marked shift of around 4 pH units was observed for the linker histone spot pattern after 2D-gel electrophoresis when comparing the proteins of 11 and 16 dap kernels. The shift from acidic......, the linker histones isolated from 16 dap kernels consistently displayed a lower affinity for DNA than the proteins isolated from 11 dap kernels. These findings suggest that the affinity for DNA of the linker histones may be regulated by post-translational modification and that the reduction in DNA affinity...

  10. Proteome analysis of protein partners to nucleosomes containing canonical H2A or the variant histones H2A.Z or H2A.X.

    Science.gov (United States)

    Fujimoto, Satoru; Seebart, Corrine; Guastafierro, Tiziana; Prenni, Jessica; Caiafa, Paola; Zlatanova, Jordanka

    2012-01-01

    Although the existence of histone variants has been known for quite some time, only recently are we grasping the breadth and diversity of the cellular processes in which they are involved. Of particular interest are the two variants of histone H2A, H2A.Z and H2A.X because of their roles in regulation of gene expression and in DNA double-strand break repair, respectively. We hypothesize that nucleosomes containing these variants may perform their distinct functions by interacting with different sets of proteins. Here, we present our proteome analysis aimed at identifying protein partners that interact with nucleosomes containing H2A.Z, H2A.X or their canonical H2A counterpart. Our development of a nucleosome-pull down assay and analysis of the recovered nucleosome-interacting proteins by mass spectrometry allowed us to directly compare nuclear partners of these variant-containing nucleosomes to those containing canonical H2A. To our knowledge, our data represent the first systematic analysis of the H2A.Z and H2A.X interactome in the context of nucleosome structure.

  11. Rapid Histone-Catalyzed DNA Lesion Excision and Accompanying Protein Modification in Nucleosomes and Nucleosome Core Particles.

    Science.gov (United States)

    Weng, Liwei; Greenberg, Marc M

    2015-09-01

    C5'-Hydrogen atoms are frequently abstracted during DNA oxidation. The oxidized abasic lesion 5'-(2-phosphoryl-1,4-dioxobutane) (DOB) is an electrophilic product of the C5'-radical. DOB is a potent irreversible inhibitor of DNA polymerase β, and forms interstrand cross-links in free DNA. We examined the reactivity of DOB within nucleosomes and nucleosome core particles (NCPs), the monomeric component of chromatin. Depending upon the position at which DOB is generated within a NCP, it is excised from nucleosomal DNA at a rate 275-1500-fold faster than that in free DNA. The half-life of DOB (7.0-16.8 min) in NCPs is shorter than any other abasic lesion. DOB's lifetime in NCPs is also significantly shorter than the estimated lifetime of an abasic site within a cell, suggesting that the observed chemistry would occur intracellularly. Histones also catalyze DOB excision when the lesion is present in the DNA linker region of a nucleosome. Schiff-base formation between DOB and histone proteins is detected in nucleosomes and NCPs, resulting in pyrrolone formation at the lysine residues. The lysines modified by DOB are often post-translationally modified. Consequently, the histone modifications described herein could affect the regulation of gene expression and may provide a chemical basis for the cytotoxicity of the DNA damaging agents that produce this lesion.

  12. Histone H1 Variants in Arabidopsis Are Subject to Numerous Post-Translational Modifications, Both Conserved and Previously Unknown in Histones, Suggesting Complex Functions of H1 in Plants.

    Science.gov (United States)

    Kotliński, Maciej; Rutowicz, Kinga; Kniżewski, Łukasz; Palusiński, Antoni; Olędzki, Jacek; Fogtman, Anna; Rubel, Tymon; Koblowska, Marta; Dadlez, Michał; Ginalski, Krzysztof; Jerzmanowski, Andrzej

    2016-01-01

    Linker histones (H1s) are conserved and ubiquitous structural components of eukaryotic chromatin. Multiple non-allelic variants of H1, which differ in their DNA/nucleosome binding properties, co-exist in animal and plant cells and have been implicated in the control of genetic programs during development and differentiation. Studies in mammals and Drosophila have revealed diverse post-translational modifications of H1s, most of which are of unknown function. So far, it is not known how this pattern compares with that of H1s from other major lineages of multicellular Eukaryotes. Here, we show that the two main H1variants of a model flowering plant Arabidopsis thaliana are subject to a rich and diverse array of post-translational modifications. The distribution of these modifications in the H1 molecule, especially in its globular domain (GH1), resembles that occurring in mammalian H1s, suggesting that their functional significance is likely to be conserved. While the majority of modifications detected in Arabidopsis H1s, including phosphorylation, acetylation, mono- and dimethylation, formylation, crotonylation and propionylation, have also been reported in H1s of other species, some others have not been previously identified in histones.

  13. Functional Characterization of C-terminal Ryanodine Receptor 1 Variants Associated with Central Core Disease or Malignant Hyperthermia.

    Science.gov (United States)

    Parker, Remai; Schiemann, Anja H; Langton, Elaine; Bulger, Terasa; Pollock, Neil; Bjorksten, Andrew; Gillies, Robyn; Hutchinson, David; Roxburgh, Richard; Stowell, Kathryn M

    2017-01-01

    Central core disease and malignant hyperthermia are human disorders of skeletal muscle resulting from aberrant Ca2+ handling. Most malignant hyperthermia and central core disease cases are associated with amino acid changes in the type 1 ryanodine receptor (RyR1), the skeletal muscle Ca2+-release channel. Malignant hyperthermia exhibits a gain-of-function phenotype, and central core disease results from loss of channel function. For a variant to be classified as pathogenic, functional studies must demonstrate a correlation with the pathophysiology of malignant hyperthermia or central core disease. We assessed the pathogenicity of four C-terminal variants of the ryanodine receptor using functional analysis. The variants were identified in families affected by either malignant hyperthermia or central core disease. Four variants were introduced separately into human cDNA encoding the skeletal muscle ryanodine receptor. Following transient expression in HEK-293T cells, functional studies were carried out using calcium release assays in response to an agonist. Two previously characterized variants and wild-type skeletal muscle ryanodine receptor were used as controls. The p.Met4640Ile variant associated with central core disease showed no difference in calcium release compared to wild-type. The p.Val4849Ile variant associated with malignant hyperthermia was more sensitive to agonist than wild-type but did not reach statistical significance and two variants (p.Phe4857Ser and p.Asp4918Asn) associated with central core disease were completely inactive. The p.Val4849Ile variant should be considered a risk factor for malignant hyperthermia, while the p.Phe4857Ser and p.Asp4918Asn variants should be classified as pathogenic for central core disease.

  14. Linker histones in hormonal gene regulation.

    Science.gov (United States)

    Vicent, G P; Wright, R H G; Beato, M

    2016-03-01

    In the present review, we summarize advances in our knowledge on the role of the histone H1 family of proteins in breast cancer cells, focusing on their response to progestins. Histone H1 plays a dual role in gene regulation by hormones, both as a structural component of chromatin and as a dynamic modulator of transcription. It contributes to hormonal regulation of the MMTV promoter by stabilizing a homogeneous nucleosome positioning, which reduces basal transcription whereas at the same time promoting progesterone receptor binding and nucleosome remodeling. These combined effects enhance hormone dependent gene transcription, which eventually requires H1 phosphorylation and displacement. Various isoforms of histone H1 have specific functions in differentiated breast cancer cells and compact nucleosomal arrays to different extents in vitro. Genome-wide studies show that histone H1 has a key role in chromatin dynamics of hormone regulated genes. A complex sequence of enzymatic events, including phosphorylation by CDK2, PARylation by PARP1 and the ATP-dependent activity of NURF, are required for H1 displacement and gene de-repression, as a prerequisite for further nucleosome remodeling. Similarly, during hormone-dependent gene repression a dedicated enzymatic mechanism controls H1 deposition at promoters by a complex containing HP1γ, LSD1 and BRG1, the ATPase of the BAF complex. Thus, a broader vision of the histone code should include histone H1, as the linker histone variants actively participate in the regulation of the chromatin structure. How modifications of the core histones tails affect H1 modifications and vice versa is one of the many questions that remains to be addressed to provide a more comprehensive view of the histone cross-talk mechanisms.

  15. New players in heterochromatin silencing: histone variant H3.3 and the ATRX/DAXX chaperone.

    Science.gov (United States)

    Voon, Hsiao P J; Wong, Lee H

    2016-02-29

    A number of studies have demonstrated that various components of the ATRX/DAXX/Histone H3.3 complex are important for heterochromatin silencing at multiple genomic regions. We provide an overview of the individual components (ATRX, DAXX and/or H3.3) tested in each study and propose a model where the ATRX/DAXX chaperone complex deposits H3.3 to maintain the H3K9me3 modification at heterochromatin throughout the genome.

  16. DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression.

    Science.gov (United States)

    Borghesan, Michela; Fusilli, Caterina; Rappa, Francesca; Panebianco, Concetta; Rizzo, Giovanni; Oben, Jude A; Mazzoccoli, Gianluigi; Faulkes, Chris; Pata, Illar; Agodi, Antonella; Rezaee, Farhad; Minogue, Shane; Warren, Alessandra; Peterson, Abigail; Sedivy, John M; Douet, Julien; Buschbeck, Marcus; Cappello, Francesco; Mazza, Tommaso; Pazienza, Valerio; Vinciguerra, Manlio

    2016-02-01

    Aging is a major risk factor for progression of liver diseases to hepatocellular carcinoma (HCC). Cellular senescence contributes to age-related tissue dysfunction, but the epigenetic basis underlying drug-induced senescence remains unclear. macroH2A1, a variant of histone H2A, is a marker of senescence-associated heterochromatic foci that synergizes with DNA methylation to silence tumor-suppressor genes in human fibroblasts. In this study, we investigated the relationship between macroH2A1 splice variants, macroH2A1.1 and macroH2A1.2, and liver carcinogenesis. We found that protein levels of both macroH2A1 isoforms were increased in the livers of very elderly rodents and humans, and were robust immunohistochemical markers of human cirrhosis and HCC. In response to the chemotherapeutic and DNA-demethylating agent 5-aza-deoxycytidine (5-aza-dC), transgenic expression of macroH2A1 isoforms in HCC cell lines prevented the emergence of a senescent-like phenotype and induced synergistic global DNA hypomethylation. Conversely, macroH2A1 depletion amplified the antiproliferative effects of 5-aza-dC in HCC cells, but failed to enhance senescence. Senescence-associated secretory phenotype and whole-transcriptome analyses implicated the p38 MAPK/IL8 pathway in mediating macroH2A1-dependent escape of HCC cells from chemotherapy-induced senescence. Furthermore, chromatin immunoprecipitation sequencing revealed that this hepatic antisenescence state also required active transcription that could not be attributed to genomic occupancy of these histones. Collectively, our findings reveal a new mechanism by which drug-induced senescence is epigenetically regulated by macroH2A1 and DNA methylation and suggest macroH2A1 as a novel biomarker of hepatic senescence that could potentially predict prognosis and disease progression. ©2016 American Association for Cancer Research.

  17. Histone H3 Variant Regulates RNA Polymerase II Transcription Termination and Dual Strand Transcription of siRNA Loci in Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    David Reynolds

    2016-01-01

    Full Text Available Base J, β-D-glucosyl-hydroxymethyluracil, is a chromatin modification of thymine in the nuclear DNA of flagellated protozoa of the order Kinetoplastida. In Trypanosoma brucei, J is enriched, along with histone H3 variant (H3.V, at sites involved in RNA Polymerase (RNAP II termination and telomeric sites involved in regulating variant surface glycoprotein gene (VSG transcription by RNAP I. Reduction of J in T. brucei indicated a role of J in the regulation of RNAP II termination, where the loss of J at specific sites within polycistronic gene clusters led to read-through transcription and increased expression of downstream genes. We now demonstrate that the loss of H3.V leads to similar defects in RNAP II termination within gene clusters and increased expression of downstream genes. Gene derepression is intensified upon the subsequent loss of J in the H3.V knockout. mRNA-seq indicates gene derepression includes VSG genes within the silent RNAP I transcribed telomeric gene clusters, suggesting an important role for H3.V in telomeric gene repression and antigenic variation. Furthermore, the loss of H3.V at regions of overlapping transcription at the end of convergent gene clusters leads to increased nascent RNA and siRNA production. Our results suggest base J and H3.V can act independently as well as synergistically to regulate transcription termination and expression of coding and non-coding RNAs in T. brucei, depending on chromatin context (and transcribing polymerase. As such these studies provide the first direct evidence for histone H3.V negatively influencing transcription elongation to promote termination.

  18. DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression

    Science.gov (United States)

    Borghesan, Michela; Fusilli, Caterina; Rappa, Francesca; Panebianco, Concetta; Rizzo, Giovanni; Oben, Jude A.; Mazzoccoli, Gianluigi; Faulkes, Chris; Pata, Illar; Agodi, Antonella; Rezaee, Farhad; Minogue, Shane; Warren, Alessandra; Peterson, Abigail; Sedivy, John M.; Douet, Julien; Buschbeck, Marcus; Cappello, Francesco; Mazza, Tommaso; Pazienza, Valerio; Vinciguerra, Manlio

    2016-01-01

    Aging is a major risk factor for progression of liver diseases to hepatocellular carcinoma (HCC). Cellular senescence contributes to age-related tissue dysfunction, but the epigenetic basis underlying drug-induced senescence remains unclear.macroH2A1, a variant of histone H2A, is a marker of senescence-associated heterochromatic foci that synergizes with DNA methylation to silence tumor-suppressor genes in human fibroblasts. In this study, we investigated the relationship between macroH2A1 splice variants, macroH2A1.1 and macroH2A1.2, and liver carcinogenesis. We found that protein levels of both macroH2A1 isoforms were increased in the livers of very elderly rodents and humans, and were robust immunohistochemical markers of human cirrhosis and HCC. In response to the chemotherapeutic and DNA-demethylating agent 5-aza-deoxycytidine (5-aza-dC), transgenic expression of macroH2A1 isoforms in HCC cell lines prevented the emergence of a senescent-like phenotype and induced synergistic global DNA hypomethylation. Conversely, macroH2A1 depletion amplified the antiproliferative effects of 5-aza-dC in HCC cells, but failed to enhance senescence. Senescence-associated secretory phenotype and whole-transcriptome analyses implicated the p38 MAPK/IL8 pathway in mediating macroH2A1-dependent escape of HCC cells from chemotherapy-induced senescence. Furthermore, chromatin immunoprecipitation sequencing revealed that this hepatic antisenescence state also required active transcription that could not be attributed to genomic occupancy of these histones. Collectively, our findings reveal a new mechanism by which drug-induced senescence is epigenetically regulated by macroH2A1 and DNA methylation and suggest macroH2A1 as a novel biomarker of hepatic senescence that could potentially predict prognosis and disease progression. PMID:26772755

  19. Intra- and inter-nucleosomal interactions of the histone H4 tail revealed with a human nucleosome core particle with genetically-incorporated H4 tetra-acetylation.

    Science.gov (United States)

    Wakamori, Masatoshi; Fujii, Yoshifumi; Suka, Noriyuki; Shirouzu, Mikako; Sakamoto, Kensaku; Umehara, Takashi; Yokoyama, Shigeyuki

    2015-11-26

    Post-translational modifications (PTMs) of histones, such as lysine acetylation of the N-terminal tails, play crucial roles in controlling gene expression. Due to the difficulty in reconstituting site-specifically acetylated nucleosomes with crystallization quality, structural analyses of histone acetylation are currently performed using synthesized tail peptides. Through engineering of the genetic code, translation termination, and cell-free protein synthesis, we reconstituted human H4-mono- to tetra-acetylated nucleosome core particles (NCPs), and solved the crystal structures of the H4-K5/K8/K12/K16-tetra-acetylated NCP and unmodified NCP at 2.4 Å and 2.2 Å resolutions, respectively. The structure of the H4-tetra-acetylated NCP resembled that of the unmodified NCP, and the DNA wrapped the histone octamer as precisely as in the unmodified NCP. However, the B-factors were significantly increased for the peripheral DNAs near the N-terminal tail of the intra- or inter-nucleosomal H4. In contrast, the B-factors were negligibly affected by the H4 tetra-acetylation in histone core residues, including those composing the acidic patch, and at H4-R23, which interacts with the acidic patch of the neighboring NCP. The present study revealed that the H4 tetra-acetylation impairs NCP self-association by changing the interactions of the H4 tail with DNA, and is the first demonstration of crystallization quality NCPs reconstituted with genuine PTMs.

  20. Telomeric heterochromatin boundaries require NuA4-dependent acetylation of histone variant H2A.Z in Saccharomycescerevisiae

    Science.gov (United States)

    Babiarz, Joshua E.; Halley, Jeffrey E.; Rine, Jasper

    2006-01-01

    SWR1-Com, which is responsible for depositing H2A.Z into chromatin, shares four subunits with the NuA4 histone acetyltransferase complex. This overlap in composition led us to test whether H2A.Z was a substrate of NuA4 in vitro and in vivo. The N-terminal tail of H2A.Z was acetylated in vivo at multiple sites by a combination of NuA4 and SAGA. H2A.Z acetylation was also dependent on SWR1-Com, causing H2A.Z to be efficiently acetylated only when incorporated in chromatin. Unacetylatable H2A.Z mutants were, like wild-type H2A.Z, enriched at heterochromatin boundaries, but were unable to block spreading of heterochromatin. A mutant version of H2A.Z that could not be acetylated, in combination with a mutation in a nonessential gene in the NuA4 complex, caused a pronounced decrease in growth rate. This H2A.Z mutation was lethal in combination with a mutant version of histone H4 that could not be acetylated by NuA4. Taken together, these results show a role for H2A.Z acetylation in restricting silent chromatin, and reveal that acetylation of H2A.Z and H4 can contribute to a common function essential to life. PMID:16543222

  1. A moist aquaplanet variant of the Held-Suarez test for atmospheric model dynamical cores

    Science.gov (United States)

    Thatcher, Diana R.; Jablonowski, Christiane

    2016-04-01

    A moist idealized test case (MITC) for atmospheric model dynamical cores is presented. The MITC is based on the Held-Suarez (HS) test that was developed for dry simulations on "a flat Earth" and replaces the full physical parameterization package with a Newtonian temperature relaxation and Rayleigh damping of the low-level winds. This new variant of the HS test includes moisture and thereby sheds light on the nonlinear dynamics-physics moisture feedbacks without the complexity of full-physics parameterization packages. In particular, it adds simplified moist processes to the HS forcing to model large-scale condensation, boundary-layer mixing, and the exchange of latent and sensible heat between the atmospheric surface and an ocean-covered planet. Using a variety of dynamical cores of the National Center for Atmospheric Research (NCAR)'s Community Atmosphere Model (CAM), this paper demonstrates that the inclusion of the moist idealized physics package leads to climatic states that closely resemble aquaplanet simulations with complex physical parameterizations. This establishes that the MITC approach generates reasonable atmospheric circulations and can be used for a broad range of scientific investigations. This paper provides examples of two application areas. First, the test case reveals the characteristics of the physics-dynamics coupling technique and reproduces coupling issues seen in full-physics simulations. In particular, it is shown that sudden adjustments of the prognostic fields due to moist physics tendencies can trigger undesirable large-scale gravity waves, which can be remedied by a more gradual application of the physical forcing. Second, the moist idealized test case can be used to intercompare dynamical cores. These examples demonstrate the versatility of the MITC approach and suggestions are made for further application areas. The new moist variant of the HS test can be considered a test case of intermediate complexity.

  2. A Ribbon-like Structure in the Ejective Organelle of the Green Microalga Pyramimonas parkeae (Prasinophyceae) Consists of Core Histones and Polymers Containing N-acetyl-glucosamine.

    Science.gov (United States)

    Yamagishi, Takahiro; Kurihara, Akira; Kawai, Hiroshi

    2015-11-01

    The green microalga, Pyramimonas parkeae (Prasinophyceae) has an ejective organelle containing a coiled ribbon structure resembling the ejectisome in Cryptophyta. This structure is discharged from the cell by a stimulus and extends to form a tube-like structure, but the molecular components of the structure have not been identified. Tricine-SDS-PAGE analysis indicated that the ribbon-like structure of P. parkeae contains some proteins and low molecular acidic polymers. Edman degradation, LC/MS/MS analyses and immunological studies demonstrated that their proteins are core histones (H3, H2A, H2B and H4). In addition, monosaccharide composition analysis of the ribbon-like structures and degradation by lysozyme strongly indicated that the ribbon-like structure consist of β (1-4) linked polymers containing N-acetyl-glucosamine. Purified polymers and recombinant histones formed glob-like or filamentous structures. Therefore we conclude that the ribbon-like structure of P. parkeae mainly consists of a complex of core histones (H3, H2A, H2B and H4) and polymers containing N-acetyl-glucosamine, and suggest to name the ejective organelle in P. parkeae the "histrosome" to distinguish it from the ejectisome in Cryptophyta. Copyright © 2015 Elsevier GmbH. All rights reserved.

  3. Comparative Assessment of Mediterranean Gorgonian-Associated Microbial Communities Reveals Conserved Core and Locally Variant Bacteria

    KAUST Repository

    van de Water, Jeroen A J M

    2016-10-10

    Gorgonians are key habitat-forming species of Mediterranean benthic communities, but their populations have suffered from mass mortality events linked to high summer seawater temperatures and microbial disease. However, our knowledge on the diversity, dynamics and function of gorgonian-associated microbial communities is limited. Here, we analysed the spatial variability of the microbiomes of five sympatric gorgonian species (Eunicella singularis, Eunicella cavolini, Eunicella verrucosa, Leptogorgia sarmentosa and Paramuricea clavata), collected from the Mediterranean Sea over a scale of ∼1100 km, using next-generation amplicon sequencing of the 16S rRNA gene. The microbiomes of all gorgonian species were generally dominated by members of the genus Endozoicomonas, which were at very low abundance in the surrounding seawater. Although the composition of the core microbiome (operational taxonomic units consistently present in a species) was found to be unique for each host species, significant overlap was observed. These spatially consistent associations between gorgonians and their core bacteria suggest intricate symbiotic relationships and regulation of the microbiome composition by the host. At the same time, local variations in microbiome composition were observed. Functional predictive profiling indicated that these differences could be attributed to seawater pollution. Taken together, our data indicate that gorgonian-associated microbiomes are composed of spatially conserved bacteria (core microbiome members) and locally variant members, and that local pollution may influence these local associations, potentially impacting gorgonian health.

  4. Co-expression as a convenient method for the production and purification of core histones in bacteria.

    Science.gov (United States)

    Anderson, Megan; Huh, Joon H; Ngo, Thien; Lee, Alice; Hernandez, Genaro; Pang, Joy; Perkins, Jennifer; Dutnall, Robert N

    2010-08-01

    Co-expression offers an important strategy for producing multiprotein complexes for biochemical and biophysical studies. We have found that co-expression of histones H2A and H2B (from yeast, chicken or Drosophila) leads to production of soluble heterodimeric H2AH2B complexes. Drosophila histones H3 and H4 can also be produced as a soluble (H3H4)(2) heterotetrameric complex if they are co-expressed with the histone chaperone Asf1. The soluble H2AH2B and (H3H4)(2) can be purified by simple chromatographic techniques and have similar properties to endogenous histones. Our methods should facilitate histone production for studies of chromatin structure and regulatory proteins that interact with histones. We describe a simple strategy for constructing co-expression plasmids, based on the T7 RNA polymerase system, which is applicable to other systems. It offers several advantages for quickly creating plasmids to express two or more proteins and for testing different combinations of proteins for optimal complex production, solubility or activity. Copyright 2010 Elsevier Inc. All rights reserved.

  5. Human GRK4γ142V Variant Promotes Angiotensin II Type I Receptor-Mediated Hypertension via Renal Histone Deacetylase Type 1 Inhibition.

    Science.gov (United States)

    Wang, Zheng; Zeng, Chunyu; Villar, Van Anthony M; Chen, Shi-You; Konkalmatt, Prasad; Wang, Xiaoyan; Asico, Laureano D; Jones, John E; Yang, Yu; Sanada, Hironobu; Felder, Robin A; Eisner, Gilbert M; Weir, Matthew R; Armando, Ines; Jose, Pedro A

    2016-02-01

    The influence of a single gene on the pathogenesis of essential hypertension may be difficult to ascertain, unless the gene interacts with other genes that are germane to blood pressure regulation. G-protein-coupled receptor kinase type 4 (GRK4) is one such gene. We have reported that the expression of its variant hGRK4γ(142V) in mice results in hypertension because of impaired dopamine D1 receptor. Signaling through dopamine D1 receptor and angiotensin II type I receptor (AT1R) reciprocally modulates renal sodium excretion and blood pressure. Here, we demonstrate the ability of the hGRK4γ(142V) to increase the expression and activity of the AT1R. We show that hGRK4γ(142V) phosphorylates histone deacetylase type 1 and promotes its nuclear export to the cytoplasm, resulting in increased AT1R expression and greater pressor response to angiotensin II. AT1R blockade and the deletion of the Agtr1a gene normalize the hypertension in hGRK4γ(142V) mice. These findings illustrate the unique role of GRK4 by targeting receptors with opposite physiological activity for the same goal of maintaining blood pressure homeostasis, and thus making the GRK4 a relevant therapeutic target to control blood pressure.

  6. Mass Spectrometric Analysis of Histone Proteoforms

    Science.gov (United States)

    Yuan, Zuo-Fei; Arnaudo, Anna M.; Garcia, Benjamin A.

    2014-06-01

    Histones play important roles in chromatin, in the forms of various posttranslational modifications (PTMs) and sequence variants, which are called histone proteoforms. Investigating modifications and variants is an ongoing challenge. Previous methods are based on antibodies, and because they usually detect only one modification at a time, they are not suitable for studying the various combinations of modifications on histones. Fortunately, mass spectrometry (MS) has emerged as a high-throughput technology for histone analysis and does not require prior knowledge about any modifications. From the data generated by mass spectrometers, both identification and quantification of modifications, as well as variants, can be obtained easily. On the basis of this information, the functions of histones in various cellular contexts can be revealed. Therefore, MS continues to play an important role in the study of histone proteoforms. In this review, we discuss the analysis strategies of MS, their applications on histones, and some key remaining challenges.

  7. Rapid purification of recombinant histones.

    Directory of Open Access Journals (Sweden)

    Henrike Klinker

    Full Text Available The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants.

  8. A genetic system to assess in vivo the functions of histones and histone modifications in higher eukaryotes.

    Science.gov (United States)

    Günesdogan, Ufuk; Jäckle, Herbert; Herzig, Alf

    2010-10-01

    Despite the fundamental role of canonical histones in nucleosome structure, there is no experimental system for higher eukaryotes in which basic questions about histone function can be directly addressed. We developed a new genetic tool for Drosophila melanogaster in which the canonical histone complement can be replaced with multiple copies of experimentally modified histone transgenes. This new histone-replacement system provides a well-defined and direct cellular assay system for histone function with which to critically test models in chromatin biology dealing with chromatin assembly, variant histone functions and the biological significance of distinct histone modifications in a multicellular organism.

  9. An age of fewer histones.

    Science.gov (United States)

    Oberdoerffer, Philipp

    2010-11-01

    Changes in chromatin structure are a conserved hallmark of ageing, and the mechanism driving these changes, as well as their functional significance, are heavily investigated. Loss of core histones is now observed in aged cells and may contribute to this phenomenon. Histone loss is coupled to cell division and seems to be triggered by telomeric DNA damage.

  10. Structure and Functions of Linker Histones.

    Science.gov (United States)

    Lyubitelev, A V; Nikitin, D V; Shaytan, A K; Studitsky, V M; Kirpichnikov, M P

    2016-03-01

    Linker histones such as variants H1, H5, and other similar proteins play an important role in regulation of chromatin structure and dynamics. However, interactions of linker histones with DNA and proteins, as well as specific functions of their different variants, are poorly studied. This is because they acquire tertiary structure only when interacting with a nucleosome, and because of limitations of currently available methods. However, deeper investigation of linker histones and their interactions with other proteins will address a number of important questions - from structure of compacted chromatin to regulation of early embryogenesis. In this review, structures of histone H1 variants and its interaction with chromatin DNA are considered. A possible functional significance of different H1 variants, a role of these proteins in maintaining interphase chromatin structure, and interactions of linker histones with other cellular proteins are also discussed.

  11. Chromosomal mapping of rRNA genes, core histone genes and telomeric sequences in Brachidontes puniceus and Brachidontes rodriguezi (Bivalvia, Mytilidae

    Directory of Open Access Journals (Sweden)

    Pasantes Juan J

    2010-12-01

    Full Text Available Abstract Background Chromosome rearrangements are an important part of the speciation process in many taxa. The study of chromosome evolution in bivalves is hampered by the absence of clear chromosomal banding patterns and the similarity in both chromosome size and morphology. For this reason, obtaining good chromosome markers is essential for reliable karyotypic comparisons. To begin this task, the chromosomes of the mussels Brachidontes puniceus and B. rodriguezi were studied by means of fluorochrome staining and fluorescent in situ hybridization (FISH. Results Brachidontes puniceus and B. rodriguezi both have 2n = 32 chromosomes but differing karyotype composition. Vertebrate-type telomeric sequences appear at both ends of every single chromosome. B. puniceus presents a single terminal major rRNA gene cluster on a chromosome pair while B. rodriguezi shows two. Both mussels present two 5S rDNA and two core histone gene clusters intercalary located on the long arms of two chromosome pairs. Double and triple-FISH experiments demonstrated that one of the 5S rDNA and one of the major rDNA clusters appear on the same chromosome pair in B. rodriguezi but not in B. puniceus. On the other hand, the second 5S rDNA cluster is located in one of the chromosome pairs also bearing one of the core histone gene clusters in the two mussel species. Conclusion Knowledge of the chromosomal distribution of these sequences in the two species of Brachidontes is a first step in the understanding of the role of chromosome changes on bivalve evolution.

  12. Implication of Posttranslational Histone Modifications in Nucleotide Excision Repair

    Directory of Open Access Journals (Sweden)

    Shisheng Li

    2012-09-01

    Full Text Available Histones are highly alkaline proteins that package and order the DNA into chromatin in eukaryotic cells. Nucleotide excision repair (NER is a conserved multistep reaction that removes a wide range of generally bulky and/or helix-distorting DNA lesions. Although the core biochemical mechanism of NER is relatively well known, how cells detect and repair lesions in diverse chromatin environments is still under intensive research. As with all DNA-related processes, the NER machinery must deal with the presence of organized chromatin and the physical obstacles it presents. A huge catalogue of posttranslational histone modifications has been documented. Although a comprehensive understanding of most of these modifications is still lacking, they are believed to be important regulatory elements for many biological processes, including DNA replication and repair, transcription and cell cycle control. Some of these modifications, including acetylation, methylation, phosphorylation and ubiquitination on the four core histones (H2A, H2B, H3 and H4 or the histone H2A variant H2AX, have been found to be implicated in different stages of the NER process. This review will summarize our recent understanding in this area.

  13. Lysine residues in N-terminal and C-terminal regions of human histone H2A are targets for biotinylation by biotinidase.

    Science.gov (United States)

    Chew, Yap Ching; Camporeale, Gabriela; Kothapalli, Nagarama; Sarath, Gautam; Zempleni, Janos

    2006-04-01

    In eukaryotic cell nuclei, DNA associates with the core histones H2A, H2B, H3 and H4 to form nucleosomal core particles. DNA binding to histones is regulated by posttranslational modifications of N-terminal tails (e.g., acetylation and methylation of histones). These modifications play important roles in the epigenetic control of chromatin structure. Recently, evidence that biotinidase and holocarboxylase synthetase (HCS) catalyze the covalent binding of biotin to histones has been provided. The primary aim of this study was to identify biotinylation sites in histone H2A and its variant H2AX. Secondary aims were to determine whether acetylation and methylation of histone H2A affect subsequent biotinylation and whether biotinidase and HCS localize to the nucleus in human cells. Biotinylation sites were identified using synthetic peptides as substrates for biotinidase. These studies provided evidence that K9 and K13 in the N-terminus of human histones H2A and H2AX are targets for biotinylation and that K125, K127 and K129 in the C-terminus of histone H2A are targets for biotinylation. Biotinylation of lysine residues was decreased by acetylation of adjacent lysines but was increased by dimethylation of adjacent arginines. The existence of biotinylated histone H2A in vivo was confirmed by using modification-specific antibodies. Antibodies to biotinidase and HCS localized primarily to the nuclear compartment, consistent with a role for these enzymes in regulating chromatin structure. Collectively, these studies have identified five novel biotinylation sites in human histones; histone H2A is unique among histones in that its biotinylation sites include amino acid residues from the C-terminus.

  14. Structures of oncogenic, suppressor and rescued p53 core-domain variants: mechanisms of mutant p53 rescue

    Energy Technology Data Exchange (ETDEWEB)

    Wallentine, Brad D.; Wang, Ying; Tretyachenko-Ladokhina, Vira; Tan, Martha; Senear, Donald F. [University of California, Irvine, Irvine, CA 92697 (United States); Luecke, Hartmut, E-mail: hudel@uci.edu [University of California, Irvine, Irvine, CA 92697 (United States); University of California, Irvine, Irvine, CA 92697 (United States); University of California, Irvine, Irvine, CA 92697 (United States); University of California, Irvine, Irvine, CA 92697 (United States); Universidad del Pais Vasco, 48940 Leioa (Spain)

    2013-10-01

    X-ray crystallographic structures of four p53 core-domain variants were determined in order to gain insights into the mechanisms by which certain second-site suppressor mutations rescue the function of a significant number of cancer mutations of the tumor suppressor protein p53. To gain insights into the mechanisms by which certain second-site suppressor mutations rescue the function of a significant number of cancer mutations of the tumor suppressor protein p53, X-ray crystallographic structures of four p53 core-domain variants were determined. These include an oncogenic mutant, V157F, two single-site suppressor mutants, N235K and N239Y, and the rescued cancer mutant V157F/N235K/N239Y. The V157F mutation substitutes a smaller hydrophobic valine with a larger hydrophobic phenylalanine within strand S4 of the hydrophobic core. The structure of this cancer mutant shows no gross structural changes in the overall fold of the p53 core domain, only minor rearrangements of side chains within the hydrophobic core of the protein. Based on biochemical analysis, these small local perturbations induce instability in the protein, increasing the free energy by 3.6 kcal mol{sup −1} (15.1 kJ mol{sup −1}). Further biochemical evidence shows that each suppressor mutation, N235K or N239Y, acts individually to restore thermodynamic stability to V157F and that both together are more effective than either alone. All rescued mutants were found to have wild-type DNA-binding activity when assessed at a permissive temperature, thus pointing to thermodynamic stability as the critical underlying variable. Interestingly, thermodynamic analysis shows that while N239Y demonstrates stabilization of the wild-type p53 core domain, N235K does not. These observations suggest distinct structural mechanisms of rescue. A new salt bridge between Lys235 and Glu198, found in both the N235K and rescued cancer mutant structures, suggests a rescue mechanism that relies on stabilizing the

  15. A histone timer for zygotic genome activation.

    Science.gov (United States)

    Siriaco, Giorgia; Tamkun, John W

    2013-09-30

    Histone H1 variants play key roles in the regulation of higher-order chromatin structure and have been implicated in numerous developmental processes. In this issue of Developmental Cell, Pérez-Montero et al. (2013) present evidence that the Drosophila histone H1 variant dBigH1 prevents premature activation of the zygotic genome during early embryogenesis. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. Specificities and genomic distribution of somatic mammalian histone H1 subtypes.

    Science.gov (United States)

    Millán-Ariño, Lluís; Izquierdo-Bouldstridge, Andrea; Jordan, Albert

    2016-03-01

    Histone H1 is a structural component of chromatin that may have a role in the regulation of chromatin dynamics. Unlike core histones, the linker histone H1 family is evolutionarily diverse and many organisms have multiple H1 variants or subtypes, distinguishable between germ-line and somatic cells. In mammals, the H1 family includes seven somatic H1 variants with a prevalence that varies between cell types and over the course of differentiation, H1.1 to H1.5 being expressed in a replication-dependent manner, whereas H1.0 and H1X are replication-independent. Until recently, it has not been known whether the different variants had specific roles in the regulation of nuclear processes or were differentially distributed across the genome. To address this, an increasing effort has been made to investigate divergent features among H1 variants, regarding their structure, expression patterns, chromatin dynamics, post-translational modifications and genome-wide distribution. Although H1 subtypes seem to have redundant functions, several reports point to the idea that they are also differently involved in specific cellular processes. Initial studies investigating the genomic distribution of H1 variants have started to suggest that despite a wide overlap, different variants may be enriched or preferentially located at different chromatin types, but this may depend on the cell type, the relative abundance of the variants, the differentiation state of the cell, or whether cells are derived from a neoplastic process. Understanding the heterogeneity of the histone H1 family is crucial to elucidate their role in chromatin organization, gene expression regulation and other cellular processes.

  17. Open and Closed: The Roles of Linker Histones in Plants and Animals

    OpenAIRE

    Over, Ryan S.; Michaels, Scott D.

    2013-01-01

    Linker histones play key roles alongside core histones in the regulation and maintenance of chromatin. Here, we illustrate our current understanding of the contributions of linker histones to the cell cycle, development, and chromatin structure in plants and animals.

  18. Absolute quantification of acetylation and phosphorylation of the histone variant H2AX upon ionizing radiation reveals distinct cellular responses in two cancer cell lines.

    Science.gov (United States)

    Matsuda, Shun; Furuya, Kanji; Ikura, Masae; Matsuda, Tomonari; Ikura, Tsuyoshi

    2015-11-01

    Histone modifications change upon the cellular response to ionizing radiation, and their cellular amounts could reflect the DNA damage response activity. We previously reported a sensitive and reliable method for the absolute quantification of γH2AX within cells, using liquid chromatography-tandem mass spectrometry (LC/MS/MS). The technique has broad adaptability to a variety of biological systems and can quantitate different modifications of histones. In this study, we applied it to quantitate the levels of γH2AX and K5-acetylated H2AX, and to compare the radiation responses between two cancer cell lines: HeLa and U-2 OS. The two cell lines have distinct properties in terms of their H2AX modifications. HeLa cells have relatively high γH2AX (3.1 %) against the total H2AX even in un-irradiated cells, while U-2 OS cells have an essentially undetectable level (nearly 0 %) of γH2AX. In contrast, the amounts of acetylated histones are lower in HeLa cells (9.3 %) and higher in U-2 OS cells (24.2 %) under un-irradiated conditions. Furthermore, after ionizing radiation exposure, the time-dependent increases and decreases in the amounts of histone modifications differed between the two cell lines, especially at the early time points. These results suggest that each biological system has distinct kinase/phosphatase and/or acetylase/deacetylase activities. In conclusion, for the first time, we have succeeded in simultaneously monitoring the absolute amounts of phosphorylated and acetylated cellular H2AX after ionizing radiation exposure. This multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems.

  19. Histones and genome integrity.

    Science.gov (United States)

    Williamson, Wes D; Pinto, Ines

    2012-01-01

    Chromosomes undergo extensive structural rearrangements during the cell cycle, from the most open chromatin state required for DNA replication to the highest level of compaction and condensation essential for mitotic segregation of sister chromatids. It is now widely accepted that chromatin is a highly dynamic structure that participates in all DNA-related functions, including transcription, DNA replication, repair, and mitosis; hence, histones have emerged as key players in these cellular processes. We review here the studies that implicate histones in functions that affect the chromosome cycle, defined as the cellular processes involved in the maintenance, replication, and segregation of chromosomal DNA. Disruption of the chromosome cycle affects the integrity of the cellular genome, leading to aneuploidy, polyploidy or cell death. Histone stoichiometry, mutations that affect the structure of the nucleosome core particle, and mutations that affect the structure and/or modifications of the histone tails, all have a direct impact on the fidelity of chromosome transmission and the integrity of the genome.

  20. The differential mobilization of histones H3.1 and H3.3 by herpes simplex virus 1 relates histone dynamics to the assembly of viral chromatin.

    Science.gov (United States)

    Conn, Kristen L; Hendzel, Michael J; Schang, Luis M

    2013-01-01

    During lytic infections, HSV-1 genomes are assembled into unstable nucleosomes. The histones required for HSV-1 chromatin assembly, however, are in the cellular chromatin. We have shown that linker (H1) and core (H2B and H4) histones are mobilized during HSV-1 infection, and proposed that the mobilized histones are available for assembly into viral chromatin. However, the actual relevance of histone mobilization remained unknown. We now show that canonical H3.1 and variant H3.3 are also mobilized during HSV-1 infection. Mobilization required no HSV-1 protein expression, although immediate early or early proteins enhanced it. We used the previously known differential association of H3.3 and H3.1 with HSV-1 DNA to test the relevance of histone mobilization. H3.3 binds to HSV-1 genomes first, whereas H3.1 only binds after HSV-1 DNA replication initiates. Consistently, H3.3 and H3.1 were differentially mobilized. H3.1 mobilization decreased with HSV-1 DNA replication, whereas H3.3 mobilization was largely unaffected by it. These results support a model in which previously mobilized H3.1 is immobilized by assembly into viral chromatin during HSV-1 DNA replication, whereas H3.3 is mobilized and assembled into HSV-1 chromatin throughout infection. The differential mobilizations of H3.3 and H3.1 are consistent with their differential assembly into viral chromatin. These data therefore relate nuclear histone dynamics to the composition of viral chromatin and provide the first evidence that histone mobilization relates to viral chromatin assembly.

  1. The evolutionary history of histone H3 suggests a deep eukaryotic root of chromatin modifying mechanisms

    Directory of Open Access Journals (Sweden)

    Postberg Jan

    2010-08-01

    Full Text Available Abstract Background The phenotype of an organism is an outcome of both its genotype, encoding the primary sequence of proteins, and the developmental orchestration of gene expression. The substrate of gene expression in eukaryotes is the chromatin, whose fundamental units are nucleosomes composed of DNA wrapped around each two of the core histone types H2A, H2B, H3 and H4. Key regulatory steps involved in the determination of chromatin conformations are posttranslational modifications (PTM at histone tails as well as the assembly of histone variants into nucleosomal arrays. Although the mechanistic background is fragmentary understood, it appears that the chromatin signature of metazoan cell types is inheritable over generations. Even less understood is the conservation of epigenetic mechanisms among eukaryotes and their origins. Results In the light of recent progress in understanding the tree of eukaryotic life we discovered the origin of histone H3 by phylogenetic analyses of variants from all supergroups, which allowed the reconstruction of ancestral states. We found that H3 variants evolved frequently but independently within related species of almost all eukaryotic supergroups. Interestingly, we found all core histone types encoded in the genome of a basal dinoflagellate and H3 variants in two other species, although is was reported that dinoflagellate chromatin is not organized into nucleosomes. Most probably one or more animal/nuclearid H3.3-like variants gave rise to H3 variants of all opisthokonts (animals, choanozoa, fungi, nuclearids, Amoebozoa. H3.2 and H3.1 as well as H3.1t are derivatives of H3.3, whereas H3.2 evolved already in early branching animals, such as Trichoplax. H3.1 and H3.1t are probably restricted to mammals. We deduced a model for protoH3 of the last eukaryotic common ancestor (LECA confirming a remarkable degree of sequence conservation in comparison to canonical human H3.1. We found evidence that multiple PTMs are

  2. The histone chaperones Nap1 and Vps75 bind histones H3 and H4 in a tetrameric conformation.

    Science.gov (United States)

    Bowman, Andrew; Ward, Richard; Wiechens, Nicola; Singh, Vijender; El-Mkami, Hassane; Norman, David George; Owen-Hughes, Tom

    2011-02-18

    Histone chaperones physically interact with histones to direct proper assembly and disassembly of nucleosomes regulating diverse nuclear processes such as DNA replication, promoter remodeling, transcription elongation, DNA damage, and histone variant exchange. Currently, the best-characterized chaperone-histone interaction is that between the ubiquitous chaperone Asf1 and a dimer of H3 and H4. Nucleosome assembly proteins (Nap proteins) represent a distinct class of histone chaperone. Using pulsed electron double resonance (PELDOR) measurements and protein crosslinking, we show that two members of this class, Nap1 and Vps75, bind histones in the tetrameric conformation also observed when they are sequestered within the nucleosome. Furthermore, H3 and H4 trapped in their tetrameric state can be used as substrates in nucleosome assembly and chaperone-mediated lysine acetylation. This alternate mode of histone interaction provides a potential means of maintaining the integrity of the histone tetramer during cycles of nucleosome reassembly.

  3. Characterization of yeast histone H3-specific type B histone acetyltransferases identifies an ADA2-independent Gcn5p activity

    Directory of Open Access Journals (Sweden)

    Parthun Mark R

    2004-07-01

    Full Text Available Abstract Background The acetylation of the core histone NH2-terminal tails is catalyzed by histone acetyltransferases. Histone acetyltransferases can be classified into two distinct groups (type A and B on the basis of cellular localization and substrate specificity. Type B histone acetyltransferases, originally defined as cytoplasmic enzymes that acetylate free histones, have been proposed to play a role in the assembly of chromatin through the acetylation of newly synthesized histones H3 and H4. To date, the only type B histone acetyltransferase activities identified are specific for histone H4. Results To better understand the role of histone acetylation in the assembly of chromatin structure, we have identified additional type B histone acetyltransferase activities specific for histone H3. One such activity, termed HatB3.1, acetylated histone H3 with a strong preference for free histones relative to chromatin substrates. Deletion of the GCN5 and ADA3 genes resulted in the loss of HatB3.1 activity while deletion of ADA2 had no effect. In addition, Gcn5p and Ada3p co-fractionated with partially purified HatB3.1 activity while Ada2p did not. Conclusions Yeast extracts contain several histone acetyltransferase activities that show a strong preference for free histone H3. One such activity, termed HatB3.1, appears to be a novel Gcn5p-containing complex which does not depend on the presence of Ada2p.

  4. Drosophila TAP/p32 is a core histone chaperone that cooperates with NAP-1, NLP, and nucleophosmin in sperm chromatin remodeling during fertilization.

    Science.gov (United States)

    Emelyanov, Alexander V; Rabbani, Joshua; Mehta, Monika; Vershilova, Elena; Keogh, Michael C; Fyodorov, Dmitry V

    2014-09-15

    Nuclear DNA in the male gamete of sexually reproducing animals is organized as sperm chromatin compacted primarily by sperm-specific protamines. Fertilization leads to sperm chromatin remodeling, during which protamines are expelled and replaced by histones. Despite our increased understanding of the factors that mediate nucleosome assembly in the nascent male pronucleus, the machinery for protamine removal remains largely unknown. Here we identify four Drosophila protamine chaperones that mediate the dissociation of protamine-DNA complexes: NAP-1, NLP, and nucleophosmin are previously characterized histone chaperones, and TAP/p32 has no known function in chromatin metabolism. We show that TAP/p32 is required for the removal of Drosophila protamine B in vitro, whereas NAP-1, NLP, and Nph share roles in the removal of protamine A. Embryos from P32-null females show defective formation of the male pronucleus in vivo. TAP/p32, similar to NAP-1, NLP, and Nph, facilitates nucleosome assembly in vitro and is therefore a histone chaperone. Furthermore, mutants of P32, Nlp, and Nph exhibit synthetic-lethal genetic interactions. In summary, we identified factors mediating protamine removal from DNA and reconstituted in a defined system the process of sperm chromatin remodeling that exchanges protamines for histones to form the nucleosome-based chromatin characteristic of somatic cells.

  5. Histone deacetylases and atherosclerosis.

    Science.gov (United States)

    Zheng, Xia-xia; Zhou, Tian; Wang, Xin-An; Tong, Xiao-hong; Ding, Jia-wang

    2015-06-01

    Atherosclerosis is the most common pathological process that leads to cardiovascular diseases, a disease of large- and medium-sized arteries that is characterized by a formation of atherosclerotic plaques consisting of necrotic cores, calcified regions, accumulated modified lipids, smooth muscle cells (SMCs), endothelial cells, leukocytes, and foam cells. Recently, the question about how to suppress the occurrence of atherosclerosis and alleviate the progress of cardiovascular disease becomes the hot topic. Accumulating evidence suggests that histone deacetylases(HDACs) play crucial roles in arteriosclerosis. This review summarizes the effect of HDACs and HDAC inhibitors(HDACi) on the progress of atherosclerosis.

  6. Histone displacement during nucleotide excision repair

    DEFF Research Database (Denmark)

    Dinant, C.; Bartek, J.; Bekker-Jensen, S.

    2012-01-01

    Nucleotide excision repair (NER) is an important DNA repair mechanism required for cellular resistance against UV light and toxic chemicals such as those found in tobacco smoke. In living cells, NER efficiently detects and removes DNA lesions within the large nuclear macromolecular complex called...... of histone variants and histone displacement (including nucleosome sliding). Here we review current knowledge, and speculate about current unknowns, regarding those chromatin remodeling activities that physically displace histones before, during and after NER. © 2012 by the authors; licensee MDPI, Basel...

  7. Histone H1 Limits DNA Methylation in Neurospora crassa.

    Science.gov (United States)

    Seymour, Michael; Ji, Lexiang; Santos, Alex M; Kamei, Masayuki; Sasaki, Takahiko; Basenko, Evelina Y; Schmitz, Robert J; Zhang, Xiaoyu; Lewis, Zachary A

    2016-07-07

    Histone H1 variants, known as linker histones, are essential chromatin components in higher eukaryotes, yet compared to the core histones relatively little is known about their in vivo functions. The filamentous fungus Neurospora crassa encodes a single H1 protein that is not essential for viability. To investigate the role of N. crassa H1, we constructed a functional FLAG-tagged H1 fusion protein and performed genomic and molecular analyses. Cell fractionation experiments showed that H1-3XFLAG is a chromatin binding protein. Chromatin-immunoprecipitation combined with sequencing (ChIP-seq) revealed that H1-3XFLAG is globally enriched throughout the genome with a subtle preference for promoters of expressed genes. In mammals, the stoichiometry of H1 impacts nucleosome repeat length. To determine if H1 impacts nucleosome occupancy or nucleosome positioning in N. crassa, we performed micrococcal nuclease digestion in the wild-type and the [Formula: see text]hH1 strain followed by sequencing (MNase-seq). Deletion of hH1 did not significantly impact nucleosome positioning or nucleosome occupancy. Analysis of DNA methylation by whole-genome bisulfite sequencing (MethylC-seq) revealed a modest but global increase in DNA methylation in the [Formula: see text]hH1 mutant. Together, these data suggest that H1 acts as a nonspecific chromatin binding protein that can limit accessibility of the DNA methylation machinery in N. crassa.

  8. Histone H1 Limits DNA Methylation in Neurospora crassa

    Directory of Open Access Journals (Sweden)

    Michael Seymour

    2016-07-01

    Full Text Available Histone H1 variants, known as linker histones, are essential chromatin components in higher eukaryotes, yet compared to the core histones relatively little is known about their in vivo functions. The filamentous fungus Neurospora crassa encodes a single H1 protein that is not essential for viability. To investigate the role of N. crassa H1, we constructed a functional FLAG-tagged H1 fusion protein and performed genomic and molecular analyses. Cell fractionation experiments showed that H1-3XFLAG is a chromatin binding protein. Chromatin-immunoprecipitation combined with sequencing (ChIP-seq revealed that H1-3XFLAG is globally enriched throughout the genome with a subtle preference for promoters of expressed genes. In mammals, the stoichiometry of H1 impacts nucleosome repeat length. To determine if H1 impacts nucleosome occupancy or nucleosome positioning in N. crassa, we performed micrococcal nuclease digestion in the wild-type and the ΔhH1 strain followed by sequencing (MNase-seq. Deletion of hH1 did not significantly impact nucleosome positioning or nucleosome occupancy. Analysis of DNA methylation by whole-genome bisulfite sequencing (MethylC-seq revealed a modest but global increase in DNA methylation in the ΔhH1 mutant. Together, these data suggest that H1 acts as a nonspecific chromatin binding protein that can limit accessibility of the DNA methylation machinery in N. crassa.

  9. Nucleosome adaptability conferred by sequence and structural variations in histone H2A-H2B dimers.

    Science.gov (United States)

    Shaytan, Alexey K; Landsman, David; Panchenko, Anna R

    2015-06-01

    Nucleosome variability is essential for their functions in compacting the chromatin structure and regulation of transcription, replication and cell reprogramming. The DNA molecule in nucleosomes is wrapped around an octamer composed of four types of core histones (H3, H4, H2A, H2B). Nucleosomes represent dynamic entities and may change their conformation, stability and binding properties by employing different sets of histone variants or by becoming post-translationally modified. There are many variants of histones H2A and H2B. Specific H2A and H2B variants may preferentially associate with each other resulting in different combinations of variants and leading to the increased combinatorial complexity of nucleosomes. In addition, the H2A-H2B dimer can be recognized and substituted by chaperones/remodelers as a distinct unit, can assemble independently and is stable during nucleosome unwinding. In this review we discuss how sequence and structural variations in H2A-H2B dimers may provide necessary complexity and confer the nucleosome functional variability.

  10. Oncogenic potential of histone-variant H2A.Z.1 and its regulatory role in cell cycle and epithelial-mesenchymal transition in liver cancer

    Science.gov (United States)

    Eun, Jung Woo; Shen, Qingyu; Kim, Hyung Seok; Shin, Woo Chan; Ahn, Young Min; Park, Won Sang; Lee, Jung Young; Nam, Suk Woo

    2016-01-01

    H2A.Z is a highly conserved H2A variant, and two distinct H2A.Z isoforms, H2A.Z.1 and H2A.Z.2, have been identified as products of two non-allelic genes, H2AFZ and H2AFV. H2A.Z has been reported to be overexpressed in breast, prostate and bladder cancers, but most studies did not clearly distinguish between isoforms. One recent study reported a unique role for the H2A.Z isoform H2A.Z.2 as a driver of malignant melanoma. Here we first report that H2A.Z.1 plays a pivotal role in the liver tumorigenesis by selectively regulating key molecules in cell cycle and epithelial-mesenchymal transition (EMT). H2AFZ expression was significantly overexpressed in a large cohort of hepatocellular carcinoma (HCC) patients, and high expression of H2AFZ was significantly associated with their poor prognosis. H2A.Z.1 overexpression was demonstrated in a subset of human HCC and cell lines. H2A.Z.1 knockdown suppressed HCC cell growth by transcriptional deregulation of cell cycle proteins and caused apoptotic cell death of HCC cells. We also observed that H2A.Z.1 knockdown reduced the metastatic potential of HCC cells by selectively modulating epithelial-mesenchymal transition regulatory proteins such as E-cadherin and fibronectin. In addition, H2A.Z.1 knockdown reduced the in vivo tumor growth rate in a mouse xenograft model. In conclusion, our findings suggest the oncogenic potential of H2A.Z.1 in liver tumorigenesis and that it plays established role in accelerating cell cycle transition and EMT during hepatocarcinogenesis. This makes H2A.Z.1 a promising target in liver cancer therapy. PMID:26863632

  11. Oncogenic potential of histone-variant H2A.Z.1 and its regulatory role in cell cycle and epithelial-mesenchymal transition in liver cancer.

    Science.gov (United States)

    Yang, Hee Doo; Kim, Pum-Joon; Eun, Jung Woo; Shen, Qingyu; Kim, Hyung Seok; Shin, Woo Chan; Ahn, Young Min; Park, Won Sang; Lee, Jung Young; Nam, Suk Woo

    2016-03-08

    H2A.Z is a highly conserved H2A variant, and two distinct H2A.Z isoforms, H2A.Z.1 and H2A.Z.2, have been identified as products of two non-allelic genes, H2AFZ and H2AFV. H2A.Z has been reported to be overexpressed in breast, prostate and bladder cancers, but most studies did not clearly distinguish between isoforms. One recent study reported a unique role for the H2A.Z isoform H2A.Z.2 as a driver of malignant melanoma. Here we first report that H2A.Z.1 plays a pivotal role in the liver tumorigenesis by selectively regulating key molecules in cell cycle and epithelial-mesenchymal transition (EMT). H2AFZ expression was significantly overexpressed in a large cohort of hepatocellular carcinoma (HCC) patients, and high expression of H2AFZ was significantly associated with their poor prognosis. H2A.Z.1 overexpression was demonstrated in a subset of human HCC and cell lines. H2A.Z.1 knockdown suppressed HCC cell growth by transcriptional deregulation of cell cycle proteins and caused apoptotic cell death of HCC cells. We also observed that H2A.Z.1 knockdown reduced the metastatic potential of HCC cells by selectively modulating epithelial-mesenchymal transition regulatory proteins such as E-cadherin and fibronectin. In addition, H2A.Z.1 knockdown reduced the in vivo tumor growth rate in a mouse xenograft model. In conclusion, our findings suggest the oncogenic potential of H2A.Z.1 in liver tumorigenesis and that it plays established role in accelerating cell cycle transition and EMT during hepatocarcinogenesis. This makes H2A.Z.1 a promising target in liver cancer therapy.

  12. Interaction of short peptides with FITC-labeled wheat histones and their complexes with deoxyribooligonucleotides.

    Science.gov (United States)

    Fedoreyeva, L I; Smirnova, T A; Kolomijtseva, G Ya; Khavinson, V Kh; Vanyushin, B F

    2013-02-01

    Judging from fluorescence modulation (quenching), short peptides (Ala-Glu-Asp-Gly, Glu-Asp-Arg, Ala-Glu-Asp-Leu, Lys-Glu-Asp-Gly, Ala-Glu-Asp-Arg, and Lys-Glu-Asp-Trp) bind with FITC-labeled wheat histones H1, H2B, H3, and H4. This results from the interaction of the peptides with the N-terminal histone regions that contain respective and seemingly homologous peptide-binding motifs. Because homologous amino acid sequences in wheat core histones were not found, the peptides seem to bind with some core histone regions having specific conformational structure. Peptide binding with histones and histone-deoxyribooligonucleotide complexes depends on the nature of the histone and the primary structures of the peptides and oligonucleotides; thus, it is site specific. Histones H1 bind preferentially with single-stranded oligonucleotides by homologous sites in the C-terminal region of the protein. Unlike histone H1, the core histones bind predominantly with double-stranded methylated oligonucleotides and methylated DNA. Stern-Volmer constants of interaction of histone H1 and core histones with double-stranded hemimethylated oligonucleotides are higher compared with that of binding with unmethylated ones. DNA or deoxyribooligonucleotides in a complex with histones can enhance or inhibit peptide binding. It is suggested that site-specific interactions of short biologically active peptides with histone tails can serve in chromatin as control epigenetic mechanisms of regulation of gene activity and cellular differentiation.

  13. Detection of NAM-A1 Natural Variants in Bread Wheat Reveals Differences in Haplotype Distribution between a Worldwide Core Collection and European Elite Germplasm

    Directory of Open Access Journals (Sweden)

    Fabien Cormier

    2015-04-01

    Full Text Available In wheat, remobilization of nitrogen absorbed before anthesis and regulation of monocarpic senescence is a major issue in breeding for nutrient use efficiency. We identified natural variants of NAM-A1, a gene having the same role as its well-characterized homoeolog NAM-B1, a NAC transcription factor associated with senescence kinetics and nutrient remobilization to the grain. Differences in haplotype frequencies between a worldwide core collection and a panel of European elite varieties were assessed and discussed. Moreover, hypotheses for the loss of function of the most common haplotype in elite European germplasm are discussed.

  14. CORE

    DEFF Research Database (Denmark)

    Krigslund, Jeppe; Hansen, Jonas; Hundebøll, Martin

    2013-01-01

    different flows. Instead of maintaining these approaches separate, we propose a protocol (CORE) that brings together these coding mechanisms. Our protocol uses random linear network coding (RLNC) for intra- session coding but allows nodes in the network to setup inter- session coding regions where flows...... intersect. Routes for unicast sessions are agnostic to other sessions and setup beforehand, CORE will then discover and exploit intersecting routes. Our approach allows the inter-session regions to leverage RLNC to compensate for losses or failures in the overhearing or transmitting process. Thus, we...... increase the benefits of XORing by exploiting the underlying RLNC structure of individual flows. This goes beyond providing additional reliability to each individual session and beyond exploiting coding opportunistically. Our numerical results show that CORE outperforms both forwarding and COPE...

  15. CORE

    DEFF Research Database (Denmark)

    Krigslund, Jeppe; Hansen, Jonas; Hundebøll, Martin

    2013-01-01

    different flows. Instead of maintaining these approaches separate, we propose a protocol (CORE) that brings together these coding mechanisms. Our protocol uses random linear network coding (RLNC) for intra- session coding but allows nodes in the network to setup inter- session coding regions where flows...... intersect. Routes for unicast sessions are agnostic to other sessions and setup beforehand, CORE will then discover and exploit intersecting routes. Our approach allows the inter-session regions to leverage RLNC to compensate for losses or failures in the overhearing or transmitting process. Thus, we...... increase the benefits of XORing by exploiting the underlying RLNC structure of individual flows. This goes beyond providing additional reliability to each individual session and beyond exploiting coding opportunistically. Our numerical results show that CORE outperforms both forwarding and COPE...

  16. HAMLET interacts with histones and chromatin in tumor cell nuclei.

    Science.gov (United States)

    Düringer, Caroline; Hamiche, Ali; Gustafsson, Lotta; Kimura, Hiroshi; Svanborg, Catharina

    2003-10-24

    HAMLET is a folding variant of human alpha-lactalbumin in an active complex with oleic acid. HAMLET selectively enters tumor cells, accumulates in their nuclei and induces apoptosis-like cell death. This study examined the interactions of HAMLET with nuclear constituents and identified histones as targets. HAMLET was found to bind histone H3 strongly and to lesser extent histones H4 and H2B. The specificity of these interactions was confirmed using BIAcore technology and chromatin assembly assays. In vivo in tumor cells, HAMLET co-localized with histones and perturbed the chromatin structure; HAMLET was found associated with chromatin in an insoluble nuclear fraction resistant to salt extraction. In vitro, HAMLET bound strongly to histones and impaired their deposition on DNA. We conclude that HAMLET interacts with histones and chromatin in tumor cell nuclei and propose that this interaction locks the cells into the death pathway by irreversibly disrupting chromatin organization.

  17. HdaA, a Major Class 2 Histone Deacetylase of Aspergillus nidulans, Affects Growth under Conditions of Oxidative Stress

    OpenAIRE

    Tribus, Martin; Galehr, Johannes; Trojer, Patrick; Brosch, Gerald; Loidl, Peter; Marx, Florentine; Haas, Hubertus; Graessle, Stefan

    2005-01-01

    Histone deacetylases (HDACs) catalyze the removal of acetyl groups from the ɛ-amino group of distinct lysine residues in the amino-terminal tail of core histones. Since the acetylation status of core histones plays a crucial role in fundamental processes in eukaryotic organisms, such as replication and regulation of transcription, recent research has focused on the enzymes responsible for the acetylation/deacetylation of core histones. Very recently, we showed that HdaA, a member of the Sacch...

  18. Replication-Uncoupled Histone Deposition during Adenovirus DNA Replication

    OpenAIRE

    Komatsu, Tetsuro; Nagata, Kyosuke

    2012-01-01

    In infected cells, the chromatin structure of the adenovirus genome DNA plays critical roles in its genome functions. Previously, we reported that in early phases of infection, incoming viral DNA is associated with both viral core protein VII and cellular histones. Here we show that in late phases of infection, newly synthesized viral DNA is also associated with histones. We also found that the knockdown of CAF-1, a histone chaperone that functions in the replication-coupled deposition of his...

  19. Readers of histone modifications

    Institute of Scientific and Technical Information of China (English)

    Miyong Yun; Jun Wu; Jerry L Workman; Bing Li

    2011-01-01

    Histone modifications not only play important roles in regulating chromatin structure and nuclear processes but also can be passed to daughter cells as epigenetic marks.Accumulating evidence suggests that the key function of histone modifications is to signal for recruitment or activity of downstream effectors. Here, we discuss the latest discovery of histone-modification readers and how the modification language is interpreted.

  20. Biochemical Analysis Reveals the Multifactorial Mechanism of Histone H3 Clipping by Chicken Liver Histone H3 Protease

    KAUST Repository

    Chauhan, Sakshi

    2016-09-02

    Proteolytic clipping of histone H3 has been identified in many organisms. Despite several studies, the mechanism of clipping, the substrate specificity, and the significance of this poorly understood epigenetic mechanism are not clear. We have previously reported histone H3 specific proteolytic clipping and a protein inhibitor in chicken liver. However, the sites of clipping are still not known very well. In this study, we attempt to identify clipping sites in histone H3 and to determine the mechanism of inhibition by stefin B protein, a cysteine protease inhibitor. By employing site-directed mutagenesis and in vitro biochemical assays, we have identified three distinct clipping sites in recombinant human histone H3 and its variants (H3.1, H3.3, and H3t). However, post-translationally modified histones isolated from chicken liver and Saccharomyces cerevisiae wild-type cells showed different clipping patterns. Clipping of histone H3 N-terminal tail at three sites occurs in a sequential manner. We have further observed that clipping sites are regulated by the structure of the N-terminal tail as well as the globular domain of histone H3. We also have identified the QVVAG region of stefin B protein to be very crucial for inhibition of the protease activity. Altogether, our comprehensive biochemical studies have revealed three distinct clipping sites in histone H3 and their regulation by the structure of histone H3, histone modifications marks, and stefin B.

  1. Histone Deacetylase Genes in Arabidopsis Development

    Institute of Scientific and Technical Information of China (English)

    Courtney Hollender; Zhongchi Liu

    2008-01-01

    Histone acetylatlon and deacetylation are directly connected with transcriptional activation and silencing in eukaryotas.Gene families for enzymes that accomplish these histone modifications show surprising complexity in domain organization,tissue-specific expression, and function. This review is focused on the family of histone deacetylases (HDACs) that remove the acetyl group from core histone tails, resulting in a "closed" chromatin and transcriptional repression. In Arabidopsis,18 HDAC genes are divided in to three different types - RPD3-1ike, HD-tuin and sirtuin - with two or more members ineach type. The structural feature of each HDAC class, the expression profile of each HDAC gene during development and functional insights of important family members are summarized here. It is clear that HDACs are an important class of global transcriptional regulators that play crucial roles in plant development, defense, and adaptation.

  2. A PHD in histone language

    Science.gov (United States)

    Chandrika, Nulu Naga Prafulla; Sundaravelpandian, Kalaipandian; Schmidt, Wolfgang

    2013-01-01

    Post-translational modifications of core histones are important for various DNA-templated processes such as transcription and repair. We recently reported that the ALFIN LIKE 6 (AL6) gene, identified in a forward genetic screen, is critical for phosphate deficiency-induced root hair formation and several other processes associated with the regulation of cellular phosphate homeostasis. AL6 contains a Plant Homeo Domain (PHD) finger that can bind to trimethylated lysine 4 of histone H3 (H3K4me3). Homozygous mutants defective in AL6 expression form very short root hairs under phosphate-deficient conditions, presumably caused by altered expression of putative primary and secondary down-stream targets of AL6. In this Addendum, we speculate about possible roles of AL6, H3K4 trimethylation and other chromatin modifications in the adaptation of plants to low phosphate availability. PMID:23531693

  3. ChIp-seq of bovine cells (MDBK) to study butyrate-induced histone modification with 10 datasets

    Science.gov (United States)

    Next-generation sequencing was combined with chromatin immunoprecipitation (ChIP) technology to analyze histone modification (acetylation) induced by butyrate and to map the epigenomic landscape of normal histone H3, H4 in rumen cells of the cow. Ten variants of histone H3 and H4 modification were m...

  4. Current Perspectives on Histone Demethylases

    Institute of Scientific and Technical Information of China (English)

    Xiaoqing TIAN; Jingyuan FANG

    2007-01-01

    The posttranslational modification of histones plays an important role in chromatin regulation.Histone methylation influences constitutive heterochromatin, genomic imprinting, X-chromosome inactivation and gene transcription. Histone demethylase catalyzes the removal of methyl groups on lysine or arginine residues of histones. Two kinds of histone lysine demethylases have been identified, including lysine specific demethylase 1 and Jumonji C (JmjC) domain family proteins. These histone demethylases are involved in the regulation of gene expression. Histone modification is a dynamic process, and the imbalance of histone methylation has been linked to cancers. Therefore, histone demethylases may represent a new target for anti-cancer therapy.

  5. Histones and nucleosomes in Archaea and Eukarya: a comparative analysis.

    Science.gov (United States)

    Pereira, S L; Reeve, J N

    1998-08-01

    Archaeal histones from mesophilic, thermophilic, and hyperthermophilic members of the Euryarchaeota have primary sequences, the histone fold, tertiary structures, and dimer formation in common with the eukaryal nucleosome core histones H2A, H2B, H3, and H4. Archaeal histones form nucleoprotein complexes in vitro and in vivo, designated archaeal nucleosomes, that contain histone tetramers and protect approximately 60 base pairs of DNA from nuclease digestion. Based on the sequence and structural homologies and experimental data reviewed here, archaeal nucleosomes appear similar, and may be homologous in evolutionary terms and function, to the structure at the center of the eukaryal nucleosome formed by the histone (H3 + H4)2 tetramer.

  6. Lysine methylation: beyond histones

    Institute of Scientific and Technical Information of China (English)

    Xi Zhang; Hong Wen; Xiaobing Shi

    2012-01-01

    Posttranslational modifications (PTMs) of histone proteins,such as acetylation,methylation,phosphorylation,and ubiquitylation,play essential roles in regulating chromatin dynamics.Combinations of different modifications on the histone proteins,termed 'histone code' in many cases,extend the information potential of the genetic code by regulating DNA at the epigenetic level.Many PTMs occur on non-histone proteins as well as histones,regulating protein-protein interactions,stability,localization,and/or enzymatic activities of proteins involved in diverse cellular processes.Although protein phosphorylation,ubiquitylation,and acetylation have been extensively studied,only a few proteins other than histones have been reported that can be modified by lysine methylation.This review summarizes the current progress on lysine methylation of nonhistone proteins,and we propose that lysine methylation,like phosphorylation and acetylation,is a common PTM that regulates proteins in diverse cellular processes.

  7. The role of histones in the immune responses of aquatic invertebrates

    OpenAIRE

    C Nikapitiya; T Dorrington; Gómez-Chiarri, M.

    2013-01-01

    Histones are primary components of eukaryotic chromatin and highly abundant in all animal cells. In addition to their important role in chromatin structure and transcriptional regulation, histones contribute to innate immune responses. In several aquatic invertebrate species, as well as in many other invertebrate and vertebrate species, the transcripts for core histones are upregulated in response to immune challenge and exposure to environmental stressors. Histones show antimicrobial activit...

  8. Readers of histone methylarginine marks.

    Science.gov (United States)

    Gayatri, Sitaram; Bedford, Mark T

    2014-08-01

    Arginine methylation is a common posttranslational modification (PTM) that alters roughly 0.5% of all arginine residues in the cells. There are three types of arginine methylation: monomethylarginine (MMA), asymmetric dimethylarginine (ADMA), and symmetric dimethylarginine (SDMA). These three PTMs are enriched on RNA-binding proteins and on histones, and also impact signal transduction cascades. To date, over thirty arginine methylation sites have been cataloged on the different core histones. These modifications alter protein structure, impact interactions with DNA, and also generate docking sites for effector molecules. The primary "readers" of methylarginine marks are Tudor domain-containing proteins. The complete family of thirty-six Tudor domain-containing proteins has yet to be fully characterized, but at least ten bind methyllysine motifs and eight bind methylarginine motifs. In this review, we will highlight the biological roles of the Tudor domains that interact with arginine methylated motifs, and also address other types of interactions that are regulated by these particular PTMs. This article is part of a Special Issue entitled: Molecular mechanisms of histone modification function. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Lateral Thinking: How Histone Modifications Regulate Gene Expression.

    Science.gov (United States)

    Lawrence, Moyra; Daujat, Sylvain; Schneider, Robert

    2016-01-01

    The DNA of each cell is wrapped around histone octamers, forming so-called 'nucleosomal core particles'. These histone proteins have tails that project from the nucleosome and many residues in these tails can be post-translationally modified, influencing all DNA-based processes, including chromatin compaction, nucleosome dynamics, and transcription. In contrast to those present in histone tails, modifications in the core regions of the histones had remained largely uncharacterised until recently, when some of these modifications began to be analysed in detail. Overall, recent work has shown that histone core modifications can not only directly regulate transcription, but also influence processes such as DNA repair, replication, stemness, and changes in cell state. In this review, we focus on the most recent developments in our understanding of histone modifications, particularly those on the lateral surface of the nucleosome. This region is in direct contact with the DNA and is formed by the histone cores. We suggest that these lateral surface modifications represent a key insight into chromatin regulation in the cell. Therefore, lateral surface modifications form a key area of interest and a focal point of ongoing study in epigenetics.

  10. Histones and histone modifications in perinuclear chromatin anchoring: from yeast to man.

    Science.gov (United States)

    Harr, Jennifer C; Gonzalez-Sandoval, Adriana; Gasser, Susan M

    2016-02-01

    It is striking that within a eukaryotic nucleus, the genome can assume specific spatiotemporal distributions that correlate with the cell's functional states. Cell identity itself is determined by distinct sets of genes that are expressed at a given time. On the level of the individual gene, there is a strong correlation between transcriptional activity and associated histone modifications. Histone modifications act by influencing the recruitment of non-histone proteins and by determining the level of chromatin compaction, transcription factor binding, and transcription elongation. Accumulating evidence also shows that the subnuclear position of a gene or domain correlates with its expression status. Thus, the question arises whether this spatial organization results from or determines a gene's chromatin status. Although the association of a promoter with the inner nuclear membrane (INM) is neither necessary nor sufficient for repression, the perinuclear sequestration of heterochromatin is nonetheless conserved from yeast to man. How does subnuclear localization influence gene expression? Recent work argues that the common denominator between genome organization and gene expression is the modification of histones and in some cases of histone variants. This provides an important link between local chromatin structure and long-range genome organization in interphase cells. In this review, we will evaluate how histones contribute to the latter, and discuss how this might help to regulate genes crucial for cell differentiation. © 2016 The Authors. Published under the terms of the CC BY NC ND 4.0 license.

  11. "Hunt"-ing for post-translational modifications that underlie the histone code

    Science.gov (United States)

    Taverna, Sean D.; David Allis, C.; Hake, Sandra B.

    2007-01-01

    Eukaryotic cells package their DNA with histone proteins to form chromatin that can be regulated to enable transcription, DNA repair and replication in response to cellular needs and external stimuli. A wealth of recent studies of post-translational histone modifications and histone variants have led to an explosion of insights into and more questions about how these processes might be regulated. Work from Donald Hunt and colleagues contributed greatly to our understanding of the "histone code" by developing novel methods to study and identify histone modifications in both generic and specialized variant histone proteins. Without his expertise, the field of chromatin biology would not be where it is today. In recognition, we are pleased to contribute to a special issue of the International Journal of Mass Spectrometry dedicated to the many advances pioneered by the Hunt laboratory, which have enhanced the science of many fields and the careers of many scientists.

  12. Histone chaperones link histone nuclear import and chromatin assembly.

    Science.gov (United States)

    Keck, Kristin M; Pemberton, Lucy F

    2013-01-01

    Histone chaperones are proteins that shield histones from nonspecific interactions until they are assembled into chromatin. After their synthesis in the cytoplasm, histones are bound by different histone chaperones, subjected to a series of posttranslational modifications and imported into the nucleus. These evolutionarily conserved modifications, including acetylation and methylation, can occur in the cytoplasm, but their role in regulating import is not well understood. As part of histone import complexes, histone chaperones may serve to protect the histones during transport, or they may be using histones to promote their own nuclear localization. In addition, there is evidence that histone chaperones can play an active role in the import of histones. Histone chaperones have also been shown to regulate the localization of important chromatin modifying enzymes. This review is focused on the role histone chaperones play in the early biogenesis of histones, the distinct cytoplasmic subcomplexes in which histone chaperones have been found in both yeast and mammalian cells and the importins/karyopherins and nuclear localization signals that mediate the nuclear import of histones. We also address the role that histone chaperone localization plays in human disease. This article is part of a Special Issue entitled: Histone chaperones and chromatin assembly.

  13. Regulation and function of histone acetyltransferase MOF.

    Science.gov (United States)

    Yang, Yang; Han, Xiaofei; Guan, Jingyun; Li, Xiangzhi

    2014-03-01

    The mammalian MOF (male absent on the first), a member of the MYST (MOZ, YBF2, SAS2, and Tip60) family of histone acetyltransferases (HATs), is the major enzyme that catalyzes the acetylation of histone H4 on lysine 16. Acetylation of K16 is a prevalent mark associated with chromatin decondensation. MOF has recently been shown to play an essential role in maintaining normal cell functions. In this study, we discuss the important roles of MOF in DNA damage repair, apoptosis, and tumorigenesis. We also analyze the role of MOF as a key regulator of the core transcriptional network of embryonic stem cells.

  14. Histone chaperone-mediated nucleosome assembly process.

    Science.gov (United States)

    Fan, Hsiu-Fang; Liu, Zi-Ning; Chow, Sih-Yao; Lu, Yi-Han; Li, Hsin

    2015-01-01

    A huge amount of information is stored in genomic DNA and this stored information resides inside the nucleus with the aid of chromosomal condensation factors. It has been reported that the repeat nucleosome core particle (NCP) consists of 147-bp of DNA and two copies of H2A, H2B, H3 and H4. Regulation of chromosomal structure is important to many processes inside the cell. In vivo, a group of histone chaperones facilitate and regulate nucleosome assembly. How NCPs are constructed with the aid of histone chaperones remains unclear. In this study, the histone chaperone-mediated nucleosome assembly process was investigated using single-molecule tethered particle motion (TPM) experiments. It was found that Asf1 is able to exert more influence than Nap1 and poly glutamate acid (PGA) on the nucleosome formation process, which highlights Asf1's specific role in tetrasome formation. Thermodynamic parameters supported a model whereby energetically favored nucleosomal complexes compete with non-nucleosomal complexes. In addition, our kinetic findings propose the model that histone chaperones mediate nucleosome assembly along a path that leads to enthalpy-favored products with free histones as reaction substrates.

  15. The specification and global reprogramming of histone epigenetic marks during gamete formation and early embryo development in C. elegans.

    Directory of Open Access Journals (Sweden)

    Mark Samson

    2014-10-01

    Full Text Available In addition to the DNA contributed by sperm and oocytes, embryos receive parent-specific epigenetic information that can include histone variants, histone post-translational modifications (PTMs, and DNA methylation. However, a global view of how such marks are erased or retained during gamete formation and reprogrammed after fertilization is lacking. To focus on features conveyed by histones, we conducted a large-scale proteomic identification of histone variants and PTMs in sperm and mixed-stage embryo chromatin from C. elegans, a species that lacks conserved DNA methylation pathways. The fate of these histone marks was then tracked using immunostaining. Proteomic analysis found that sperm harbor ∼2.4 fold lower levels of histone PTMs than embryos and revealed differences in classes of PTMs between sperm and embryos. Sperm chromatin repackaging involves the incorporation of the sperm-specific histone H2A variant HTAS-1, a widespread erasure of histone acetylation, and the retention of histone methylation at sites that mark the transcriptional history of chromatin domains during spermatogenesis. After fertilization, we show HTAS-1 and 6 histone PTM marks distinguish sperm and oocyte chromatin in the new embryo and characterize distinct paternal and maternal histone remodeling events during the oocyte-to-embryo transition. These include the exchange of histone H2A that is marked by ubiquitination, retention of HTAS-1, removal of the H2A variant HTZ-1, and differential reprogramming of histone PTMs. This work identifies novel and conserved features of paternal chromatin that are specified during spermatogenesis and processed in the embryo. Furthermore, our results show that different species, even those with diverged DNA packaging and imprinting strategies, use conserved histone modification and removal mechanisms to reprogram epigenetic information.

  16. Citrullination regulates pluripotency and histone H1 binding to chromatin

    Science.gov (United States)

    Christophorou, Maria A.; Castelo-Branco, Gonçalo; Halley-Stott, Richard P.; Oliveira, Clara Slade; Loos, Remco; Radzisheuskaya, Aliaksandra; Mowen, Kerri A.; Bertone, Paul; Silva, José C. R.; Zernicka-Goetz, Magdalena; Nielsen, Michael L.; Gurdon, John B.; Kouzarides, Tony

    2014-03-01

    Citrullination is the post-translational conversion of an arginine residue within a protein to the non-coded amino acid citrulline. This modification leads to the loss of a positive charge and reduction in hydrogen-bonding ability. It is carried out by a small family of tissue-specific vertebrate enzymes called peptidylarginine deiminases (PADIs) and is associated with the development of diverse pathological states such as autoimmunity, cancer, neurodegenerative disorders, prion diseases and thrombosis. Nevertheless, the physiological functions of citrullination remain ill-defined, although citrullination of core histones has been linked to transcriptional regulation and the DNA damage response. PADI4 (also called PAD4 or PADV), the only PADI with a nuclear localization signal, was previously shown to act in myeloid cells where it mediates profound chromatin decondensation during the innate immune response to infection. Here we show that the expression and enzymatic activity of Padi4 are also induced under conditions of ground-state pluripotency and during reprogramming in mouse. Padi4 is part of the pluripotency transcriptional network, binding to regulatory elements of key stem-cell genes and activating their expression. Its inhibition lowers the percentage of pluripotent cells in the early mouse embryo and significantly reduces reprogramming efficiency. Using an unbiased proteomic approach we identify linker histone H1 variants, which are involved in the generation of compact chromatin, as novel PADI4 substrates. Citrullination of a single arginine residue within the DNA-binding site of H1 results in its displacement from chromatin and global chromatin decondensation. Together, these results uncover a role for citrullination in the regulation of pluripotency and provide new mechanistic insights into how citrullination regulates chromatin compaction.

  17. The intrinsically disordered distal face of nucleoplasmin recognizes distinct oligomerization states of histones

    Science.gov (United States)

    Ramos, Isbaal; Fernández-Rivero, Noelia; Arranz, Rocío; Aloria, Kerman; Finn, Ron; Arizmendi, Jesús M.; Ausió, Juan; Valpuesta, José María; Muga, Arturo; Prado, Adelina

    2014-01-01

    The role of Nucleoplasmin (NP) as a H2A-H2B histone chaperone has been extensively characterized. To understand its putative interaction with other histone ligands, we have characterized its ability to bind H3-H4 and histone octamers. We find that the chaperone forms distinct complexes with histones, which differ in the number of molecules that build the assembly and in their spatial distribution. When complexed with H3-H4 tetramers or histone octamers, two NP pentamers form an ellipsoidal particle with the histones located at the center of the assembly, in stark contrast with the NP/H2A-H2B complex that contains up to five histone dimers bound to one chaperone pentamer. This particular assembly relies on the ability of H3-H4 to form tetramers either in solution or as part of the octamer, and it is not observed when a variant of H3 (H3C110E), unable to form stable tetramers, is used instead of the wild-type protein. Our data also suggest that the distal face of the chaperone is involved in the interaction with distinct types of histones, as supported by electron microscopy analysis of the different NP/histone complexes. The use of the same structural region to accommodate all type of histones could favor histone exchange and nucleosome dynamics. PMID:24121686

  18. Cloning and characterization of the major histone H2A genes completes the cloning and sequencing of known histone genes of Tetrahymena thermophila.

    Science.gov (United States)

    Liu, X; Gorovsky, M A

    1996-01-01

    A truncated cDNA clone encoding Tetrahymena thermophila histone H2A2 was isolated using synthetic degenerate oligonucleotide probes derived from H2A protein sequences of Tetrahymena pyriformis. The cDNA clone was used as a homologous probe to isolate a truncated genomic clone encoding H2A1. The remaining regions of the genes for H2A1 (HTA1) and H2A2 (HTA2) were then isolated using inverse PCR on circularized genomic DNA fragments. These partial clones were assembled into intact HTA1 and HTA2 clones. Nucleotide sequences of the two genes were highly homologous within the coding region but not in the noncoding regions. Comparison of the deduced amino acid sequences with protein sequences of T. pyriformis H2As showed only two and three differences respectively, in a total of 137 amino acids for H2A1, and 132 amino acids for H2A2, indicating the two genes arose before the divergence of these two species. The HTA2 gene contains a TAA triplet within the coding region, encoding a glutamine residue. In contrast with the T. thermophila HHO and HTA3 genes, no introns were identified within the two genes. The 5'- and 3'-ends of the histone H2A mRNAs; were determined by RNase protection and by PCR mapping using RACE and RLM-RACE methods. Both genes encode polyadenylated mRNAs and are highly expressed in vegetatively growing cells but only weakly expressed in starved cultures. With the inclusion of these two genes, T. thermophila is the first organism whose entire complement of known core and linker histones, including replication-dependent and basal variants, has been cloned and sequenced. PMID:8760889

  19. Application of histone modification-specific interaction domains as an alternative to antibodies

    Science.gov (United States)

    Kungulovski, Goran; Kycia, Ina; Tamas, Raluca; Jurkowska, Renata Z.; Kudithipudi, Srikanth; Henry, Chisato; Reinhardt, Richard; Labhart, Paul

    2014-01-01

    Post-translational modifications (PTMs) of histones constitute a major chromatin indexing mechanism, and their proper characterization is of highest biological importance. So far, PTM-specific antibodies have been the standard reagent for studying histone PTMs despite caveats such as lot-to-lot variability of specificity and binding affinity. Herein, we successfully employed naturally occurring and engineered histone modification interacting domains for detection and identification of histone PTMs and ChIP-like enrichment of different types of chromatin. Our results demonstrate that histone interacting domains are robust and highly specific reagents that can replace or complement histone modification antibodies. These domains can be produced recombinantly in Escherichia coli at low cost and constant quality. Protein design of reading domains allows for generation of novel specificities, addition of affinity tags, and preparation of PTM binding pocket variants as matching negative controls, which is not possible with antibodies. PMID:25301795

  20. Critical Role of Histone Turnover in Neuronal Transcription and Plasticity.

    Science.gov (United States)

    Maze, Ian; Wenderski, Wendy; Noh, Kyung-Min; Bagot, Rosemary C; Tzavaras, Nikos; Purushothaman, Immanuel; Elsässer, Simon J; Guo, Yin; Ionete, Carolina; Hurd, Yasmin L; Tamminga, Carol A; Halene, Tobias; Farrelly, Lorna; Soshnev, Alexey A; Wen, Duancheng; Rafii, Shahin; Birtwistle, Marc R; Akbarian, Schahram; Buchholz, Bruce A; Blitzer, Robert D; Nestler, Eric J; Yuan, Zuo-Fei; Garcia, Benjamin A; Shen, Li; Molina, Henrik; Allis, C David

    2015-07-01

    Turnover and exchange of nucleosomal histones and their variants, a process long believed to be static in post-replicative cells, remains largely unexplored in brain. Here, we describe a novel mechanistic role for HIRA (histone cell cycle regulator) and proteasomal degradation-associated histone dynamics in the regulation of activity-dependent transcription, synaptic connectivity, and behavior. We uncover a dramatic developmental profile of nucleosome occupancy across the lifespan of both rodents and humans, with the histone variant H3.3 accumulating to near-saturating levels throughout the neuronal genome by mid-adolescence. Despite such accumulation, H3.3-containing nucleosomes remain highly dynamic-in a modification-independent manner-to control neuronal- and glial-specific gene expression patterns throughout life. Manipulating H3.3 dynamics in both embryonic and adult neurons confirmed its essential role in neuronal plasticity and cognition. Our findings establish histone turnover as a critical and previously undocumented regulator of cell type-specific transcription and plasticity in mammalian brain. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Histone deacetylase inhibitors promote the tumoricidal effect of HAMLET.

    Science.gov (United States)

    Brest, Patrick; Gustafsson, Mattias; Mossberg, Ann-Kristin; Gustafsson, Lotta; Duringer, Caroline; Hamiche, Ali; Svanborg, Catharina

    2007-12-01

    Histone deacetylase inhibitors (HDIs) and HAMLET (human alpha-lactalbumin made lethal to tumor cells) interact with histones, modify the structure of chromatin, and trigger tumor cell death. This study investigated how the combination of HDIs and HAMLET influences cell viability, histone acetylation, and DNA integrity. The pretreatment of tumor cells with HDIs was shown to enhance the lethal effect of HAMLET and the histone hyperacetylation response to HDIs increased even further after HAMLET treatment. HDIs and HAMLET were shown to target different histone domains as HAMLET bound tailless core histones, whereas HDIs modify the acetylation of the histone tail. DNA damage in response to HAMLET was increased by HDIs. The DNA repair response (p21WAFI expression) was induced by both agonists but abolished when the two agonists were combined. The results suggest that the synergy of HDIs and HAMLET is based on different but converging death pathways, both involving chromatin alterations. We speculate that HAMLET and HDIs might be combined to promote tumor cell death in vivo.

  2. Inhibitors of histone deacetylase

    DEFF Research Database (Denmark)

    2015-01-01

    The present invention relates to compounds of formula (I) or a pharmaceutically acceptable salt, hydrate, solvate, or prodrug thereof, wherein X1, X2, X3, X4, X5, W1, W2, W3, and W4 are as described. The present invention relates generally to inhibitors of histone deacetylase and to methods...

  3. Structural plasticity of histones H3-H4 facilitates their allosteric exchange between RbAp48 and ASF1.

    Science.gov (United States)

    Zhang, Wei; Tyl, Marek; Ward, Richard; Sobott, Frank; Maman, Joseph; Murthy, Andal S; Watson, Aleksandra A; Fedorov, Oleg; Bowman, Andrew; Owen-Hughes, Tom; El Mkami, Hassane; Murzina, Natalia V; Norman, David G; Laue, Ernest D

    2013-01-01

    The mechanisms by which histones are disassembled and reassembled into nucleosomes and chromatin structure during DNA replication, repair and transcription are poorly understood. A better understanding of the processes involved is, however, crucial if we are to understand whether and how histone variants and post-translationally modified histones are inherited in an epigenetic manner. To this end we have studied the interaction of the histone H3-H4 complex with the human retinoblastoma-associated protein RbAp48 and their exchange with a second histone chaperone, anti-silencing function protein 1 (ASF1). Exchange of histones H3-H4 between these two histone chaperones has a central role in the assembly of new nucleosomes, and we show here that the H3-H4 complex has an unexpected structural plasticity, which is important for this exchange.

  4. High-resolution genome-wide mapping of histone modifications.

    Science.gov (United States)

    Roh, Tae-young; Ngau, Wing Chi; Cui, Kairong; Landsman, David; Zhao, Keji

    2004-08-01

    The expression patterns of eukaryotic genomes are controlled by their chromatin structure, consisting of nucleosome subunits in which DNA of approximately 146 bp is wrapped around a core of 8 histone molecules. Post-translational histone modifications play an essential role in modifying chromatin structure. Here we apply a combination of SAGE and chromatin immunoprecipitation (ChIP) protocols to determine the distribution of hyperacetylated histones H3 and H4 in the Saccharomyces cerevisiae genome. We call this approach genome-wide mapping technique (GMAT). Using GMAT, we find that the highest acetylation levels are detected in the 5' end of a gene's coding region, but not in the promoter. Furthermore, we show that the histone acetyltransferase, GCN5p, regulates H3 acetylation in the promoter and 5' end of the coding regions. These findings indicate that GMAT should find valuable applications in mapping target sites of chromatin-modifying enzymes.

  5. Histone H2A.Z subunit exchange controls consolidation of recent and remote memory.

    Science.gov (United States)

    Zovkic, Iva B; Paulukaitis, Brynna S; Day, Jeremy J; Etikala, Deepa M; Sweatt, J David

    2014-11-27

    Memory formation is a multi-stage process that initially requires cellular consolidation in the hippocampus, after which memories are downloaded to the cortex for maintenance, in a process termed systems consolidation. Epigenetic mechanisms regulate both types of consolidation, but histone variant exchange, in which canonical histones are replaced with their variant counterparts, is an entire branch of epigenetics that has received limited attention in the brain and has never, to our knowledge, been studied in relation to cognitive function. Here we show that histone H2A.Z, a variant of histone H2A, is actively exchanged in response to fear conditioning in the hippocampus and the cortex, where it mediates gene expression and restrains the formation of recent and remote memory. Our data provide evidence for H2A.Z involvement in cognitive function and specifically implicate H2A.Z as a negative regulator of hippocampal consolidation and systems consolidation, probably through downstream effects on gene expression. Moreover, alterations in H2A.Z binding at later stages of systems consolidation suggest that this histone has the capacity to mediate stable molecular modifications required for memory retention. Overall, our data introduce histone variant exchange as a novel mechanism contributing to the molecular basis of cognitive function and implicate H2A.Z as a potential therapeutic target for memory disorders.

  6. Involvement of histone acetylation in the regulation of choline acetyltransferase gene in NG108-15 neuronal cells.

    Science.gov (United States)

    Aizawa, Shu; Yamamuro, Yutaka

    2010-03-01

    Post-translational modification of histone such as acetylation of N-terminal of lysine residues influences gene expression by modulating the accessibility of specific transcription factors to the promoter region, and is essential for a wide variety of cellular processes in the development of individual tissues, including the brain. However, few details concerning the acquisition of specific neurotransmitter phenotype have been obtained. In the present study, we investigated the possible involvement of histone acetylation in the gene expression of choline acetyltransferase (ChAT), a specific marker for cholinergic neuron and its function, in NG108-15 neuronal cells as an in vitro model of cholinergic neuron. Treatment with the histone deacetylase (HDAC) inhibitor trichostatin A (TSA), which induces global histone hyper-acetylation of the cells, resulted in marked increase in the expression of ChAT gene in proliferating NG108-15 cells. Furthermore, RT-PCR analysis using primer pairs for individual variants of ChAT mRNA (R1-4, N1, and M type) revealed that M type, not R1-4 and N1 type, ChAT mRNA were mainly transcribed, and chromatin immunoprecipitation assay indicated that the promoter region of M type ChAT gene was highly acetylated, in the dibutyryl cyclic AMP-induced neuronal differentiation of NG108-15 cells. The present findings demonstrate that the acquisition of neurotransmitter phenotype is epigenetically, at least the hyper-acetylation on the core promoter region of ChAT gene, regulated in NG108-15 neuronal cells.

  7. Directed evolution of a histone acetyltransferase--enhancing thermostability, whilst maintaining catalytic activity and substrate specificity.

    Science.gov (United States)

    Leemhuis, Hans; Nightingale, Karl P; Hollfelder, Florian

    2008-11-01

    Histone acetylation plays an integral role in the epigenetic regulation of gene expression. Transcriptional activity reflects the recruitment of opposing classes of enzymes to promoter elements; histone acetyltransferases (EC 2.3.1.48) that deposit acetyl marks at a subset of histone residues and histone deacetylases that remove them. Many histone acetyltransferases are difficult to study in solution because of their limited stability once purified. We have developed a directed evolution protocol that allows the screening of hundreds of histone acetyltransferase mutants for histone acetylating activity, and used this to enhance the thermostability of the human P/CAF histone acetyltransferase. Two rounds of directed evolution significantly stabilized the enzyme without lowering the catalytic efficiency and substrate specificity of the enzyme. Twenty-four variants with higher thermostability were identified. Detailed analysis revealed twelve single amino acid mutants that were found to possess a higher thermostability. The residues affected are scattered over the entire protein structure, and are different from mutations predicted by sequence alignment approaches, suggesting that sequence comparison and directed evolution methods are complementary strategies in engineering increased protein thermostability. The stabilizing mutations are predominately located at surface of the enzyme, suggesting that the protein's surface is important for stability. The directed evolution approach described in the present study is easily adapted to other histone modifying enzymes, requiring only appropriate peptide substrates and antibodies, which are available from commercial suppliers.

  8. A nucleosome turnover map reveals that the stability of histone H4 Lys20 methylation depends on histone recycling in transcribed chromatin.

    Science.gov (United States)

    Svensson, J Peter; Shukla, Manu; Menendez-Benito, Victoria; Norman-Axelsson, Ulrika; Audergon, Pauline; Sinha, Indranil; Tanny, Jason C; Allshire, Robin C; Ekwall, Karl

    2015-06-01

    Nucleosome composition actively contributes to chromatin structure and accessibility. Cells have developed mechanisms to remove or recycle histones, generating a landscape of differentially aged nucleosomes. This study aimed to create a high-resolution, genome-wide map of nucleosome turnover in Schizosaccharomyces pombe. The recombination-induced tag exchange (RITE) method was used to study replication-independent nucleosome turnover through the appearance of new histone H3 and the disappearance or preservation of old histone H3. The genome-wide location of histones was determined by chromatin immunoprecipitation-exonuclease methodology (ChIP-exo). The findings were compared with diverse chromatin marks, including histone variant H2A.Z, post-translational histone modifications, and Pol II binding. Finally, genome-wide mapping of the methylation states of H4K20 was performed to determine the relationship between methylation (mono, di, and tri) of this residue and nucleosome turnover. Our analysis showed that histone recycling resulted in low nucleosome turnover in the coding regions of active genes, stably expressed at intermediate levels. High levels of transcription resulted in the incorporation of new histones primarily at the end of transcribed units. H4K20 was methylated in low-turnover nucleosomes in euchromatic regions, notably in the coding regions of long genes that were expressed at low levels. This transcription-dependent accumulation of histone methylation was dependent on the histone chaperone complex FACT. Our data showed that nucleosome turnover is highly dynamic in the genome and that several mechanisms are at play to either maintain or suppress stability. In particular, we found that FACT-associated transcription conserves histones by recycling them and is required for progressive H4K20 methylation.

  9. Histone deacetylation is required for progression through mitosis in tobacco cells.

    Science.gov (United States)

    Li, Yan; Butenko, Yana; Grafi, Gideon

    2005-02-01

    Post-translational modifications of core histone proteins play a key role in chromatin structure and function. Here, we study histone post-translational modifications during reentry of protoplasts derived from tobacco mesophyll cells into the cell cycle and evaluate their significance for progression through mitosis. Methylation of histone H3 at lysine residues 4 and 9 persisted in chromosomes during all phases of the cell cycle. However, acetylation of H4 and H3 was dramatically reduced during mitosis in a stage-specific manner; while deacetylation of histone H4 commenced at prophase and persisted up to telophase, histone H3 remained acetylated up to metaphase but was deacetylated at anaphase and telophase. Phosphorylation of histone H3 at serine 10 was initiated at prophase, concomitantly with deacetylation of histone H4, and persisted up to telophase. Preventing histone deacetylation by the histone deacetylase inhibitor trichostatin A (TSA) led to accumulation of protoplasts at metaphase-anaphase, and reduced S10 phosphorylation during anaphase and telophase; in cultured tobacco cells, TSA significantly reduced the frequency of mitotic figures. Our results indicate that deacetylation of histone H4 and H3 in tobacco protoplasts occurs during mitosis in a phase-specific manner, and is important for progression through mitosis.

  10. Histones and lung cancer: Are the histone deacetylases a promising therapeutic target?

    Science.gov (United States)

    Petta, Vasiliki; Gkiozos, Ioannis; Strimpakos, Alex; Syrigos, Konstantinos

    2013-11-01

    Deoxyribonucleic acid is wrapped around an octamer of core histone proteins to form a nucleosome, the basic structure of chromatin. Two main families of enzymes maintain the equilibrium of acetyl groups added to or removed from lysine residues. Histone deacetylases (HDACs) catalyze the removal of acetyl groups from lysine residues in histone amino termini and non-histone proteins also, leading to chromatin condensation and transcriptional repression. HDAC overexpression, resulting in tumor suppressor genes silencing, has been found in several human cancer tissues, indicating that aberrant epigenetic activity is associated with cancer development. Therefore, inhibitors of these enzymes are emerging anticancer agents and there is evidence supporting their role in hematological malignancies. The minimal efficacy of conventional chemotherapy has prompted a renewed focus on targeted therapy based on pathways altered during the pathogenesis of lung cancer. We identify the pleiotropic antitumor effects of HDAC inhibitors in lung cancer, focusing on the result caused by their use individually, as well as in combination with other chemotherapeutic agents, in lung cancer cell lines and in clinical trials. We searched reviews and original papers in Pubmed over the last 10 years. We identified 76 original papers on this topic. Numerous preclinical studies have shown that HDAC inhibitors exhibit impressive antitumor activity in lung cancer cell lines. Nevertheless, Phase III randomized studies do not support HDAC inhibitors use in lung cancer patients in everyday practice. Ongoing and future studies would help determine their role in lung cancer treatment.

  11. On the origin of the histone fold

    Directory of Open Access Journals (Sweden)

    Söding Johannes

    2007-03-01

    Full Text Available Abstract Background Histones organize the genomic DNA of eukaryotes into chromatin. The four core histone subunits consist of two consecutive helix-strand-helix motifs and are interleaved into heterodimers with a unique fold. We have searched for the evolutionary origin of this fold using sequence and structure comparisons, based on the hypothesis that folded proteins evolved by combination of an ancestral set of peptides, the antecedent domain segments. Results Our results suggest that an antecedent domain segment, corresponding to one helix-strand-helix motif, gave rise divergently to the N-terminal substrate recognition domain of Clp/Hsp100 proteins and to the helical part of the extended ATPase domain found in AAA+ proteins. The histone fold arose subsequently from the latter through a 3D domain-swapping event. To our knowledge, this is the first example of a genetically fixed 3D domain swap that led to the emergence of a protein family with novel properties, establishing domain swapping as a mechanism for protein evolution. Conclusion The helix-strand-helix motif common to these three folds provides support for our theory of an 'ancient peptide world' by demonstrating how an ancestral fragment can give rise to 3 different folds.

  12. Histones and chromatin structure in hyperthermophilic Archaea.

    Science.gov (United States)

    Grayling, R A; Sandman, K; Reeve, J N

    1996-05-01

    HMf is a histone from the hyperthermophile Methanothermus fervidus. It is the archetype and most studied member of a family of archaeal histones that have primary sequences and three-dimensional structures in common with the eukaryal nucleosome core histones and that bind and compact DNA molecules into nucleosome-like structures (NLS). HMf preparations are mixtures of two similar, small (approximately 7.5 kDa) polypeptides designated HMfA and HMfB that in vivo form both homodimers and heterodimers. HMfA synthesis predominates during exponential growth but the relative amount of HMfB increases as M. fervidus cells enter the stationary growth phase. Analyses of homogeneous preparations of recombinant (r) (HMfA)2 and (rHMfB)2 have demonstrated that these proteins have different DNA-binding and compaction properties in vitro, consistent with different roles in vivo for the (HMfA)2, (HMfB)2 and HMfA. HmfB dimers, and for the NLS that they form, in regulating gene expression and in genome compaction and stability.

  13. Structural and Histone Binding Ability Characterizations of Human PWWP Domains

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Hong; Zeng, Hong; Lam, Robert; Tempel, Wolfram; Amaya, Maria F.; Xu, Chao; Dombrovski, Ludmila; Qiu, Wei; Wang, Yanming; Min, Jinrong (Toronto); (Penn)

    2013-09-25

    The PWWP domain was first identified as a structural motif of 100-130 amino acids in the WHSC1 protein and predicted to be a protein-protein interaction domain. It belongs to the Tudor domain 'Royal Family', which consists of Tudor, chromodomain, MBT and PWWP domains. While Tudor, chromodomain and MBT domains have long been known to bind methylated histones, PWWP was shown to exhibit histone binding ability only until recently. The PWWP domain has been shown to be a DNA binding domain, but sequence analysis and previous structural studies show that the PWWP domain exhibits significant similarity to other 'Royal Family' members, implying that the PWWP domain has the potential to bind histones. In order to further explore the function of the PWWP domain, we used the protein family approach to determine the crystal structures of the PWWP domains from seven different human proteins. Our fluorescence polarization binding studies show that PWWP domains have weak histone binding ability, which is also confirmed by our NMR titration experiments. Furthermore, we determined the crystal structures of the BRPF1 PWWP domain in complex with H3K36me3, and HDGF2 PWWP domain in complex with H3K79me3 and H4K20me3. PWWP proteins constitute a new family of methyl lysine histone binders. The PWWP domain consists of three motifs: a canonical {beta}-barrel core, an insertion motif between the second and third {beta}-strands and a C-terminal {alpha}-helix bundle. Both the canonical {beta}-barrel core and the insertion motif are directly involved in histone binding. The PWWP domain has been previously shown to be a DNA binding domain. Therefore, the PWWP domain exhibits dual functions: binding both DNA and methyllysine histones.

  14. Histone methyltransferases in cancer

    DEFF Research Database (Denmark)

    Albert, Mareike; Helin, Kristian

    2009-01-01

    Cancer is perceived as a heterogeneous group of diseases that is characterized by aberrant patterns of gene expression. In the last decade, an increasing amount of data has pointed to a key role for epigenetic alterations in human cancer. In this review, we focus on a subclass of epigenetic...... regulators, namely histone methyltransferases (HMTs). Several HMTs have been linked to different types of cancer; however, in most cases we only have limited knowledge regarding the molecular mechanisms by which the HMTs contribute to disease development. We summarize the current knowledge regarding some...

  15. Histone Deacetylases and Cardiometabolic Diseases

    OpenAIRE

    Yiew, Kan Hui; Chatterjee, Tapan K.; Hui, David Y.; Weintraub, Neal L.

    2015-01-01

    Cardiometabolic disease, emerging as a worldwide epidemic, is a combination of metabolic derangements leading to type 2 diabetes and cardiovascular disease. Genetic and environmental factors are linked through epigenetic mechanisms to the pathogenesis of cardiometabolic disease. Post-translational modifications of histone tails, including acetylation and deacetylation, epigenetically alter chromatin structure and dictate cell-specific gene expression patterns. The histone deacetylase (HDAC) f...

  16. Cellulase variants

    Energy Technology Data Exchange (ETDEWEB)

    Blazej, Robert; Toriello, Nicholas; Emrich, Charles; Cohen, Richard N.; Koppel, Nitzan

    2015-07-14

    This invention provides novel variant cellulolytic enzymes having improved activity and/or stability. In certain embodiments the variant cellulotyic enzymes comprise a glycoside hydrolase with or comprising a substitution at one or more positions corresponding to one or more of residues F64, A226, and/or E246 in Thermobifida fusca Cel9A enzyme. In certain embodiments the glycoside hydrolase is a variant of a family 9 glycoside hydrolase. In certain embodiments the glycoside hydrolase is a variant of a theme B family 9 glycoside hydrolase.

  17. Mutagenesis of tGCN5 core region reveals two critical surface residues F90 and R140

    Energy Technology Data Exchange (ETDEWEB)

    Mehta, Kinjal Rajesh; Chan, Yan M.; Lee, Man X.; Yang, Ching Yao; Voloshchuk, Natalya [Department of Chemical and Biological Sciences, Polytechnic Institute of New York University, 6 MetroTech Center, Brooklyn, NY 11201 (United States); Montclare, Jin Kim, E-mail: jmontcla@poly.edu [Department of Chemical and Biological Sciences, Polytechnic Institute of New York University, 6 MetroTech Center, Brooklyn, NY 11201 (United States); Department of Biochemistry, SUNY-Downstate Medical Center, 450 Clarkson Avenue, Brooklyn, NY 11203 (United States)

    2010-09-24

    Research highlights: {yields} Mutagenesis of the tGCN5 core region reveals two residues important for function. {yields} Developed a fluorescent lysate-based activity assay to assess mutants. {yields} Surface-exposed residues F90 and R140 of tGCN5 are critical for H3 acetylation. -- Abstract: Tetrahymena General Control Non-Derepressor 5 (tGCN5) is a critical regulator of gene transcription via acetylation of histones. Since the acetylation ability has been attributed to the 'core region', we perform mutagenesis of residues within the tGCN5 'core region' in order to identify those critical for function and stability. Residues that do not participate in catalysis are identified, mutated and characterized for activity, structure and thermodynamic stability. Variants I107V, Q114L, A121T and A130S maintain the acetylation function relative to wild-type tGCN5, while variants F90Y, F112R and R140H completely abolish function. Of the three non-functional variants, since F112 is mutated into a non-homologous charged residue, a loss in function is expected. However, the remaining two variants are mutated into homologous residues, suggesting that F90 and R140 are critical for the activity of tGCN5. While mutation to homologous residue maintains acetylation of histone H3 for the majority of the variants, the two surface-exposed residues, F90 and R140, appear to be essential for tGCN5 function, structure or stability.

  18. Recognition and classification of histones using support vector machine

    OpenAIRE

    Bhasin, Manoj; Reinherz, Ellis L.; Reche, Pedro A.

    2006-01-01

    Histones are DNA-binding proteins found in the chromatin of all eukaryotic cells. They are highly conserved and can be grouped into five major classes: H1/H5, H2A, H2B, H3, and H4. Two copies of H2A, H2B, H3, and H4 bind to about 160 base pairs of DNA forming the core of the nucleosome (the repeating structure of chromatin) and H1/H5 bind to its DNA linker sequence. Overall, histones have a high arginine/lysine content that is optimal for interaction with DNA. This sequence bias can make the ...

  19. Direct interplay among histones, histone chaperones, and a chromatin boundary protein in the control of histone gene expression.

    Science.gov (United States)

    Zunder, Rachel M; Rine, Jasper

    2012-11-01

    In Saccharomyces cerevisiae, the histone chaperone Rtt106 binds newly synthesized histone proteins and mediates their delivery into chromatin during transcription, replication, and silencing. Rtt106 is also recruited to histone gene regulatory regions by the HIR histone chaperone complex to ensure S-phase-specific expression. Here we showed that this Rtt106:HIR complex included Asf1 and histone proteins. Mutations in Rtt106 that reduced histone binding reduced Rtt106 enrichment at histone genes, leading to their increased transcription. Deletion of the chromatin boundary element Yta7 led to increased Rtt106:H3 binding, increased Rtt106 enrichment at histone gene regulatory regions, and decreased histone gene transcription at the HTA1-HTB1 locus. These results suggested a unique regulatory mechanism in which Rtt106 sensed the level of histone proteins to maintain the proper level of histone gene transcription. The role of these histone chaperones and Yta7 differed markedly among the histone gene loci, including the two H3-H4 histone gene pairs. Defects in silencing in rtt106 mutants could be partially accounted for by Rtt106-mediated changes in histone gene repression. These studies suggested that feedback mediated by histone chaperone complexes plays a pivotal role in regulating histone gene transcription.

  20. Histone-modifying enzymes, histone modifications and histone chaperones in nucleosome assembly: Lessons learned from Rtt109 histone acetyltransferases.

    Science.gov (United States)

    Dahlin, Jayme L; Chen, Xiaoyue; Walters, Michael A; Zhang, Zhiguo

    2015-01-01

    During DNA replication, nucleosomes ahead of replication forks are disassembled to accommodate replication machinery. Following DNA replication, nucleosomes are then reassembled onto replicated DNA using both parental and newly synthesized histones. This process, termed DNA replication-coupled nucleosome assembly (RCNA), is critical for maintaining genome integrity and for the propagation of epigenetic information, dysfunctions of which have been implicated in cancers and aging. In recent years, it has been shown that RCNA is carefully orchestrated by a series of histone modifications, histone chaperones and histone-modifying enzymes. Interestingly, many features of RCNA are also found in processes involving DNA replication-independent nucleosome assembly like histone exchange and gene transcription. In yeast, histone H3 lysine K56 acetylation (H3K56ac) is found in newly synthesized histone H3 and is critical for proper nucleosome assembly and for maintaining genomic stability. The histone acetyltransferase (HAT) regulator of Ty1 transposition 109 (Rtt109) is the sole enzyme responsible for H3K56ac in yeast. Much research has centered on this particular histone modification and histone-modifying enzyme. This Critical Review summarizes much of our current understanding of nucleosome assembly and highlights many important insights learned from studying Rtt109 HATs in fungi. We highlight some seminal features in nucleosome assembly conserved in mammalian systems and describe some of the lingering questions in the field. Further studying fungal and mammalian chromatin assembly may have important public health implications, including deeper understandings of human cancers and aging as well as the pursuit of novel anti-fungal therapies.

  1. Modification of histones by sugar β-N-acetylglucosamine (GlcNAc) occurs on multiple residues, including histone H3 serine 10, and is cell cycle-regulated.

    Science.gov (United States)

    Zhang, Suisheng; Roche, Kevin; Nasheuer, Heinz-Peter; Lowndes, Noel Francis

    2011-10-28

    The monosaccharide, β-N-acetylglucosamine (GlcNAc), can be added to the hydroxyl group of either serines or threonines to generate an O-linked β-N-acetylglucosamine (O-GlcNAc) residue (Love, D. C., and Hanover, J. A. (2005) Sci. STKE 2005 312, 1-14; Hart, G. W., Housley, M. P., and Slawson, C. (2007) Nature 446, 1017-1022). This post-translational protein modification, termed O-GlcNAcylation, is reversible, analogous to phosphorylation, and has been implicated in many cellular processes. Here, we present evidence that in human cells all four core histones of the nucleosome are substrates for this glycosylation in the relative abundance H3, H4/H2B, and H2A. Increasing the intracellular level of UDP-GlcNAc, the nucleotide sugar donor substrate for O-GlcNAcylation enhanced histone O-GlcNAcylation and partially suppressed phosphorylation of histone H3 at serine 10 (H3S10ph). Expression of recombinant H3.3 harboring an S10A mutation abrogated histone H3 O-GlcNAcylation relative to its wild-type version, consistent with H3S10 being a site of histone O-GlcNAcylation (H3S10glc). Moreover, O-GlcNAcylated histones were lost from H3S10ph immunoprecipitates, whereas immunoprecipitation of either H3K4me3 or H3K9me3 (active or inactive histone marks, respectively) resulted in co-immunoprecipitation of O-GlcNAcylated histones. We also examined histone O-GlcNAcylation during cell cycle progression. Histone O-GlcNAcylation is high in G(1) cells, declines throughout the S phase, increases again during late S/early G(2), and persists through late G(2) and mitosis. Thus, O-GlcNAcylation is a novel histone post-translational modification regulating chromatin conformation during transcription and cell cycle progression.

  2. Epigenetic regulation: methylation of histone and non-histone proteins

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Histone methylation is believed to play important roles in epigenetic memory in various biological processes. However, questions like whether the methylation marks themselves are faithfully transmit- ted into daughter cells and through what mechanisms are currently under active investigation. Previ- ously, methylation was considered to be irreversible, but the recent discovery of histone lysine de- methylases revealed a dynamic nature of histone methylation regulation on four of the main sites of methylation on histone H3 and H4 tails (H3K4, H3K9, H3K27 and H3K36). Even so, it is still unclear whether demethylases specific for the remaining two sites, H3K79 and H4K20, exist. Furthermore, be- sides histone proteins, the lysine methylation and demethylation also occur on non-histone proteins, which are probably subjected to similar regulation as histones. This review discusses recent pro- gresses in protein lysine methylation regulation focusing on the above topics, while referring readers to a number of recent reviews for the biochemistry and biology of these enzymes.

  3. Histone Acetylation near the Nucleosome Dyad Axis Enhances Nucleosome Disassembly by RSC and SWI/SNF.

    Science.gov (United States)

    Chatterjee, Nilanjana; North, Justin A; Dechassa, Mekonnen Lemma; Manohar, Mridula; Prasad, Rashmi; Luger, Karolin; Ottesen, Jennifer J; Poirier, Michael G; Bartholomew, Blaine

    2015-12-01

    Signaling associated with transcription activation occurs through posttranslational modification of histones and is best exemplified by lysine acetylation. Lysines are acetylated in histone tails and the core domain/lateral surface of histone octamers. While acetylated lysines in histone tails are frequently recognized by other factors referred to as "readers," which promote transcription, the mechanistic role of the modifications in the lateral surface of the histone octamer remains unclear. By using X-ray crystallography, we found that acetylated lysines 115 and 122 in histone H3 are solvent accessible, but in biochemical assays they appear not to interact with the bromodomains of SWI/SNF and RSC to enhance recruitment or nucleosome mobilization, as previously shown for acetylated lysines in H3 histone tails. Instead, we found that acetylation of lysines 115 and 122 increases the predisposition of nucleosomes for disassembly by SWI/SNF and RSC up to 7-fold, independent of bromodomains, and only in conjunction with contiguous nucleosomes. Thus, in combination with SWI/SNF and RSC, acetylation of lateral surface lysines in the histone octamer serves as a crucial regulator of nucleosomal dynamics distinct from the histone code readers and writers.

  4. Synthesis of histone proteins by CPE ligation using a recombinant peptide as the C-terminal building block.

    Science.gov (United States)

    Kawakami, Toru; Yoshikawa, Ryo; Fujiyoshi, Yuki; Mishima, Yuichi; Hojo, Hironobu; Tajima, Shoji; Suetake, Isao

    2015-11-01

    The post-translational modification of histones plays an important role in gene expression. We report herein on a method for synthesizing such modified histones by ligating chemically prepared N-terminal peptides and C-terminal recombinant peptide building blocks. Based on their chemical synthesis, core histones can be categorized as two types; histones H2A, H2B and H4 which contain no Cys residues, and histone H3 which contains a Cys residue(s) in the C-terminal region. A combination of native chemical ligation and desulphurization can be simply used to prepare histones without Cys residues. For the synthesis of histone H3, the endogenous Cys residue(s) must be selectively protected, while keeping the N-terminal Cys residue of the C-terminal building block that is introduced for purposes of chemical ligation unprotected. To this end, a phenacyl group was successfully utilized to protect endogenous Cys residue(s), and the recombinant peptide was ligated with a peptide containing a Cys-Pro ester (CPE) sequence as a thioester precursor. Using this approach it was possible to prepare all of the core histones H2A, H2B, H3 and H4 with any modifications. The resulting proteins could then be used to prepare a core histone library of proteins that have been post-translationally modified.

  5. Novel E3 ubiquitin ligases that regulate histone protein levels in the budding yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Rakesh Kumar Singh

    Full Text Available Core histone proteins are essential for packaging the genomic DNA into chromatin in all eukaryotes. Since multiple genes encode these histone proteins, there is potential for generating more histones than what is required for chromatin assembly. The positively charged histones have a very high affinity for negatively charged molecules such as DNA, and any excess of histone proteins results in deleterious effects on genomic stability and cell viability. Hence, histone levels are known to be tightly regulated via transcriptional, posttranscriptional and posttranslational mechanisms. We have previously elucidated the posttranslational regulation of histone protein levels by the ubiquitin-proteasome pathway involving the E2 ubiquitin conjugating enzymes Ubc4/5 and the HECT (Homologous to E6-AP C-Terminus domain containing E3 ligase Tom1 in the budding yeast. Here we report the identification of four additional E3 ligases containing the RING (Really Interesting New Gene finger domains that are involved in the ubiquitylation and subsequent degradation of excess histones in yeast. These E3 ligases are Pep5, Snt2 as well as two previously uncharacterized Open Reading Frames (ORFs YKR017C and YDR266C that we have named Hel1 and Hel2 (for Histone E3 Ligases respectively. Mutants lacking these E3 ligases are sensitive to histone overexpression as they fail to degrade excess histones and accumulate high levels of endogenous histones on histone chaperones. Co-immunoprecipitation assays showed that these E3 ligases interact with the major E2 enzyme Ubc4 that is involved in the degradation related ubiquitylation of histones. Using mutagenesis we further demonstrate that the RING domains of Hel1, Hel2 and Snt2 are required for histone regulation. Lastly, mutants corresponding to Hel1, Hel2 and Pep5 are sensitive to replication inhibitors. Overall, our results highlight the importance of posttranslational histone regulatory mechanisms that employ multiple E3

  6. Histone deacetylase inhibition as an alternative strategy against invasive aspergillosis

    Directory of Open Access Journals (Sweden)

    Frederic eLamoth

    2015-02-01

    Full Text Available Invasive aspergillosis (IA is a life-threatening infection due to Aspergillus fumigatus and other Aspergillus spp. Drugs targeting the fungal cell membrane (triazoles, amphotericin B or cell wall (echinocandins are currently the sole therapeutic options against IA. Their limited efficacy and the emergence of resistance warrant the identification of new antifungal targets. Histone deacetylases (HDACs are enzymes responsible of the deacetylation of lysine residues of core histones, thus controlling chromatin remodeling and transcriptional activation. HDACs also control the acetylation and activation status of multiple non-histone proteins, including the heat shock protein 90 (Hsp90, an essential molecular chaperone for fungal virulence and antifungal resistance. This review provides an overview of the different HDACs in Aspergillus spp. as well as their respective contribution to total HDAC activity, fungal growth, stress responses, and virulence. The potential of HDAC inhibitors, currently under development for cancer therapy, as novel alternative antifungal agents against IA is discussed.

  7. Why are histones dynamic?

    OpenAIRE

    Ferrari, Paolo

    2015-01-01

    Afin de pouvoir être contenu dans le noyau des cellules eucaryotes, l’ADN doit être compacté en un ensemble très structuré. L’ADN eucaryote est généralement associé à des protéines, principalement à des histones, pour former cette structure très compact appelée chromatine. La chromatine régule de nombreux processus cellulaires de bases reposant sur les interactions ADN-protéines, y compris la transcription des gènes. La liaison des facteurs de transcription aux séquences régulatrices des gène...

  8. Replication stress interferes with histone recycling and predeposition marking of new histones

    DEFF Research Database (Denmark)

    Jasencakova, Zuzana; Scharf, Annette N D; Ask, Katrine

    2010-01-01

    To restore chromatin on new DNA during replication, recycling of histones evicted ahead of the fork is combined with new histone deposition. The Asf1 histone chaperone, which buffers excess histones under stress, is a key player in this process. Yet how histones handled by human Asf1 are modified...

  9. Chromatin dynamics: interplay between remodeling enzymes and histone modifications.

    Science.gov (United States)

    Swygert, Sarah G; Peterson, Craig L

    2014-08-01

    Chromatin dynamics play an essential role in regulating the accessibility of genomic DNA for a variety of nuclear processes, including gene transcription and DNA repair. The posttranslational modification of the core histones and the action of ATP-dependent chromatin remodeling enzymes represent two primary mechanisms by which chromatin dynamics are controlled and linked to nuclear events. Although there are examples in which a histone modification or a remodeling enzyme may be sufficient to drive a chromatin transition, these mechanisms typically work in concert to integrate regulatory inputs, leading to a coordinated alteration in chromatin structure and function. Indeed, site-specific histone modifications can facilitate the recruitment of chromatin remodeling enzymes to particular genomic regions, or they can regulate the efficiency or the outcome of a chromatin remodeling reaction. Conversely, chromatin remodeling enzymes can also influence, and sometimes directly modulate, the modification state of histones. These functional interactions are generally complex, frequently transient, and often require the association of myriad additional factors. This article is part of a Special Issue entitled: Molecular mechanisms of histone modification function. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Identification of novel post-translational modifications in linker histones from chicken erythrocytes.

    Science.gov (United States)

    Sarg, Bettina; Lopez, Rita; Lindner, Herbert; Ponte, Inma; Suau, Pedro; Roque, Alicia

    2015-01-15

    Chicken erythrocyte nuclei were digested with micrococcal nuclease and fractionated by centrifugation in low-salt buffer into soluble and insoluble fractions. Post-translational modifications of the purified linker histones of both fractions were analyzed by LC-ESI-MS/MS. All six histone H1 subtypes (H1.01, H1.02, H1.03, H1.10, H1.1L and H1.1R) and histone H5 were identified. Mass spectrometry analysis enabled the identification of a wide range of PTMs, including N(α)-terminal acetylation, acetylation, formylation, phosphorylation and oxidation. A total of nine new modifications in chicken linker histones were mapped, most of them located in the N-terminal and globular domains. Relative quantification of the modified peptides showed that linker histone PTMs were differentially distributed among both chromatin fractions, suggesting their relevance in the regulation of chromatin structure. The analysis of our results combined with previously reported data for chicken and some mammalian species showed that most of the modified positions were conserved throughout evolution, highlighting their importance in specific linker histone functions and epigenetics. Post-translational modifications of linker histones could have a role in the regulation of gene expression through the modulation of chromatin higher-order structure and chromatin remodeling. Finding new PTMs in linker histones is the first step to elucidate their role in the histone code. In this manuscript we report nine new post-translational modifications of the linker histones from chicken erythrocytes, one in H5 and eight in the H1 subtypes. Chromatin fractionated by centrifugation in low-salt buffer resulted in two fractions with different contents and compositions of linker histones and enriched in specific core histone PTMs. Of particular interest is the fact that linker histone PTMs were differentially distributed in both chromatin fractions, suggesting specific functions. Future studies are needed to

  11. Enhanced stability of histone octamers from plant nucleosomes: role of H2A and H2B histones.

    Science.gov (United States)

    Moehs, C P; Baxevanis, A D; Moudrianakis, E N; Spiker, S

    1992-11-10

    Gel filtration and sedimentation studies have previously established that the vertebrate animal core histone octamer is in equilibrium with an (H3-H4)2 tetramer and an H2A-H2B dimer [Eickbush, T. H., & Moudrianakis, E. N. (1978) Biochemistry 17, 4955-4964; Godfrey, J. E., Eickbush, T. H., & Moudrianakis, E. N. (1980) Biochemistry 19, 1339-1346]. We have investigated the core histone octamer of wheat (Triticum aestivum L.) and have found it to be much more stable than its vertebrate animal counterpart. When vertebrate animal histone octamers are subjected to gel filtration in 2 M NaCl, a trailing peak of H2A-H2B dimer can be clearly resolved from the main octamer peak. When the plant octamer is subjected to the identical procedure, there is no trailing peak of H2A-H2B dimer, but rather a single peak containing the octamer. A sampling across the octamer peak from leading to trailing edge shows no change in the ratio of H2A-H2B to (H3-H4)2. Surprisingly, the plant octamer shows the same stability at 0.6 M NaCl, a salt concentration in which the vertebrate animal octamer dissociates into dimers and tetramers. Equilibrium sedimentation data indicate that the assembly potential of the wheat histones in 2 M NaCl is very high at all protein concentrations above 0.1 mg mL-1. In order to disrupt the forces stabilizing the plant histone octamer at high histone concentrations, the concentration of NaCl must be lowered to approximately 0.3 M.(ABSTRACT TRUNCATED AT 250 WORDS)

  12. Epigenetic regulation: methylation of histone and non-histone proteins

    Institute of Scientific and Technical Information of China (English)

    LAN Fei; SHI Yang

    2009-01-01

    Histone methylation is believed to play important roles In epigenetic memory in various biological processes. However, questions like whether the methylation marks themselves are faithfully transmit-ted into daughter cells and through what mechanisms are currently under active investigation. Previ-ously, methylation was considered to be irreversible, but the recent discovery of histone lysine de-methylases revealed a dynamic nature of histone methylation regulation on four of the main sites of methylation on histone H3 and H4 tails (H3K4, H3K9, H3K27 and H3K36). Even so, it is stlll unclear whether demethylases specific for the remaining two sites, H3K79 and H4K20, exist. Furthermore, be-sides hlstone proteins, the lysine methylation and demethylation also occur on non-histone proteins,which are probably subjected to similar regulation as histones. This review discusses recent pro-gresses In protein lysine methylation regulation focusing on the above topics, while referring readers to a number of recent reviews for the biochemistry and biology of these enzymes.

  13. Progressive methylation of ageing histones by Dot1 functions as a timer.

    Science.gov (United States)

    De Vos, Dirk; Frederiks, Floor; Terweij, Marit; van Welsem, Tibor; Verzijlbergen, Kitty F; Iachina, Ekaterina; de Graaf, Erik L; Altelaar, A F Maarten; Oudgenoeg, Gideon; Heck, Albert J R; Krijgsveld, Jeroen; Bakker, Barbara M; van Leeuwen, Fred

    2011-09-01

    Post-translational modifications of histone proteins have a crucial role in regulating gene expression. If efficiently re-established after chromosome duplication, histone modifications could help propagate gene expression patterns in dividing cells by epigenetic mechanisms. We used an integrated approach to investigate the dynamics of the conserved methylation of histone H3 Lys 79 (H3K79) by Dot1. Our results show that methylation of H3K79 progressively changes after histone deposition, which is incompatible with a rapid copy mechanism. Instead, methylation accumulates on ageing histones, providing the cell with a timer mechanism to directly couple cell-cycle length to changes in chromatin modification on the nucleosome core.

  14. Balancing chromatin remodeling and histone modifications in transcription.

    Science.gov (United States)

    Petty, Emily; Pillus, Lorraine

    2013-11-01

    Chromatin remodelers use the energy of ATP hydrolysis to reposition or evict nucleosomes or to replace canonical histones with histone variants. By regulating nucleosome dynamics, remodelers gate access to the underlying DNA for replication, repair, and transcription. Nucleosomes are subject to extensive post-translational modifications that can recruit regulatory proteins or alter the local chromatin structure. Just as extensive crosstalk has been observed between different histone post-translational modifications, there is growing evidence for both coordinated and antagonistic functional relations between nucleosome remodeling and modifying machineries. Defining the combined functions of the complexes that alter nucleosome interactions, position, and stability is key to understanding processes that require access to DNA, particularly with growing appreciation of their contributions to human health and disease. Here, we highlight recent advances in the interactions between histone modifications and the imitation-switch (ISWI) and chromodomain helicase DNA-binding protein 1 (CHD1) chromatin remodelers from studies in budding yeast, fission yeast, flies, and mammalian cells, with a focus on yeast.

  15. The organization structure and regulatory elements of Chlamydomonas histone genes reveal features linking plant and animal genes.

    Science.gov (United States)

    Fabry, S; Müller, K; Lindauer, A; Park, P B; Cornelius, T; Schmitt, R

    1995-09-01

    The genome of the green alga Chlamydomonas reinhardtii contains approximately 15 gene clusters of the nucleosomal (or core) histone H2A, H2B, H3 and H4 genes and at least one histone H1 gene. Seven non-allelic histone gene loci were isolated from a genomic library, physically mapped, and the nucleotide sequences of three isotypes of each core histone gene species and one linked H1 gene determined. The core histone genes are organized in clusters of H2A-H2B and H3-H4 pairs, in which each gene pair shows outwardly divergent transcription from a short (< 300 bp) intercistronic region. These intercistronic regions contain typically conserved promoter elements, namely a TATA-box and the three motifs TGGCCAG-G(G/C)-CGAG, CGTTGACC and CGGTTG. Different from the genes of higher plants, but like those of animals and the related alga Volvox, the 3' untranslated regions contain no poly A signal, but a palindromic sequence (3' palindrome) essential for mRNA processing is present. One single H1 gene was found in close linkage to a H2A-H2B pair. The H1 upstream region contains the octameric promoter element GGTTGACC (also found upstream of the core histone genes) and two specific sequence motifs that are shared only with the Volvox H1 promoters. This suggests differential transcription of the H1 and the core histone genes. The H1 gene is interrupted by two introns. Unlike Volvox H3 genes, the three sequenced H3 isoforms are intron-free. Primer-directed PCR of genomic DNA demonstrated, however, that at least 8 of the about 15 H3 genes do contain one intron at a conserved position. In synchronized C. reinhardtii cells, H4 mRNA levels (representative of all core histone mRNAs) peak during cell division, suggesting strict replication-dependent gene control. The derived peptide sequences place C. reinhardtii core histones closer to plants than to animals, except that the H2A histones are more animal-like. The peptide sequence of histone H1 is closely related to the V. carteri VH1-II

  16. Imaging local deposition of newly synthesized histones in UVC-damaged chromatin.

    Science.gov (United States)

    Adam, Salomé; Dabin, Juliette; Bai, Siau-Kun; Polo, Sophie E

    2015-01-01

    DNA damage not only jeopardizes genome integrity but also challenges the well-organized association of DNA with histone proteins into chromatin, which is key for regulating gene expression and cell functions. The extent to which the original chromatin structure is altered after repair of DNA lesions is thus a critical issue. Dissecting histone dynamics at sites of DNA damage has provided mechanistic insights into chromatin plasticity in response to genotoxic stress. Here, we present an experimental protocol for visualizing the deposition of newly synthesized histone H3 variants at sites of UVC damage in human cells that couples SNAP-tag based labeling of new histones with local UVC irradiation of cells through micropore filters.

  17. Drug-induced histone eviction from open chromatin contributes to the chemotherapeutic effects of doxorubicin

    Science.gov (United States)

    Pang, Baoxu; Qiao, Xiaohang; Janssen, Lennert; Velds, Arno; Groothuis, Tom; Kerkhoven, Ron; Nieuwland, Marja; Ovaa, Huib; Rottenberg, Sven; van Tellingen, Olaf; Janssen, Jeroen; Huijgens, Peter; Zwart, Wilbert; Neefjes, Jacques

    2013-01-01

    DNA topoisomerase II inhibitors are a major class of cancer chemotherapeutics, which are thought to eliminate cancer cells by inducing DNA double-strand breaks. Here we identify a novel activity for the anthracycline class of DNA topoisomerase II inhibitors: histone eviction from open chromosomal areas. We show that anthracyclines promote histone eviction irrespective of their ability to induce DNA double-strand breaks. The histone variant H2AX, which is a key component of the DNA damage response, is also evicted by anthracyclines, and H2AX eviction is associated with attenuated DNA repair. Histone eviction deregulates the transcriptome in cancer cells and organs such as the heart, and can drive apoptosis of topoisomerase-negative acute myeloid leukaemia blasts in patients. We define a novel mechanism of action of anthracycline anticancer drugs doxorubicin and daunorubicin on chromatin biology, with important consequences for DNA damage responses, epigenetics, transcription, side effects and cancer therapy. PMID:23715267

  18. Communication of brain network core connections altered in behavioral variant frontotemporal dementia but possibly preserved in early-onset Alzheimer's disease

    Science.gov (United States)

    Daianu, Madelaine; Jahanshad, Neda; Mendez, Mario F.; Bartzokis, George; Jimenez, Elvira E.; Thompson, Paul M.

    2015-03-01

    Diffusion imaging and brain connectivity analyses can assess white matter deterioration in the brain, revealing the underlying patterns of how brain structure declines. Fiber tractography methods can infer neural pathways and connectivity patterns, yielding sensitive mathematical metrics of network integrity. Here, we analyzed 1.5-Tesla wholebrain diffusion-weighted images from 64 participants - 15 patients with behavioral variant frontotemporal dementia (bvFTD), 19 with early-onset Alzheimer's disease (EOAD), and 30 healthy elderly controls. Using whole-brain tractography, we reconstructed structural brain connectivity networks to map connections between cortical regions. We evaluated the brain's networks focusing on the most highly central and connected regions, also known as hubs, in each diagnostic group - specifically the "high-cost" structural backbone used in global and regional communication. The high-cost backbone of the brain, predicted by fiber density and minimally short pathways between brain regions, accounted for 81-92% of the overall brain communication metric in all diagnostic groups. Furthermore, we found that the set of pathways interconnecting high-cost and high-capacity regions of the brain's communication network are globally and regionally altered in bvFTD, compared to healthy participants; however, the overall organization of the high-cost and high-capacity networks were relatively preserved in EOAD participants, relative to controls. Disruption of the major central hubs that transfer information between brain regions may impair neural communication and functional integrity in characteristic ways typical of each subtype of dementia.

  19. Stage-specific histone modification profiles reveal global transitions in the Xenopus embryonic epigenome.

    Directory of Open Access Journals (Sweden)

    Tobias D Schneider

    Full Text Available Vertebrate embryos are derived from a transitory pool of pluripotent cells. By the process of embryonic induction, these precursor cells are assigned to specific fates and differentiation programs. Histone post-translational modifications are thought to play a key role in the establishment and maintenance of stable gene expression patterns underlying these processes. While on gene level histone modifications are known to change during differentiation, very little is known about the quantitative fluctuations in bulk histone modifications during development. To investigate this issue we analysed histones isolated from four different developmental stages of Xenopus laevis by mass spectrometry. In toto, we quantified 59 modification states on core histones H3 and H4 from blastula to tadpole stages. During this developmental period, we observed in general an increase in the unmodified states, and a shift from histone modifications associated with transcriptional activity to transcriptionally repressive histone marks. We also compared these naturally occurring patterns with the histone modifications of murine ES cells, detecting large differences in the methylation patterns of histone H3 lysines 27 and 36 between pluripotent ES cells and pluripotent cells from Xenopus blastulae. By combining all detected modification transitions we could cluster their patterns according to their embryonic origin, defining specific histone modification profiles (HMPs for each developmental stage. To our knowledge, this data set represents the first compendium of covalent histone modifications and their quantitative flux during normogenesis in a vertebrate model organism. The HMPs indicate a stepwise maturation of the embryonic epigenome, which may be causal to the progressing restriction of cellular potency during development.

  20. Enhanced top-down characterization of histone post-translational modifications

    Energy Technology Data Exchange (ETDEWEB)

    Tian, Zhixin; Tolić, Nikola; Zhao, Rui; Moore, Ronald J.; Hengel, Shawna M.; Robinson, Errol W.; Stenoien, David L.; Wu, Si; Smith, Richard D.; Paša-Tolić, Ljiljana

    2012-01-01

    Background: Multiple post-translational modifications (PTMs) on core histones often work synergistically to fine tune chromatin structure and functions, generating a “histone code” that can be interpreted by a variety of chromatin interacting proteins. Although previous bottom-up and middle-down proteomic approaches have been developed for limited characterization of PTMs on histone N-terminal tails, high-throughput methods for comprehensive identification of PTMs distributed along the entire primary amino acid sequence are yet to be implemented. Results: Here we report a novel online two-dimensional liquid chromatography - tandem mass spectrometry (2D LC–MS/MS) platform for high-throughput and sensitive characterization of histone PTMs at the intact protein level. The metal-free LC system with reverse phase separation followed by weak cation exchange – hydrophilic interaction chromatography (WCX-HILIC) and online Orbitrap Velos tandem mass spectrometry allowed for unambiguous identification of over 700 histone isoforms from a single 2D LC–MS/MS analysis of 7.5 µg of purified core histones. In comparison with previous offline top-down analysis of H4, this online study identified 100 additional isoforms from 100-fold less sample. This platform enabled comprehensive characterization of histone modifications, including those beyond tail regions, with dramatically improved throughput and sensitivity compared to more traditional platforms. Isoforms identified included those with combinatorial PTMs extending well beyond the N-terminal tail regions as well as a large number of phosphorylated isoforms.

  1. Local compartment changes and regulatory landscape alterations in histone H1-depleted cells

    NARCIS (Netherlands)

    Geeven, Geert; Zhu, Yun; Kim, Byung Ju; Bartholdy, Boris A; Yang, Seung-Min; Macfarlan, Todd S; Gifford, Wesley D; Pfaff, Samuel L; Verstegen, Marjon J A M; Pinto, Hugo; Vermunt, Marit W; Creyghton, Menno P; Wijchers, Patrick J; Stamatoyannopoulos, John A; Skoultchi, Arthur I; de Laat, Wouter|info:eu-repo/dai/nl/169934497

    2015-01-01

    BACKGROUND: Linker histone H1 is a core chromatin component that binds to nucleosome core particles and the linker DNA between nucleosomes. It has been implicated in chromatin compaction and gene regulation and is anticipated to play a role in higher-order genome structure. Here we have used a combi

  2. spFRET studies of nucleosome dynamics modulated by histone modifications, histone variants and neighboring nucleosomes

    NARCIS (Netherlands)

    Buning, Ruth

    2015-01-01

    At the basis of the regulation of the genetic code (DNA) in eukaryotes is its organization into nucleosomes. Nucleosomes modulate DNA accessibility through conformational dynamics like DNA breathing - the transient unwrapping of DNA from the nucleosome. Single-pair Fluorescence Resonance Energy Tran

  3. The role of histones in the immune responses of aquatic invertebrates

    Directory of Open Access Journals (Sweden)

    C Nikapitiya

    2013-10-01

    Full Text Available Histones are primary components of eukaryotic chromatin and highly abundant in all animal cells. In addition to their important role in chromatin structure and transcriptional regulation, histones contribute to innate immune responses. In several aquatic invertebrate species, as well as in many other invertebrate and vertebrate species, the transcripts for core histones are upregulated in response to immune challenge and exposure to environmental stressors. Histones show antimicrobial activity against bacteria and parasites in vitro and in vivo and have the ability to bind bacterial lipopolysaccharide and other pathogen-associated molecules. Several mechanisms regulating and facilitating the antimicrobial action of histones against pathogens have been described in vertebrate and some invertebrate species, including the production of Extracellular Traps (ETs and the accumulation of histones in lipid droplets that can be selectively released in response to immune stimuli. Further studies are needed to determine the mechanisms of action of histones in immune responses in aquatic invertebrates and investigate the potential use of histones in the treatment of infectious diseases in aquaculture

  4. Changes in nuclear content of protein conjugate histone H2A-ubiquitin during rooster spermatogenesis.

    Science.gov (United States)

    Agell, N; Chiva, M; Mezquita, C

    1983-05-08

    Electrophoretic analysis of acid-soluble chromosomal proteins isolated from rooster testis cell nuclei at different stages of spermatogenesis, revealed that the nuclear content of a protein identified by its solubility, electrophoretic mobility and amino acid analysis as the protein conjugate histone H2A-ubiquitin (uH2A, A24) changed markedly from meiotic cells to late spermatids. The protein was not detectable in tetraploid primary spermatocytes; it was present in 1.7% of the total amount of nucleosomal core histones in early spermatids and reached its maximum level (3.5% and 11%) at the end of spermiogenesis, when histones are replaced by the protamine galline.

  5. A novel, enigmatic histone modification: biotinylation of histones by holocarboxylase synthetase.

    Science.gov (United States)

    Hassan, Yousef I; Zempleni, Janos

    2008-12-01

    Holocarboxylase synthetase catalyzes the covalent binding of biotin to histones in humans and other eukaryotes. Eleven biotinylation sites have been identified in histones H2A, H3, and H4. K12-biotinylated histone H4 is enriched in heterochromatin, repeat regions, and plays a role in gene repression. About 30% of the histone H4 molecules are biotinylated at K12 in histone H4 in human fibroblast telomeres. The abundance of biotinylated histones at distinct genomic loci depends on biotin availability. Decreased histone biotinylation decreases life span and stress resistance in Drosophila. Low enrichment of biotinylated histones at transposable elements impairs repression of these elements.

  6. Chromatin remodeling and cancer, Part I: Covalent histone modifications.

    Science.gov (United States)

    Wang, Gang G; Allis, C David; Chi, Ping

    2007-09-01

    Dynamic chromatin remodeling underlies many, if not all, DNA-templated biological processes, including gene transcription; DNA replication and repair; chromosome condensation; and segregation and apoptosis. Disruption of these processes has been linked to the development and progression of cancer. The mechanisms of dynamic chromatin remodeling include the use of covalent histone modifications, histone variants, ATP-dependent complexes and DNA methylation. Together, these mechanisms impart variation into the chromatin fiber, and this variation gives rise to an 'epigenetic landscape' that extends the biological output of DNA alone. Here, we review recent advances in chromatin remodeling, and pay particular attention to mechanisms that appear to be linked to human cancer. Where possible, we discuss the implications of these advances for disease-management strategies.

  7. Histone code or not? Combinatorial pattern analyses of histone modifications

    Science.gov (United States)

    Zang, Chongzhi; Peng, Weiqun; Wang, Zhibin; Schones, Dustin E.; Barski, Artem; Cuddapah, Suresh; Cui, Kairong; Roh, Tae-Young; Zhao, Keji; Rosenfeld, Jeffrey; Zhang, Michael

    2008-03-01

    Eukaryotic genomes are organized into chromatin, the structure of which plays critical role in the program of gene expression. Chromatin structure and function is regulated by a myriad of posttranslational modifications on histone tails of the nucleosomes, the fundamental unit of chromatin. It remains unclear how different modifications interact. Based on high- resolution genomic maps of close to 40 histone methylations and acetylations in human T-cells obtained experimentally by ChIP- Seq technique, we investigated the combinatorial patterns of histone modifications at gene promoter regions. We found that a very limited number of patterns dominate. Modifications within a pattern are strongly correlated and each pattern is associated with a distinct gene expression distribution. Our results suggest that it is the patterns rather than the individual modifications that affect the downstream readout.

  8. Tandem affinity purification of histones, coupled to mass spectrometry, identifies associated proteins and new sites of post-translational modification in Saccharomyces cerevisiae.

    Science.gov (United States)

    Valero, M Luz; Sendra, Ramon; Pamblanco, Mercè

    2016-03-16

    Histones and their post-translational modifications contribute to regulating fundamental biological processes in all eukaryotic cells. We have applied a conventional tandem affinity purification strategy to histones H3 and H4 of the yeast Saccharomyces cerevisiae. Mass spectrometry analysis of the co-purified proteins revealed multiple associated proteins, including core histones, which indicates that tagged histones may be incorporated to the nucleosome particle. Among the many other co-isolated proteins there are histone chaperones, elements of chromatin remodeling, of nucleosome assembly/disassembly, and of histone modification complexes. The histone chaperone Rtt106p, two members of chromatin assembly FACT complex and Psh1p, an ubiquitin ligase, were the most abundant proteins obtained with both H3-TAP and H4-TAP, regardless of the cell extraction medium stringency. Our mass spectrometry analyses have also revealed numerous novel post-translational modifications, including 30 new chemical modifications in histones, mainly by ubiquitination. We have discovered not only new sites of ubiquitination but that, besides lysine, also serine and threonine residues are targets of ubiquitination on yeast histones. Our results show the standard tandem affinity purification procedure is suitable for application to yeast histones, in order to isolate and characterize histone-binding proteins and post-translational modifications, avoiding the bias caused by histone purification from a chromatin-enriched fraction.

  9. Analysis of Myc-induced histone modifications on target chromatin.

    Directory of Open Access Journals (Sweden)

    Francesca Martinato

    Full Text Available The c-myc proto-oncogene is induced by mitogens and is a central regulator of cell growth and differentiation. The c-myc product, Myc, is a transcription factor that binds a multitude of genomic sites, estimated to be over 10-15% of all promoter regions. Target promoters generally pre-exist in an active or poised chromatin state that is further modified by Myc, contributing to fine transcriptional regulation (activation or repression of the afferent gene. Among other mechanisms, Myc recruits histone acetyl-transferases to target chromatin and locally promotes hyper-acetylation of multiple lysines on histones H3 and H4, although the identity and combination of the modified lysines is unknown. Whether Myc dynamically regulates other histone modifications (or marks at its binding sites also remains to be addressed. Here, we used quantitative chromatin immunoprecipitation (qChIP to profile a total of 24 lysine-acetylation and -methylation marks modulated by Myc at target promoters in a human B-cell line with a regulatable c-myc transgene. Myc binding promoted acetylation of multiple lysines, primarily of H3K9, H3K14, H3K18, H4K5 and H4K12, but significantly also of H4K8, H4K91 and H2AK5. Dimethylation of H3K79 was also selectively induced at target promoters. A majority of target promoters showed co-induction of multiple marks - in various combinations - correlating with recruitment of the two HATs tested (Tip60 and HBO1, incorporation of the histone variant H2A.Z and transcriptional activation. Based on this and previous findings, we surmise that Myc recruits the Tip60/p400 complex to achieve a coordinated histone acetylation/exchange reaction at activated promoters. Our data are also consistent with the additive and redundant role of multiple acetylation events in transcriptional activation.

  10. Centromere domain organization and histone modifications

    Directory of Open Access Journals (Sweden)

    P. Bjerling

    2002-05-01

    Full Text Available Centromere function requires the proper coordination of several subfunctions, such as kinetochore assembly, sister chromatid cohesion, binding of kinetochore microtubules, orientation of sister kinetochores to opposite spindle poles, and their movement towards the spindle poles. Centromere structure appears to be organized in different, separable domains in order to accomplish these functions. Despite the conserved nature of centromere functions, the molecular genetic definition of the DNA sequences that form a centromere in the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, in the fruit fly Drosophila melanogaster, and in humans has revealed little conservation at the level of centromere DNA sequences. Also at the protein level few centromere proteins are conserved in all of these four organisms and many are unique to the different organisms. The recent analysis of the centromere structure in the yeast S. pombe by electron microscopy and detailed immunofluorescence microscopy of Drosophila centromeres have brought to light striking similarities at the overall structural level between these centromeres and the human centromere. The structural organization of the centromere is generally multilayered with a heterochromatin domain and a central core/inner plate region, which harbors the outer plate structures of the kinetochore. It is becoming increasingly clear that the key factors for assembly and function of the centromere structure are the specialized histones and modified histones which are present in the centromeric heterochromatin and in the chromatin of the central core. Thus, despite the differences in the DNA sequences and the proteins that define a centromere, there is an overall structural similarity between centromeres in evolutionarily diverse eukaryotes.

  11. Genetic variants of methyl metabolizing enzymes and epigenetic regulators: Associations with promoter CpG island hypermethylation in colorectal cancer

    NARCIS (Netherlands)

    Vogel, S. de; Wouters, K.A.D.; Gottschalk, R.W.H.; Schooten, F.J. van; Goeij, A.F.P.M. de; Bruïne, A.P. de; Goldbohm, R.A.; Brandt, P.A. van den; Weijenberg, M.P.; Engeland, M. van

    2009-01-01

    Aberrant DNA methylation affects carcinogenesis of colorectal cancer. Folate metabolizing enzymes may influence the bioavailability of methyl groups, whereas DNA and histone methyltransferases are involved in epigenetic regulation of gene expression. We studied associations of genetic variants of fo

  12. Histone acetylation: from code to web and router via intrinsically disordered regions.

    Science.gov (United States)

    Horikoshi, Masami

    2013-01-01

    Structural changes of chromatin, which consists of nucleosomes and nucleosome-associated factors, lead to functional changes that are important determinants of eukaryotic gene regulation. These structural changes are regulated by modifications of histones and DNA, both of which are components of nucleosomes, as well as by replacement of histone variants and the actions of noncoding RNAs. In studies of chromatin modifications, a great deal of attention has been paid to histone acetylation. Progress in understanding this subject has been extensive, including i) elucidation of the relationship of histone acetylation and gene activity; ii) the first isolation of a histonemodifying enzyme; iii) the first identification of a factor that recognizes a modified site; iv) elucidation of the mechanism by which histone modification leads to structural changes in nucleosomes; and v) elucidation of the mechanism of border formation between euchromatin and heterochromatin. Histone acetylation is considered to be fundamental in several fields, including studies of a) the role of chromatin and epigenetics in higher-order biochemical systems such as transcription, DNA replication, and repair; b) biological phenomena such as cell proliferation and differentiation; and c) cancer and aging, potentially leading to clinical applications. In this review, I will discuss the histone code hypothesis, at one time believed to represent a unified theory regarding the functions of histone modification. In addition, I will describe the "modification web theory, " by which the problems in the histone code hypothesis can be overcome, as well as the "signal router theory, " which explains the mechanisms of formation, development, and evolution of the modification web from a structural viewpoint. Lastly, I will illustrate how these novel theories partially explain the robustness of biological systems against various perturbations, and elucidate the strategy that a cell employs to avoid fatal

  13. Structural and histone binding ability characterizations of human PWWP domains.

    Directory of Open Access Journals (Sweden)

    Hong Wu

    Full Text Available BACKGROUND: The PWWP domain was first identified as a structural motif of 100-130 amino acids in the WHSC1 protein and predicted to be a protein-protein interaction domain. It belongs to the Tudor domain 'Royal Family', which consists of Tudor, chromodomain, MBT and PWWP domains. While Tudor, chromodomain and MBT domains have long been known to bind methylated histones, PWWP was shown to exhibit histone binding ability only until recently. METHODOLOGY/PRINCIPAL FINDINGS: The PWWP domain has been shown to be a DNA binding domain, but sequence analysis and previous structural studies show that the PWWP domain exhibits significant similarity to other 'Royal Family' members, implying that the PWWP domain has the potential to bind histones. In order to further explore the function of the PWWP domain, we used the protein family approach to determine the crystal structures of the PWWP domains from seven different human proteins. Our fluorescence polarization binding studies show that PWWP domains have weak histone binding ability, which is also confirmed by our NMR titration experiments. Furthermore, we determined the crystal structures of the BRPF1 PWWP domain in complex with H3K36me3, and HDGF2 PWWP domain in complex with H3K79me3 and H4K20me3. CONCLUSIONS: PWWP proteins constitute a new family of methyl lysine histone binders. The PWWP domain consists of three motifs: a canonical β-barrel core, an insertion motif between the second and third β-strands and a C-terminal α-helix bundle. Both the canonical β-barrel core and the insertion motif are directly involved in histone binding. The PWWP domain has been previously shown to be a DNA binding domain. Therefore, the PWWP domain exhibits dual functions: binding both DNA and methyllysine histones. ENHANCED VERSION: This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web

  14. The histone acetyltransferase p300 inhibitor C646 reduces pro-inflammatory gene expression and inhibits histone deacetylases

    NARCIS (Netherlands)

    van den Bosch, Thea; Boichenko, Alexander; Leus, Niek G J; Ourailidou, Maria Eleni; Wapenaar, Hannah; Rotili, Dante; Mai, Antonello; Imhof, Axel; Bischoff, Rainer; Haisma, Hidde J; Dekker, Frank J

    2016-01-01

    Lysine acetylations are reversible posttranslational modifications of histone and non-histone proteins that play important regulatory roles in signal transduction cascades and gene expression. Lysine acetylations are regulated by histone acetyltransferases as writers and histone deacetylases as eras

  15. The C terminus of the histone chaperone Asf1 cross-links to histone H3 in yeast and promotes interaction with histones H3 and H4.

    Science.gov (United States)

    Dennehey, Briana K; Noone, Seth; Liu, Wallace H; Smith, Luke; Churchill, Mair E A; Tyler, Jessica K

    2013-02-01

    The central histone H3/H4 chaperone Asf1 comprises a highly conserved globular core and a divergent C-terminal tail. While the function and structure of the Asf1 core are well known, the function of the tail is less well understood. Here, we have explored the role of the yeast (yAsf1) and human (hAsf1a and hAsf1b) Asf1 tails in Saccharomyces cerevisiae. We show, using a photoreactive, unnatural amino acid, that Asf1 tail residue 210 cross-links to histone H3 in vivo and, further, that loss of C-terminal tail residues 211 to 279 weakens yAsf1-histone binding affinity in vitro nearly 200-fold. Via several yAsf1 C-terminal truncations and yeast-human chimeric proteins, we found that truncations at residue 210 increase transcriptional silencing and that the hAsf1a tail partially substitutes for full-length yAsf1 with respect to silencing but that full-length hAsf1b is a better overall substitute for full-length yAsf1. In addition, we show that the C-terminal tail of Asf1 is phosphorylated at T270 in yeast. Loss of this phosphorylation site does not prevent coimmunoprecipitation of yAsf1 and Rad53 from yeast extracts, whereas amino acid residue substitutions at the Asf1-histone H3/H4 interface do. Finally, we show that residue substitutions in yAsf1 near the CAF-1/HIRA interface also influence yAsf1's function in silencing.

  16. Histone chaperone networks shaping chromatin function

    DEFF Research Database (Denmark)

    Hammond, Colin; Strømme, Caroline Bianchi; Huang, Hongda

    2017-01-01

    and fate, which affects all chromosomal processes, including gene expression, chromosome segregation and genome replication and repair. Here, we review the distinct structural and functional properties of the expanding network of histone chaperones. We emphasize how chaperones cooperate in the histone...... chaperone network and via co-chaperone complexes to match histone supply with demand, thereby promoting proper nucleosome assembly and maintaining epigenetic information by recycling modified histones evicted from chromatin....

  17. A unique binding mode enables MCM2 to chaperone histones H3-H4 at replication forks.

    Science.gov (United States)

    Huang, Hongda; Strømme, Caroline B; Saredi, Giulia; Hödl, Martina; Strandsby, Anne; González-Aguilera, Cristina; Chen, Shoudeng; Groth, Anja; Patel, Dinshaw J

    2015-08-01

    During DNA replication, chromatin is reassembled by recycling of modified old histones and deposition of new ones. How histone dynamics integrates with DNA replication to maintain genome and epigenome information remains unclear. Here, we reveal how human MCM2, part of the replicative helicase, chaperones histones H3-H4. Our first structure shows an H3-H4 tetramer bound by two MCM2 histone-binding domains (HBDs), which hijack interaction sites used by nucleosomal DNA. Our second structure reveals MCM2 and ASF1 cochaperoning an H3-H4 dimer. Mutational analyses show that the MCM2 HBD is required for MCM2-7 histone-chaperone function and normal cell proliferation. Further, we show that MCM2 can chaperone both new and old canonical histones H3-H4 as well as H3.3 and CENPA variants. The unique histone-binding mode of MCM2 thus endows the replicative helicase with ideal properties for recycling histones genome wide during DNA replication.

  18. A unique binding mode enables MCM2 to chaperone histones H3–H4 at replication forks

    Science.gov (United States)

    Huang, Hongda; Strømme, Caroline B; Saredi, Giulia; Hödl, Martina; Strandsby, Anne; González-Aguilera, Cristina; Chen, Shoudeng; Groth, Anja; Patel, Dinshaw J

    2015-01-01

    During DNA replication, chromatin is reassembled by recycling of modified old histones and deposition of new ones. How histone dynamics integrates with DNA replication to maintain genome and epigenome information remains unclear. Here, we reveal how human MCM2, part of the replicative helicase, chaperones histones H3–H4. Our first structure shows an H3–H4 tetramer bound by two MCM2 histone-binding domains (HBDs), which hijack interaction sites used by nucleosomal DNA. Our second structure reveals MCM2 and ASF1 cochaperoning an H3–H4 dimer. Mutational analyses show that the MCM2 HBD is required for MCM2–7 histone-chaperone function and normal cell proliferation. Further, we show that MCM2 can chaperone both new and old canonical histones H3–H4 as well as H3.3 and CENPA variants. The unique histone-binding mode of MCM2 thus endows the replicative helicase with ideal properties for recycling histones genome wide during DNA replication. PMID:26167883

  19. Coupling between Histone Conformations and DNA Geometry in Nucleosomes on a Microsecond Timescale: Atomistic Insights into Nucleosome Functions.

    Science.gov (United States)

    Shaytan, Alexey K; Armeev, Grigoriy A; Goncearenco, Alexander; Zhurkin, Victor B; Landsman, David; Panchenko, Anna R

    2016-01-16

    An octamer of histone proteins wraps about 200bp of DNA into two superhelical turns to form nucleosomes found in chromatin. Although the static structure of the nucleosomal core particle has been solved, details of the dynamic interactions between histones and DNA remain elusive. We performed extensively long unconstrained, all-atom microsecond molecular dynamics simulations of nucleosomes including linker DNA segments and full-length histones in explicit solvent. For the first time, we were able to identify and characterize the rearrangements in nucleosomes on a microsecond timescale including the coupling between the conformation of the histone tails and the DNA geometry. We found that certain histone tail conformations promoted DNA bulging near its entry/exit sites, resulting in the formation of twist defects within the DNA. This led to a reorganization of histone-DNA interactions, suggestive of the formation of initial nucleosome sliding intermediates. We characterized the dynamics of the histone tails upon their condensation on the core and linker DNA and showed that tails may adopt conformationally constrained positions due to the insertion of "anchoring" lysines and arginines into the DNA minor grooves. Potentially, these phenomena affect the accessibility of post-translationally modified histone residues that serve as important sites for epigenetic marks (e.g., at H3K9, H3K27, H4K16), suggesting that interactions of the histone tails with the core and linker DNA modulate the processes of histone tail modifications and binding of the effector proteins. We discuss the implications of the observed results on the nucleosome function and compare our results to different experimental studies.

  20. Histone H1 of Trypanosoma cruzi is concentrated in the nucleolus region and disperses upon phosphorylation during progression to mitosis.

    Science.gov (United States)

    Gutiyama, Luciana M; da Cunha, Julia P Chagas; Schenkman, Sergio

    2008-04-01

    Phosphorylation of histone H1 is intimately related to the cell cycle progression in higher eukaryotes, reaching maximum levels during mitosis. We have previously shown that in the flagellated protozoan Trypanosoma cruzi, which does not condense chromatin during mitosis, histone H1 is phosphorylated at a single cyclin-dependent kinase site. By using an antibody that recognizes specifically the phosphorylated T. cruzi histone H1 site, we have now confirmed that T. cruzi histone H1 is also phosphorylated in a cell cycle-dependent manner. Differently from core histones, the bulk of nonphosphorylated histone H1 in G(1) and S phases of the cell cycle is concentrated in the central regions of the nucleus, which contains the nucleolus and less densely packed chromatin. When cells pass G(2), histone H1 becomes phosphorylated and starts to diffuse. At the onset of mitosis, histone H1 phosphorylation is maximal and found in the entire nuclear space. As permeabilized parasites preferentially lose phosphorylated histone H1, we conclude that this modification promotes its release from less condensed and nucleolar chromatin after G(2).

  1. CSR-1 RNAi pathway positively regulates histone expression in C. elegans.

    Science.gov (United States)

    Avgousti, Daphne C; Palani, Santhosh; Sherman, Yekaterina; Grishok, Alla

    2012-10-03

    Endogenous small interfering RNAs (endo-siRNAs) have been discovered in many organisms, including mammals. In C. elegans, depletion of germline-enriched endo-siRNAs found in complex with the CSR-1 Argonaute protein causes sterility and defects in chromosome segregation in early embryos. We discovered that knockdown of either csr-1, the RNA-dependent RNA polymerase (RdRP) ego-1, or the dicer-related helicase drh-3, leads to defects in histone mRNA processing, resulting in severe depletion of core histone proteins. The maturation of replication-dependent histone mRNAs, unlike that of other mRNAs, requires processing of their 3'UTRs through an endonucleolytic cleavage guided by the U7 snRNA, which is lacking in C. elegans. We found that CSR-1-bound antisense endo-siRNAs match histone mRNAs and mRNA precursors. Consistently, we demonstrate that CSR-1 directly binds to histone mRNA in an ego-1-dependent manner using biotinylated 2'-O-methyl RNA oligonucleotides. Moreover, we demonstrate that increasing the dosage of histone genes rescues the lethality associated with depletion of CSR-1 and EGO-1. These results support a positive and direct effect of RNAi on histone gene expression.

  2. Nucleosome alterations caused by mutations at modifiable histone residues in Saccharomyces cerevisiae.

    Science.gov (United States)

    Liu, Hongde; Wang, Pingyan; Liu, Lingjie; Min, Zhu; Luo, Kun; Wan, Yakun

    2015-10-26

    Nucleosome organization exhibits dynamic properties depending on the cell state and environment. Histone proteins, fundamental components of nucleosomes, are subject to chemical modifications on particular residues. We examined the effect of substituting modifiable residues of four core histones with the non-modifiable residue alanine on nucleosome dynamics. We mapped the genome-wide nucleosomes in 22 histone mutants of Saccharomyces cerevisiae and compared the nucleosome alterations relative to the wild-type strain. Our results indicated that different types of histone mutation resulted in different phenotypes and a distinct reorganization of nucleosomes. Nucleosome occupancy was altered at telomeres, but not at centromeres. The first nucleosomes upstream (-1) and downstream (+1) of the transcription start site (TSS) were more dynamic than other nucleosomes. Mutations in histones affected the nucleosome array downstream of the TSS. Highly expressed genes, such as ribosome genes and genes involved in glycolysis, showed increased nucleosome occupancy in many types of histone mutant. In particular, the H3K56A mutant exhibited a high percentage of dynamic genomic regions, decreased nucleosome occupancy at telomeres, increased occupancy at the +1 and -1 nucleosomes, and a slow growth phenotype under stress conditions. Our findings provide insight into the influence of histone mutations on nucleosome dynamics.

  3. Biochemical profiling of histone binding selectivity of the yeast bromodomain family.

    Directory of Open Access Journals (Sweden)

    Qiang Zhang

    Full Text Available BACKGROUND: It has been shown that molecular interactions between site-specific chemical modifications such as acetylation and methylation on DNA-packing histones and conserved structural modules present in transcriptional proteins are closely associated with chromatin structural changes and gene activation. Unlike methyl-lysine that can interact with different protein modules including chromodomains, Tudor and MBT domains, as well as PHD fingers, acetyl-lysine (Kac is known thus far to be recognized only by bromodomains. While histone lysine acetylation plays a crucial role in regulation of chromatin-mediated gene transcription, a high degree of sequence variation of the acetyl-lysine binding site in the bromodomains has limited our understanding of histone binding selectivity of the bromodomain family. Here, we report a systematic family-wide analysis of 14 yeast bromodomains binding to 32 lysine-acetylated peptides derived from known major acetylation sites in four core histones that are conserved in eukaryotes. METHODOLOGY: The histone binding selectivity of purified recombinant yeast bromodomains was assessed by using the native core histones in an overlay assay, as well as N-terminally biotinylated lysine-acetylated histone peptides spotted on streptavidin-coated nitrocellulose membrane in a dot blot assay. NMR binding analysis further validated the interactions between histones and selected bromodomain. Structural models of all yeast bromodomains were built using comparative modeling to provide insights into the molecular basis of their histone binding selectivity. CONCLUSIONS: Our study reveals that while not all members of the bromodomain family are privileged to interact with acetylated-lysine, identifiable sequence features from those that bind histone emerge. These include an asparagine residue at the C-terminus of the third helix in the 4-helix bundle, negatively charged residues around the ZA loop, and preponderance of aromatic

  4. Strategies for crystallizing a chromatin protein in complex with the nucleosome core particle.

    Science.gov (United States)

    Makde, Ravindra D; Tan, Song

    2013-11-15

    The molecular details of how chromatin factors and enzymes interact with the nucleosome are critical to understanding fundamental genetic processes including cell division and gene regulation. A structural understanding of such processes has been hindered by the difficulty in producing diffraction-quality crystals of chromatin proteins in complex with the nucleosome. We describe here the steps used to grow crystals of the 300-kDa RCC1 chromatin factor/nucleosome core particle complex that diffract to 2.9-Å resolution. These steps include both pre- and postcrystallization strategies potentially useful to other complexes. We screened multiple variant RCC1/nucleosome core particle complexes assembled using different RCC1 homologs and deletion variants, and nucleosomes containing nucleosomal DNA with different sequences and lengths, as well as histone deletion variants. We found that using RCC1 from different species produced different crystal forms of the RCC1/nucleosome complex consistent with key crystal packing interactions mediated by RCC1. Optimization of postcrystallization soaks to dehydrate the crystals dramatically improved the diffraction quality of the RCC1/nucleosome crystal from 5.0- to 2.9-Å resolution. Copyright © 2013 Elsevier Inc. All rights reserved.

  5. Daunomycin, an antitumor DNA intercalator, influences histone-DNA interactions.

    Science.gov (United States)

    Wójcik, Krzysztof; Zarębski, Mirosław; Cossarizza, Andrea; Dobrucki, Jurek W

    2013-09-01

    Although daunomycin and adriamycin are considered effective antitumor drugs and have been used in the clinic for over 40 years, their mechanism of action is still a matter of debate. We investigated the influence of daunomycin on interaction between linker or core histones and DNA in live HeLa cells in vitro, using image and flow cytometry. Exposure to daunomycin at clinically relevant concentrations (25-250 nM) caused dissociation of wild-type H1.1 as well as 4 H1 point mutants from DNA, followed by their accumulation in nucleoli and aggregation of chromatin. A detectable dissociation of H2B core histones occurred only at much higher concentrations of the drug (500 nM). Replication of DNA and synthesis of RNA were not halted by daunomycin (up to 2500 nM); however the characteristic subnuclear distribution of sites of transcription and replication was lost. Dissociation of the H1.1 linker histones and subsequent loss of higher order chromatin structures may constitute an important component of the mechanism of cytotoxicity of daunomycin.

  6. Electrostatic effect of H1-histone protein binding on nucleosome repeat length

    Science.gov (United States)

    Cherstvy, Andrey G.; Teif, Vladimir B.

    2014-08-01

    Within a simple biophysical model we describe the effect of electrostatic binding of H1 histone proteins on the nucleosome repeat length in chromatin. The length of wrapped DNA optimizes its binding energy to the histone core and the elastic energy penalty of DNA wrapping. The magnitude of the effect predicted from our model is in agreement with the systematic experimental data on the linear variation of nucleosome repeat lengths with H1/nucleosome ratio (Woodcock C L et al 2006 Chromos. Res. 14 17-25). We compare our model to the data for different cell types and organisms, with a widely varying ratio of bound H1 histones per nucleosome. We underline the importance of this non-specific histone-DNA charge-balance mechanism in regulating the positioning of nucleosomes and the degree of compaction of chromatin fibers in eukaryotic cells.

  7. Importance of electrostatic interactions in the association of intrinsically disordered histone chaperone Chz1 and histone H2A.Z-H2B.

    Directory of Open Access Journals (Sweden)

    Xiakun Chu

    Full Text Available Histone chaperones facilitate assembly and disassembly of nucleosomes. Understanding the process of how histone chaperones associate and dissociate from the histones can help clarify their roles in chromosome metabolism. Some histone chaperones are intrinsically disordered proteins (IDPs. Recent studies of IDPs revealed that the recognition of the biomolecules is realized by the flexibility and dynamics, challenging the century-old structure-function paradigm. Here we investigate the binding between intrinsically disordered chaperone Chz1 and histone variant H2A.Z-H2B by developing a structure-based coarse-grained model, in which Debye-Hückel model is implemented for describing electrostatic interactions due to highly charged characteristic of Chz1 and H2A.Z-H2B. We find that major structural changes of Chz1 only occur after the rate-limiting electrostatic dominant transition state and Chz1 undergoes folding coupled binding through two parallel pathways. Interestingly, although the electrostatic interactions stabilize bound complex and facilitate the recognition at first stage, the rate for formation of the complex is not always accelerated due to slow escape of conformations with non-native electrostatic interactions at low salt concentrations. Our studies provide an ionic-strength-controlled binding/folding mechanism, leading to a cooperative mechanism of "local collapse or trapping" and "fly-casting" together and a new understanding of the roles of electrostatic interactions in IDPs' binding.

  8. Identification and Interrogation of Combinatorial Histone Modifications

    Directory of Open Access Journals (Sweden)

    Kelly R Karch

    2013-12-01

    Full Text Available Histone proteins are dynamically modified to mediate a variety of cellular processes including gene transcription, DNA damage repair, and apoptosis. Regulation of these processes occurs through the recruitment of non-histone proteins to chromatin by specific combinations of histone post-translational modifications (PTMs. Mass spectrometry has emerged as an essential tool to discover and quantify histone PTMs both within and between samples in an unbiased manner. Developments in mass spectrometry that allow for characterization of large histone peptides or intact protein has made it possible to determine which modifications occur simultaneously on a single histone polypeptide. A variety of techniques from biochemistry, biophysics, and chemical biology have been employed to determine the biological relevance of discovered combinatorial codes. This review first describes advancements in the field of mass spectrometry that have facilitated histone PTM analysis and then covers notable approaches to probe the biological relevance of these modifications in their nucleosomal context.

  9. SWI/SNF chromatin remodeling and linker histones in plants.

    Science.gov (United States)

    Jerzmanowski, Andrzej

    2007-01-01

    In yeast and mammals, ATP-dependent chromatin remodeling complexes belonging to the SWI/SNF family play critical roles in the regulation of transcription, cell proliferation, differentiation and development. Homologs of conserved subunits of SWI/SNF-type complexes, including several putative ATPases and other core subunits, have been identified in plants. Here I summarize recent insights in structural organization and functional diversification of putative plant SWI/SNF-type chromatin remodeling complexes and discuss in a broader evolutionary perspective the similarities and differences between plant and yeast/animal SWI/SNF remodeling. I also summarize the current view of localization in nucleosome and dynamic behaviour in chromatin of linker (H1) histones and discuss significance of recent findings indicating that in both plants and mammals histone H1 is involved in determining patterns of DNA methylation at selected loci.

  10. Histone gene expression and histone mRNA 3' end structure in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Pettitt Jonathan

    2007-06-01

    Full Text Available Abstract Background Histone protein synthesis is essential for cell proliferation and required for the packaging of DNA into chromatin. In animals, histone proteins are provided by the expression of multicopy replication-dependent histone genes. Histone mRNAs that are processed by a histone-specific mechanism to end after a highly conserved RNA hairpin element, and lack a poly(A tail. In vertebrates and Drosophila, their expression is dependent on HBP/SLBP that binds to the RNA hairpin element. We showed previously that these cis and trans acting regulators of histone gene expression are conserved in C. elegans. Here we report the results of an investigation of the histone mRNA 3' end structure and of histone gene expression during C. elegans development. Results Sequence analysis of replication-dependent histone genes revealed the presence of several highly conserved sequence elements in the 3' untranslated region of histone pre-mRNAs, including an RNA hairpin element and a polyadenylation signal. To determine whether in C. elegans histone mRNA 3' end formation occurs at this polyadenylation signal and results in polyadenylated histone mRNA, we investigated the mRNA 3' end structure of histone mRNA. Using poly(A selection, RNAse protection and sequencing of histone mRNA ends, we determined that a majority of C. elegans histone mRNAs lack a poly(A tail and end three to six nucleotides after the hairpin structure, after an A or a U, and have a 3' OH group. RNAi knock down of CDL-1, the C. elegans HBP/SLBP, does not significantly affect histone mRNA levels but severely depletes histone protein levels. Histone gene expression varies during development and is reduced in L3 animals compared to L1 animals and adults. In adults, histone gene expression is restricted to the germ line, where cell division occurs. Conclusion Our findings indicate that the expression of C. elegans histone genes is subject to control mechanisms similar to the ones in other

  11. Role of H1 linker histones in mammalian development and stem cell differentiation.

    Science.gov (United States)

    Pan, Chenyi; Fan, Yuhong

    2016-03-01

    H1 linker histones are key chromatin architectural proteins facilitating the formation of higher order chromatin structures. The H1 family constitutes the most heterogeneous group of histone proteins, with eleven non-allelic H1 variants in mammals. H1 variants differ in their biochemical properties and exhibit significant sequence divergence from one another, yet most of them are highly conserved during evolution from mouse to human. H1 variants are differentially regulated during development and their cellular compositions undergo dramatic changes in embryogenesis, gametogenesis, tissue maturation and cellular differentiation. As a group, H1 histones are essential for mouse development and proper stem cell differentiation. Here we summarize our current knowledge on the expression and functions of H1 variants in mammalian development and stem cell differentiation. Their diversity, sequence conservation, complex expression and distinct functions suggest that H1s mediate chromatin reprogramming and contribute to the large variations and complexity of chromatin structure and gene expression in the mammalian genome. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Histone H4 hyperacetylation and rapid turnover of its acetyl groups in transcriptionally inactive rooster testis spermatids.

    Science.gov (United States)

    Oliva, R; Mezquita, C

    1982-12-20

    In order to study the relationship between acetylation of histones, chromatin structure and gene activity, the distribution and turnover of acetyl groups among nucleosomal core histones and the extent of histone H4 acetylation were examined in rooster testis cell nuclei at different stages of spermatogenesis. Histone H4 was the predominant acetylated histone in mature testes. Hyperacetylation of H4 and rapid turnover of its acetyl groups are not univocally correlated with transcriptional activity since they were detected in both genetically active testicular cells and genetically inactive elongated spermatids. During the transition from nucleohistone to nucleoprotamine in elongated spermatids the chromatin undergoes dramatic structural changes with exposition of binding sites on DNA (1). Hyperacetylation of H4 and rapid turnover of its acetyl groups could be correlated with the particular conformation of chromatin in elongated spermatids and might represent a necessary condition for binding of chromosomal proteins to DNA.

  13. A PHD in histone language: on the role of histone methylation in plant responses to phosphate deficiency.

    Science.gov (United States)

    Chandrika, Nulu Naga Prafulla; Sundaravelpandian, Kalaipandian; Schmidt, Wolfgang

    2013-06-01

    Post-translational modifications of core histones are important for various DNA-templated processes such as transcription and repair. We recently reported that the ALFIN LIKE 6 (AL6) gene, identified in a forward genetic screen, is critical for phosphate deficiency-induced root hair formation and several other processes associated with the regulation of cellular phosphate homeostasis. AL6 contains a Plant Homeo Domain (PHD) finger that can bind to trimethylated lysine 4 of histone H3 (H3K4me3). Homozygous mutants defective in AL6 expression form very short root hairs under phosphate-deficient conditions, presumably caused by altered expression of putative primary and secondary down-stream targets of AL6. In this Addendum, we speculate about possible roles of AL6, H3K4 trimethylation and other chromatin modifications in the adaptation of plants to low phosphate availability.

  14. Genetic variants in epigenetic genes and breast cancer risk.

    Science.gov (United States)

    Cebrian, Arancha; Pharoah, Paul D; Ahmed, Shahana; Ropero, Santiago; Fraga, Mario F; Smith, Paula L; Conroy, Don; Luben, Robert; Perkins, Barbara; Easton, Douglas F; Dunning, Alison M; Esteller, Manel; Ponder, Bruce A J

    2006-08-01

    Epigenetic events, resulting changes in gene expression capacity, are important in tumour progression, and variation in genes involved in epigenetic mechanisms might therefore be important in cancer susceptibility. To evaluate this hypothesis, we examined common variants in 12 genes coding for DNA methyltransferases (DNMT), histone acetyltransferases, histone deacetyltransferases, histone methyltrasferases and methyl-CpG binding domain proteins, for association with breast cancer in a large case-control study (N cases = 4474 and N controls = 4580). We identified 63 single nucleotide polymorphisms (SNPs) that efficiently tag all the known common variants in these genes, and are also expected to tag any unknown SNP in each gene. We found some evidence for association for six SNPs: DNMT3b-c31721t [P (2 df) = 0.007], PRDM2-c99243 t [P (2 df) = 0.03] and t105413c [P-recessive = 0.05], EHMT1-g-9441a [P (2df) = 0.05] and g41451t (P-trend = 0.04), and EHMT2-S237S [P (2df) = 0.04]. The most significant result was for DNMT3b-c31721t (P-trend = 0.124 after adjusting for multiple testing). However, there were three other results with P variants in histone methyltransferases, and warrant the design of larger epidemiological and biochemical studies to establish the true meaning of these findings.

  15. Two arginine residues suppress the flexibility of nucleosomal DNA in the canonical nucleosome core.

    Science.gov (United States)

    Kono, Hidetoshi; Shirayama, Kazuyoshi; Arimura, Yasuhiro; Tachiwana, Hiroaki; Kurumizaka, Hitoshi

    2015-01-01

    The dynamics of nucleosomes containing either canonical H3 or its centromere-specific variant CENP-A were investigated using molecular dynamics simulations. The simulations showed that the histone cores were structurally stable during simulation periods of 100 ns and 50 ns, while DNA was highly flexible at the entry and exit regions and partially dissociated from the histone core. In particular, approximately 20-25 bp of DNA at the entry and exit regions of the CENP-A nucleosome exhibited larger fluctuations than DNA at the entry and exit regions of the H3 nucleosome. Our detailed analysis clarified that this difference in dynamics was attributable to a difference in two basic amino acids in the αN helix; two arginine (Arg) residues in H3 were substituted by lysine (Lys) residues at the corresponding sites in CENP-A. The difference in the ability to form hydrogen bonds with DNA of these two residues regulated the flexibility of nucleosomal DNA at the entry and exit regions. Our exonuclease III assay consistently revealed that replacement of these two Arg residues in the H3 nucleosome by Lys enhanced endonuclease susceptibility, suggesting that the DNA ends of the CENP-A nucleosome are more flexible than those of the H3 nucleosome. This difference in the dynamics between the two types of nucleosomes may be important for forming higher order structures in different phases.

  16. Single molecule DNA compaction by purified histones

    Institute of Scientific and Technical Information of China (English)

    RAN ShiYong; WANG XiaoLing; FU WenBo; WANG WeiChi; LI Ming

    2008-01-01

    The compaction of single DNA molecules by purified histones is studied using magnetic tweezers, The compaction rate increases rapidly when the histone concentration is increased from 0.002 to 0.2 mmol/L, and saturates when the concentration is beyond 0.2 mmol/L, The time course of compaction is exponential at low histone concentrations. It becomes sigmoidal at high concentrations. Cooperativity between the histones bound to DNA is proposed to be responsible for the transition. The histones are loaded onto DNA randomly at low concentrations. They tend to bind DNA cooperatively at high con-centrations because the structural torsions of DNA induced by the bound histones become overlapping so that the binding of one histone facilitates the binding of others. Under very large forces, the com-pacted histone-DNA complex can be disrupted in a discrete manner with a step size of ~60 nm. But the histones cannot be completely stripped off DNA, as is revealed by the lowered B-S transition plateau of the histone-bound DNA.

  17. Dissecting the Molecular Roles of Histone Chaperones in Histone Acetylation by Type B Histone Acetyltransferases (HAT-B).

    Science.gov (United States)

    Haigney, Allison; Ricketts, M Daniel; Marmorstein, Ronen

    2015-12-18

    The HAT-B enzyme complex is responsible for acetylating newly synthesized histone H4 on lysines K5 and K12. HAT-B is a multisubunit complex composed of the histone acetyltransferase 1 (Hat1) catalytic subunit and the Hat2 (rbap46) histone chaperone. Hat1 is predominantly localized in the nucleus as a member of a trimeric NuB4 complex containing Hat1, Hat2, and a histone H3-H4 specific histone chaperone called Hif1 (NASP). In addition to Hif1 and Hat2, Hat1 interacts with Asf1 (anti-silencing function 1), a histone chaperone that has been reported to be involved in both replication-dependent and -independent chromatin assembly. To elucidate the molecular roles of the Hif1 and Asf1 histone chaperones in HAT-B histone binding and acetyltransferase activity, we have characterized the stoichiometry and binding mode of Hif1 and Asf1 to HAT-B and the effect of this binding on the enzymatic activity of HAT-B. We find that Hif1 and Asf1 bind through different modes and independently to HAT-B, whereby Hif1 binds directly to Hat2, and Asf1 is only capable of interactions with HAT-B through contacts with histones H3-H4. We also demonstrate that HAT-B is significantly more active against an intact H3-H4 heterodimer over a histone H4 peptide, independent of either Hif1 or Asf1 binding. Mutational studies further demonstrate that HAT-B binding to the histone tail regions is not sufficient for this enhanced activity. Based on these data, we propose a model for HAT-B/histone chaperone assembly and acetylation of H3-H4 complexes.

  18. Robust methods for purification of histones from cultured mammalian cells with the preservation of their native modifications.

    Science.gov (United States)

    Rodriguez-Collazo, Pedro; Leuba, Sanford H; Zlatanova, Jordanka

    2009-06-01

    Post-translational modifications (PTMs) of histones play a role in modifying chromatin structure for DNA-templated processes in the eukaryotic nucleus, such as transcription, replication, recombination and repair; thus, histone PTMs are considered major players in the epigenetic control of these processes. Linking specific histone PTMs to gene expression is an arduous task requiring large amounts of highly purified and natively modified histones to be analyzed by various techniques. We have developed robust and complementary procedures, which use strong protein denaturing conditions and yield highly purified core and linker histones from unsynchronized proliferating, M-phase arrested and butyrate-treated cells, fully preserving their native PTMs without using enzyme inhibitors. Cell hypotonic swelling and lysis, nuclei isolation/washing and chromatin solubilization under mild conditions are bypassed to avoid compromising the integrity of histone native PTMs. As controls for our procedures, we tested the most widely used conventional methodologies and demonstrated that they indeed lead to drastic histone dephosphorylation. Additionally, we have developed methods for preserving acid-labile histone modifications by performing non-acid extractions to obtain highly purified H3 and H4. Importantly, isolation of histones H3, H4 and H2A/H2B is achieved without the use of HPLC. Functional supercoiling assays reveal that both hyper- and hypo-phosphorylated histones can be efficiently assembled into polynucleosomes. Notably, the preservation of fully phosphorylated mitotic histones and their assembly into polynucleosomes should open new avenues to investigate an important but overlooked question: the impact of mitotic phosphorylation in chromatin structure and function.

  19. SIRT1-metabolite binding histone macroH2A1.1 protects hepatocytes against lipid accumulation

    NARCIS (Netherlands)

    Pazienza, Valerio; Borghesan, Michela; Mazza, Tommaso; Sheedfar, Fareeba; Panebianco, Concetta; Williams, Roger; Mazzoccoli, Gianluigi; Andriulli, Angelo; Nakanishi, Tomoko; Vinciguerra, Manlio

    2014-01-01

    Non-alcoholic-fatty-liver-disease (NAFLD) encompasses conditions associated to fat deposition in the liver, which are generally deteriorated during the aging process. MacroH2A1, a variant of histone H2A, is a key transcriptional regulator involved in tumorigenic processes and cell senescence, and fe

  20. SIRT1-metabolite binding histone macroH2A1.1 protects hepatocytes against lipid accumulation.

    NARCIS (Netherlands)

    Pazienza, V.; Borghesan, M.; Mazza, T.; Sheedfar, F.; Panebianco, C.; Williams, R.; Mazzoccoli, G.; Andriulli, A.; Nakanishi, T.; Vinciguerra, M.

    2014-01-01

    Non-alcoholic-fatty-liver-disease (NAFLD) encompasses conditions associated to fat deposition in the liver, which are generally deteriorated during the aging process. MacroH2A1, a variant of histone H2A, is a key transcriptional regulator involved in tumorigenic processes and cell senescence, and

  1. SIRT1-metabolite binding histone macroH2A1.1 protects hepatocytes against lipid accumulation

    NARCIS (Netherlands)

    Pazienza, Valerio; Borghesan, Michela; Mazza, Tommaso; Sheedfar, Fareeba; Panebianco, Concetta; Williams, Roger; Mazzoccoli, Gianluigi; Andriulli, Angelo; Nakanishi, Tomoko; Vinciguerra, Manlio

    Non-alcoholic-fatty-liver-disease (NAFLD) encompasses conditions associated to fat deposition in the liver, which are generally deteriorated during the aging process. MacroH2A1, a variant of histone H2A, is a key transcriptional regulator involved in tumorigenic processes and cell senescence, and

  2. A unique binding mode enables MCM2 to chaperone histones H3-H4 at replication forks

    DEFF Research Database (Denmark)

    Huang, Hongda; Strømme, Caroline B; Saredi, Giulia

    2015-01-01

    , chaperones histones H3-H4. Our first structure shows an H3-H4 tetramer bound by two MCM2 histone-binding domains (HBDs), which hijack interaction sites used by nucleosomal DNA. Our second structure reveals MCM2 and ASF1 cochaperoning an H3-H4 dimer. Mutational analyses show that the MCM2 HBD is required......During DNA replication, chromatin is reassembled by recycling of modified old histones and deposition of new ones. How histone dynamics integrates with DNA replication to maintain genome and epigenome information remains unclear. Here, we reveal how human MCM2, part of the replicative helicase...... for MCM2-7 histone-chaperone function and normal cell proliferation. Further, we show that MCM2 can chaperone both new and old canonical histones H3-H4 as well as H3.3 and CENPA variants. The unique histone-binding mode of MCM2 thus endows the replicative helicase with ideal properties for recycling...

  3. Reduced Euchromatin histone methyltransferase 1 causes developmental delay, hypotonia, and cranial abnormalities associated with increased bone gene expression in Kleefstra syndrome mice

    NARCIS (Netherlands)

    Balemans, M.C.M.; Ansar, M.; Oudakker, A.R.; Caam, A.P.M. van; Bakker, B.; Vitters, E.L.; Kraan, P.M. van der; Bruijn, D.R.H. de; Janssen, S.M.; Kuipers, A.J.; Huibers, M.M.; Maliepaard, Eliza M.; Walboomers, X.F.; Benevento, M.; Nadif Kasri, N.; Kleefstra, T.; Zhou, H.; Zee, C.E.E.M. van der; Bokhoven, H. van

    2014-01-01

    Haploinsufficiency of Euchromatin histone methyltransferase 1 (EHMT1), a chromatin modifying enzyme, is the cause of Kleefstra syndrome (KS). KS is an intellectual disability (ID) syndrome, with general developmental delay, hypotonia, and craniofacial dysmorphisms as additional core features. Recent

  4. Human tNASP promotes in vitro nucleosome assembly with histone H3.3.

    Science.gov (United States)

    Kato, Daiki; Osakabe, Akihisa; Tachiwana, Hiroaki; Tanaka, Hiroki; Kurumizaka, Hitoshi

    2015-02-10

    Nuclear autoantigenic sperm proteins (NASPs) are members of the acidic histone chaperones, which promote nucleosome assembly. In humans, two splicing variants proposed for the somatic and testicular isoforms, sNASP and tNASP, respectively, have been found, and the shorter form, sNASP, reportedly promotes nucleosome assembly with the histone H3 isoforms, H3.1, H3.2, and H3.3. However, the biochemical properties of the longer form, tNASP, have not been reported. tNASP is considered to exist specifically in the testis. Our present results revealed that the tNASP protein is ubiquitously produced in various human tissues, in addition to testis. Unexpectedly, we found that the nucleosome assembly activity of purified tNASP was extremely low with the canonical histone H3.1 or H3.2, but was substantially detected with the replacement histone H3.3 variant. A mutational analysis revealed that the H3.3 Ile89 residue, corresponding to the H3.1 Val89 residue, is responsible for the tNASP-mediated nucleosome assembly with H3.3. A histone deposition assay showed that the H3.3-H4 complex is more efficiently deposited onto DNA by tNASP than the H3.1-H4 complex. These results provide evidence that tNASP is ubiquitously produced in various types of human tissues and promotes in vitro nucleosome assembly with H3 variant specificity.

  5. Methylglyoxal mediated conformational changes in histone H2A-generation of carboxyethylated advanced glycation end products.

    Science.gov (United States)

    Mir, Abdul Rouf; uddin, Moin; Alam, Khursheed; Ali, Asif

    2014-08-01

    Methylglyoxal, an oxo-aldehyde has been implicated as a potential precursor in non enzymatic glycation reactions. Its role in the modification of extra cellular proteins has been extensively reported, but little is known about its modification of nuclear proteins, like histones. Here, we report the methylglyoxal induced modification of histone H2A which forms an essential part of intact core nucleosome. In this study commercially available histone H2A was subjected to in vitro non-enzymatic glycation by methylglyoxal. The structural alterations in the histone were characterised by various biophysical and biochemical techniques. The modified histone showed hyperchromicity at 276nm, loss in intrinsic tyrosine fluorescence intensity at 305nm along with a red shift, cross linking and dimer formation in SDS PAGE, induction of α-helix in CD spectroscopy, reduced hydrophobicity in ANS binding studies, accumulation of AGE products, increased carbonyl content, and appearance of a novel peak showing carboxyethylation complemented by a shift in amide I and amide II bands in ATR-FTIR spectroscopy. The modified histone exhibited increased melting temperatures (Tm) and enhanced heat capacities (Cp) in differential scanning calorimetric analysis. The results suggest that methylglyoxal significantly altered the structure of the nuclear histone H2A by non enzymatic glycation reaction. The conformational changes in histone H2A may influence the chromatin integrity which may have implications in various pathological conditions.

  6. Histone Acetylation in Fungal Pathogens of Plants

    Directory of Open Access Journals (Sweden)

    Junhyun Jeon

    2014-03-01

    Full Text Available Acetylation of histone lysine residues occurs in different organisms ranging from yeast to plants and mammals for the regulation of diverse cellular processes. With the identification of enzymes that create or reverse this modification, our understanding on histone acetylation has expanded at an amazing pace during the last two decades. In fungal pathogens of plants, however, the importance of such modification has only just begun to be appreciated in the recent years and there is a dearth of information on how histone acetylation is implicated in fungal pathogenesis. This review covers the current status of research related to histone acetylation in plant pathogenic fungi and considers relevant findings in the interaction between fungal pathogens and host plants. We first describe the families of histone acetyltransferases and deacetylases. Then we provide the cases where histone acetylation was investigated in the context of fungal pathogenesis. Finally, future directions and perspectives in epigenetics of fungal pathogenesis are discussed.

  7. Basic nuclear processes affected by histone acetyltransferases and histone deacetylase inhibitors

    NARCIS (Netherlands)

    Legartová, Soňa; Stixová, Lenka; Strnad, Hynek; Kozubek, Stanislav; Martinet, Nadine; Dekker, Frank J; Franek, Michal; Bártová, Eva

    2013-01-01

    AIM: The optimal balance between histone acetylation and deacetylation is important for proper gene function. Therefore, we addressed how inhibitors of histone-modifying enzymes can modulate nuclear events, including replication, transcription, splicing and DNA repair. MATERIALS & METHODS: Changes i

  8. Structural insight into how the human helicase subunit MCM2 may act as a histone chaperone together with ASF1 at the replication fork

    Science.gov (United States)

    Richet, Nicolas; Liu, Danni; Legrand, Pierre; Velours, Christophe; Corpet, Armelle; Gaubert, Albane; Bakail, May; Moal-Raisin, Gwenaelle; Guerois, Raphael; Compper, Christel; Besle, Arthur; Guichard, Berengère; Almouzni, Genevieve; Ochsenbein, Françoise

    2015-01-01

    MCM2 is a subunit of the replicative helicase machinery shown to interact with histones H3 and H4 during the replication process through its N-terminal domain. During replication, this interaction has been proposed to assist disassembly and assembly of nucleosomes on DNA. However, how this interaction participates in crosstalk with histone chaperones at the replication fork remains to be elucidated. Here, we solved the crystal structure of the ternary complex between the histone-binding domain of Mcm2 and the histones H3-H4 at 2.9 Å resolution. Histones H3 and H4 assemble as a tetramer in the crystal structure, but MCM2 interacts only with a single molecule of H3-H4. The latter interaction exploits binding surfaces that contact either DNA or H2B when H3-H4 dimers are incorporated in the nucleosome core particle. Upon binding of the ternary complex with the histone chaperone ASF1, the histone tetramer dissociates and both MCM2 and ASF1 interact simultaneously with the histones forming a 1:1:1:1 heteromeric complex. Thermodynamic analysis of the quaternary complex together with structural modeling support that ASF1 and MCM2 could form a chaperoning module for histones H3 and H4 protecting them from promiscuous interactions. This suggests an additional function for MCM2 outside its helicase function as a proper histone chaperone connected to the replication pathway. PMID:25618846

  9. Pathway analysis of whole exome sequence data provides further support for the involvement of histone modification in the aetiology of schizophrenia.

    Science.gov (United States)

    Curtis, David

    2016-10-01

    Weighted burden pathway analysis was applied to whole exome sequence data for 2045 schizophrenic patients and 2045 controls. Overall, there was a statistically significant excess of pathways with more rare, functional variants in cases than in controls. Among the highest ranked were pathways relating to histone modification, as well as neuron differentiation and membrane and vesicle function. This bolsters the evidence from previous studies that histone modification pathways may be important in the aetiology of schizophrenia.

  10. Dynamic fuzziness during linker histone action.

    Science.gov (United States)

    McBryant, Steven J; Hansen, Jeffrey C

    2012-01-01

    Linker histones are multi-domain nucleosome binding proteins that stabilize higher order chromatin structures and engage in specific protein-protein interactions. Here we emphasize the structural and functional properties of the linker histone C-terminal domain (CTD), focusing on its intrinsic disorder, interaction-induced secondary structure formation and dynamic fuzziness. We argue that the fuzziness inherent in the CTD is a primary molecular mechanism underlying linker histone function in the nucleus.

  11. The role of histone H4 biotinylation in the structure of nucleosomes.

    Directory of Open Access Journals (Sweden)

    Nina A Filenko

    Full Text Available BACKGROUND: Post-translational modifications of histones play important roles in regulating nucleosome structure and gene transcription. It has been shown that biotinylation of histone H4 at lysine-12 in histone H4 (K12Bio-H4 is associated with repression of a number of genes. We hypothesized that biotinylation modifies the physical structure of nucleosomes, and that biotin-induced conformational changes contribute to gene silencing associated with histone biotinylation. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis we used atomic force microscopy to directly analyze structures of nucleosomes formed with biotin-modified and non-modified H4. The analysis of the AFM images revealed a 13% increase in the length of DNA wrapped around the histone core in nucleosomes with biotinylated H4. This statistically significant (p<0.001 difference between native and biotinylated nucleosomes corresponds to adding approximately 20 bp to the classical 147 bp length of nucleosomal DNA. CONCLUSIONS/SIGNIFICANCE: The increase in nucleosomal DNA length is predicted to stabilize the association of DNA with histones and therefore to prevent nucleosomes from unwrapping. This provides a mechanistic explanation for the gene silencing associated with K12Bio-H4. The proposed single-molecule AFM approach will be instrumental for studying the effects of various epigenetic modifications of nucleosomes, in addition to biotinylation.

  12. Restriction of histone gene transcription to S phase by phosphorylation of a chromatin boundary protein.

    Science.gov (United States)

    Kurat, Christoph F; Lambert, Jean-Philippe; van Dyk, Dewald; Tsui, Kyle; van Bakel, Harm; Kaluarachchi, Supipi; Friesen, Helena; Kainth, Pinay; Nislow, Corey; Figeys, Daniel; Fillingham, Jeffrey; Andrews, Brenda J

    2011-12-01

    The cell cycle-regulated expression of core histone genes is required for DNA replication and proper cell cycle progression in eukaryotic cells. Although some factors involved in histone gene transcription are known, the molecular mechanisms that ensure proper induction of histone gene expression during S phase remain enigmatic. Here we demonstrate that S-phase transcription of the model histone gene HTA1 in yeast is regulated by a novel attach-release mechanism involving phosphorylation of the conserved chromatin boundary protein Yta7 by both cyclin-dependent kinase 1 (Cdk1) and casein kinase 2 (CK2). Outside S phase, integrity of the AAA-ATPase domain is required for Yta7 boundary function, as defined by correct positioning of the histone chaperone Rtt106 and the chromatin remodeling complex RSC. Conversely, in S phase, Yta7 is hyperphosphorylated, causing its release from HTA1 chromatin and productive transcription. Most importantly, abrogation of Yta7 phosphorylation results in constitutive attachment of Yta7 to HTA1 chromatin, preventing efficient transcription post-recruitment of RNA polymerase II (RNAPII). Our study identified the chromatin boundary protein Yta7 as a key regulator that links S-phase kinases with RNAPII function at cell cycle-regulated histone gene promoters.

  13. Human borna disease virus infection impacts host proteome and histone lysine acetylation in human oligodendroglia cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xia [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Department of Neurology, The Fifth People' s Hospital of Shanghai, School of Medicine, Fudan University, Shanghai, 200240 (China); Zhao, Libo [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Department of Neurology, The Third People' s Hospital of Chongqing, 400014 (China); Yang, Yongtao [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Bode, Liv [Bornavirus Research Group affiliated to the Free University of Berlin, Berlin (Germany); Huang, Hua [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Liu, Chengyu [Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Huang, Rongzhong [Department of Rehabilitative Medicine, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010 (China); Zhang, Liang [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); and others

    2014-09-15

    Background: Borna disease virus (BDV) replicates in the nucleus and establishes persistent infections in mammalian hosts. A human BDV strain was used to address the first time, how BDV infection impacts the proteome and histone lysine acetylation (Kac) of human oligodendroglial (OL) cells, thus allowing a better understanding of infection-driven pathophysiology in vitro. Methods: Proteome and histone lysine acetylation were profiled through stable isotope labeling for cell culture (SILAC)-based quantitative proteomics. The quantifiable proteome was annotated using bioinformatics. Histone acetylation changes were validated by biochemistry assays. Results: Post BDV infection, 4383 quantifiable differential proteins were identified and functionally annotated to metabolism pathways, immune response, DNA replication, DNA repair, and transcriptional regulation. Sixteen of the thirty identified Kac sites in core histones presented altered acetylation levels post infection. Conclusions: BDV infection using a human strain impacted the whole proteome and histone lysine acetylation in OL cells. - Highlights: • A human strain of BDV (BDV Hu-H1) was used to infect human oligodendroglial cells (OL cells). • This study is the first to reveal the host proteomic and histone Kac profiles in BDV-infected OL cells. • BDV infection affected the expression of many transcription factors and several HATs and HDACs.

  14. Archaeal histones: dynamic and versatile genome architects

    Directory of Open Access Journals (Sweden)

    Bram Henneman

    2015-12-01

    Full Text Available Genome organization and compaction in Archaea involves different chromatin proteins, among which homologues of eukaryotic histones. Archaeal histones are considered the ancestors of their eukaryotic counterparts, which isreflected in the way they position along the genome and wrap DNA. Evolution from the archaeal modes of action to the prototypical eukaryotic nucleosome may be attributed to altered histone-histone interactions and DNA sequence determinants cooperating to yield stable multimeric structures. The identification of a new candidate phylum, proposed to be a missing link between archaea and eukaryotes, Lokiarchaeaota, may be instrumental in addressing this hypothesis.

  15. Uncoupling histone turnover from transcription-associated histone H3 modifications.

    Science.gov (United States)

    Ferrari, Paolo; Strubin, Michel

    2015-04-30

    Transcription in eukaryotes is associated with two major changes in chromatin organization. Firstly, nucleosomal histones are continuously replaced by new histones, an event that in yeast occurs predominantly at transcriptionally active promoters. Secondly, histones become modified post-translationally at specific lysine residues. Some modifications, including histone H3 trimethylation at lysine 4 (H3K4me3) and acetylation at lysines 9 (H3K9ac) and 14 (H3K14ac), are specifically enriched at active promoters where histones exchange, suggesting a possible causal relationship. Other modifications accumulate within transcribed regions and one of them, H3K36me3, is thought to prevent histone exchange. Here we explored the relationship between these four H3 modifications and histone turnover at a few selected genes. Using lysine-to-arginine mutants and a histone exchange assay, we found that none of these modifications plays a major role in either promoting or preventing histone turnover. Unexpectedly, mutation of H3K56, whose acetylation occurs prior to chromatin incorporation, had an effect only when introduced into the nucleosomal histone. Furthermore, we used various genetic approaches to show that histone turnover can be experimentally altered with no major consequence on the H3 modifications tested. Together, these results suggest that transcription-associated histone turnover and H3 modification are two correlating but largely independent events.

  16. Crystal Structure and Function of Human Nucleoplasmin (Npm2): A Histone Chaperone in Oocytes and Embryos

    Energy Technology Data Exchange (ETDEWEB)

    O Platonova; I Akey; J Head; C Akey

    2011-12-31

    Human Npm2 is an ortholog of Xenopus nucleoplasmin (Np), a chaperone that binds histones. We have determined the crystal structure of a truncated Npm2-core at 1.9 {angstrom} resolution and show that the N-terminal domains of Npm2 and Np form similar pentamers. This allowed us to model an Npm2 decamer which may be formed by hydrogen bonds between quasi-conserved residues in the interface between two pentamers. Interestingly, the Npm2 pentamer lacks a prototypical A1-acidic tract in each of its subunits. This feature may be responsible for the inability of Npm2-core to bind histones. However, Npm2 with a large acidic tract in its C-terminal tail (Npm2-A2) is able to bind histones and form large complexes. Fluorescence resonance energy transfer experiments and biochemical analysis of loop mutations support the premise that nucleoplasmins form decamers when they bind H2A-H2B dimers and H3-H4 tetramers simultaneously. In the absence of histone tetramers, these chaperones bind H2A-H2B dimers with a single pentamer forming the central hub. When taken together, our data provide insights into the mechanism of histone binding by nucleoplasmins.

  17. Transcription initiation factor IID-interactive histone chaperone CIA-II implicated in mammalian spermatogenesis.

    Science.gov (United States)

    Umehara, Takashi; Horikoshi, Masami

    2003-09-12

    Histones are thought to have specific roles in mammalian spermatogenesis, because several subtypes of histones emerge that are post-translationally modified during spermatogenesis. Though regular assembly of nucleosome is guaranteed by histone chaperones, their involvement in spermatogenesis is yet to be characterized. Here we identified a histone chaperone-related factor, which we designated as CCG1-interacting factor A-II (CIA-II), through interaction with bromodomains of TAFII250/CCG1, which is the largest subunit of human transcription initiation factor IID (TFIID). We found that human CIA-II (hCIA-II) localizes in HeLa nuclei and is highly expressed in testis and other proliferating cell-containing tissues. Expression of mouse CIA-II (mCIA-II) does not occur in the germ cell-lacking testes of adult WBB6F1-W/Wv mutant mice, indicating its expression in testis to be specific to germ cells. Fractionation of testicular germ cells revealed that mCIA-II transcripts accumulate in pachytene spermatocytes but not in spermatids. In addition, the mCIA-II transcripts in testis were present as early as 4 days after birth and decreased at 56 days after birth. These findings indicate that mCIA-II expression in testis is restricted to premeiotic to meiotic stages during spermatogenesis. Also, we found that hCIA-II interacts with histone H3 in vivo and with histones H3/H4 in vitro and that it facilitates supercoiling of circular DNA when it is incubated with core histones and topoisomerase I in vitro. These data suggest that CIA-II is a histone chaperone and is implicated in the regulation of mammalian spermatogenesis.

  18. MAP kinases and histone modification

    Institute of Scientific and Technical Information of China (English)

    Tamaki Suganuma; Jerry L. Workman

    2012-01-01

    Signal transduction pathways alter the gene expression program in response to extracellular or intracellular cues.Mitogen-activated protein kinases (MAPKs) govern numerous cellular processes including cell growth,stress response,apoptosis,and differentiation.In the past decade,MAPKs have been shown to regulate the transcription machinery and associate with chromatin-modifying complexes.Moreover,recent studies demonstrate that several MAPKs bind directly to chromatin at target genes.This review highlights the recent discoveries of MAPK signaling in regard to histone modifications and chromatin regulation.Evidence suggesting that further unknown mechanisms integrate signal transduction with chromatin biology is discussed.

  19. Histone acetyltransferases: challenges in targeting bi-substrate enzymes

    National Research Council Canada - National Science Library

    Wapenaar, Hannah; Dekker, Frank J

    2016-01-01

    Histone acetyltransferases (HATs) are epigenetic enzymes that install acetyl groups onto lysine residues of cellular proteins such as histones, transcription factors, nuclear receptors, and enzymes...

  20. Ethanol precipitation analysis of thymus histone

    NARCIS (Netherlands)

    Bijvoet, P.

    1957-01-01

    An analytical ethanol precipitation technique, similar to 's salting-out procedure, was used for the characterisation of whole thymus histone and the products obtained by preparative ethanol fractionation. The analysis was carried out at —5° C and pH 6.5. Whole histone prepared according to et al.,

  1. Histone modifications: Cycling with chromosomal replication

    DEFF Research Database (Denmark)

    Thon, Genevieve

    2008-01-01

    Histone modifications tend to be lost during chromosome duplication. Several recent studies suggest that the RNA interference pathway becomes active during the weakened transcriptional repression occurring at centromeres in S phase, resulting in the re-establishment of histone modifications that ...

  2. Recognition of the centromere-specific histone Cse4 by the chaperone Scm3

    OpenAIRE

    Cho, Uhn-Soo; Harrison, Stephen C.

    2011-01-01

    A specialized nucleosome is a component of all eukaryotic kinetochores. The core of this nucleosome contains a centromere-specific histone, CENP-A (the Cse4 gene product in budding yeast), instead of the usual H3. Assembly of a centromeric nucleosome depends on a specific chaperone, called Scm3 in yeast and HJURP in higher eukaryotes. We describe here the structure of a complex formed by an N-terminal fragment of Scm3 with the histone-fold domains of Cse4, and H4, all prepared as recombinant ...

  3. Acetylated histone H3 increases nucleosome dissociation

    Science.gov (United States)

    Simon, Marek; Manohar, Mridula; Ottesen, Jennifer; Poirier, Michael

    2009-03-01

    Chromatin's basic unit structure is the nucleosome, i.e. genomic DNA wrapped around a particular class of proteins -- histones -- which due to their physical hindrance, block vital biological processes, such as DNA repair, DNA replication, and RNA transcription. Histone post-translational modifications, which are known to exist in vivo, are hypothesized to regulate these biological processes by directly altering DNA-histone interactions and thus nucleosome structure and stability. Using magnetic tweezers technique we studied the acetylation of histone H3 in the dyad region, i.e. at K115 and K122, on reconstituted arrays of nucleosomes under constant external force. Based on the measured increase in the probability of dissociation of modified nucleosomes, we infer that this double modification could facilitate histone chaperone mediated nucleosome disassembly in vivo.

  4. Recognition of the centromere-specific histone Cse4 by the chaperone Scm3

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Uhn-Soo; Harrison, Stephen C. (Harvard-Med)

    2011-09-20

    A specialized nucleosome is a component of all eukaryotic kinetochores. The core of this nucleosome contains a centromere-specific histone, CENP-A (the Cse4 gene product in budding yeast), instead of the usual H3. Assembly of a centromeric nucleosome depends on a specific chaperone, called Scm3 in yeast and HJURP in higher eukaryotes. We describe here the structure of a complex formed by an N-terminal fragment of Scm3 with the histone-fold domains of Cse4, and H4, all prepared as recombinant proteins derived from the budding yeast Kluyveromyces lactis. The contacts of Scm3 with Cse4 explain its selectivity for the centromere-specific histone; key residues at the interface are conserved in HJURP, indicating a common mechanism for centromeric-histone deposition. We also report the structure of a (Cse4 : H4)2 heterotetramer; comparison with the structure of the Scm3:Cse4:H4 complex shows that tetramer formation and DNA-binding require displacement of Scm3 from the nucleosome core. The two structures together suggest that specific contacts between the chaperone and Cse4, rather than an altered overall structure of the nucleosome core, determine the selective presence of Cse4 at centromeres.

  5. Structure and function of human histone H3.Y nucleosome.

    Science.gov (United States)

    Kujirai, Tomoya; Horikoshi, Naoki; Sato, Koichi; Maehara, Kazumitsu; Machida, Shinichi; Osakabe, Akihisa; Kimura, Hiroshi; Ohkawa, Yasuyuki; Kurumizaka, Hitoshi

    2016-07-27

    Histone H3.Y is a primate-specific, distant H3 variant. It is evolutionarily derived from H3.3, and may function in transcription regulation. However, the mechanism by which H3.Y regulates transcription has not been elucidated. In the present study, we determined the crystal structure of the H3.Y nucleosome, and found that many H3.Y-specific residues are located on the entry/exit sites of the nucleosome. Biochemical analyses revealed that the DNA ends of the H3.Y nucleosome were more flexible than those of the H3.3 nucleosome, although the H3.Y nucleosome was stable in vitro and in vivo Interestingly, the linker histone H1, which compacts nucleosomal DNA, appears to bind to the H3.Y nucleosome less efficiently, as compared to the H3.3 nucleosome. These characteristics of the H3.Y nucleosome are also conserved in the H3.Y/H3.3 heterotypic nucleosome, which may be the predominant form in cells. In human cells, H3.Y preferentially accumulated around transcription start sites (TSSs). Taken together, H3.Y-containing nucleosomes around transcription start sites may form relaxed chromatin that allows transcription factor access, to regulate the transcription status of specific genes.

  6. Inhibition of mitotic-specific histone phophorylation by sodium arsenite

    Energy Technology Data Exchange (ETDEWEB)

    Cobo, J.M. [Universidad de Alcala de Henares, Madrid (Spain); Valdez, J.G.; Gurley, L.R. [Los Alamos National Lab., NM (United States)

    1994-10-01

    Synchronized cultures of Chinese hamster cells (line CHO) were used to measure the effects of 10{mu}M sodium arsenite on histone phosphorylation. This treatment caused cell proliferation to be temporarily arrested, after which the cells spontaneously resumed cell proliferation in a radiomimetric manner. Immediately following treatment, it was found that sodium arsenite affected only mitotic-specific HI and H3 phosphorylations. Neither interphase, nor mitotic, H2A and H4 phosphorylations were affected, nor was interphase HI Phosphorylation affected. The phosphorylation of HI was inhibited only in mitosis, reducing HI phosphorylation to 38.1% of control levels, which was the level of interphase HI phosphorylation. The phosphorylation of both H3 variants was inhibited in mitosis, the less hydrophobic H3 to 19% and the more hydrophobic H3 to 24% of control levels. These results suggest that sodium arsenite may inhibite cell proliferation by interfering with the cyclin B/p34{sup cdc2} histone kinase activity which is thought to play a key role in regulating the cell cycle. It has been proposed by our laboratory that HI and H3 phosphorylations play a role in restructuring interphase chromatin into metaphase chromosomes. Interference of this process by sodium arsenite may lead to structurally damaged chromosomes resulting in the increased cancer risks known to be produced by arsenic exposure from the environment.

  7. Chromatin condensation in terminally differentiating mouse erythroblasts does not involve special architectural proteins but depends on histone deacetylation

    Energy Technology Data Exchange (ETDEWEB)

    Popova, Evgenya Y.; Krauss, Sharon Wald; Short, Sarah A.; Lee, Gloria; Villalobos, Jonathan; Etzell, Joan; Koury, Mark J.; Ney, Paul A.; Chasis, Joel Anne; Grigoryev, Sergei A.

    2008-08-21

    Terminal erythroid differentiation in vertebrates is characterized by progressive heterochromatin formation, chromatin condensation and, in mammals, culminates in nuclear extrusion. To date, although mechanisms regulating avian erythroid chromatin condensation have been identified, little is known regarding this process during mammalian erythropoiesis. To elucidate the molecular basis for mammalian erythroblast chromatin condensation, we used Friend virus-infected murine spleen erythroblasts that undergo terminal differentiation in vitro. Chromatin isolated from early and late stage erythroblasts had similar levels of linker and core histones, only a slight difference in nucleosome repeats, and no significant accumulation of known developmentally-regulated architectural chromatin proteins. However, histone H3(K9) dimethylation markedly increased while histone H4(K12) acetylation dramatically decreased and became segregated from the histone methylation as chromatin condensed. One histone deacetylase, HDAC5, was significantly upregulated during the terminal stages of Friend virus-infected erythroblast differentiation. Treatment with histone deacetylase inhibitor, trichostatin A, blocked both chromatin condensation and nuclear extrusion. Based on our data, we propose a model for a unique mechanism in which extensive histone deacetylation at pericentromeric heterochromatin mediates heterochromatin condensation in vertebrate erythroblasts that would otherwise be mediated by developmentally-regulated architectural proteins in nucleated blood cells.

  8. Cell differentiation and development in Arabidopsis are associated with changes in histone dynamics at the single-cell level.

    Science.gov (United States)

    Rosa, Stefanie; Ntoukakis, Vardis; Ohmido, Nobuko; Pendle, Ali; Abranches, Rita; Shaw, Peter

    2014-12-01

    The mechanism whereby the same genome can give rise to different cell types with different gene expression profiles is a fundamental problem in biology. Chromatin organization and dynamics have been shown to vary with altered gene expression in different cultured animal cell types, but there is little evidence yet from whole organisms linking chromatin dynamics with development. Here, we used both fluorescence recovery after photobleaching and two-photon photoactivation to show that in stem cells from Arabidopsis thaliana roots the mobility of the core histone H2B, as judged by exchange dynamics, is lower than in the surrounding cells of the meristem. However, as cells progress from meristematic to fully differentiated, core histones again become less mobile and more strongly bound to chromatin. We show that these transitions are largely mediated by changes in histone acetylation. We further show that altering histone acetylation levels, either in a mutant or by drug treatment, alters both the histone mobility and markers of development and differentiation. We propose that plant stem cells have relatively inactive chromatin, but they keep the potential to divide and differentiate into more dynamic states, and that these states are at least in part determined by histone acetylation levels. © 2014 American Society of Plant Biologists. All rights reserved.

  9. Histone tail modifications and noncanonical functions of histones: perspectives in cancer epigenetics.

    Science.gov (United States)

    Hadnagy, Annamaria; Beaulieu, Raymond; Balicki, Danuta

    2008-04-01

    Over the past few years, the histone deacetylase (HDAC) inhibitors have occupied an important place in the effort to develop novel, but less toxic, anticancer therapy. HDAC inhibitors block HDACs, which are the enzymes responsible for histone deacetylation, and therefore they modulate gene expression. The cellular effects of HDAC inhibitors include growth arrest and the induction of differentiation. Early successes in cancer therapeutics obtained using these drugs alone or in combination with other anticancer drugs emphasize the important place of posttranslational modifications of histones in cancer therapy. Histone tail modifications along with DNA methylation are the most studied epigenetic events related to cancer progression. Moreover, extranuclear functions of histones have also been described. Because HDAC inhibitors block HDACs and thereby increase histone acetylation, we propose a model wherein exogenous acetylated histones or other related acetylated proteins that are introduced into the nucleus become HDAC substrates and thereby compete with endogenous histones for HDACs. This competition may lead to the increased acetylation of the endogenous histones, as in the case of HDAC inhibitor therapy. Moreover, other mechanisms of action, such as binding to chromatin and modulating gene expression, are also possible for exogenously introduced histones.

  10. Preferential Phosphorylation on Old Histones during Early Mitosis in Human Cells.

    Science.gov (United States)

    Lin, Shu; Yuan, Zuo-Fei; Han, Yumiao; Marchione, Dylan M; Garcia, Benjamin A

    2016-07-15

    How histone post-translational modifications (PTMs) are inherited through the cell cycle remains poorly understood. Canonical histones are made in the S phase of the cell cycle. Combining mass spectrometry-based technologies and stable isotope labeling by amino acids in cell culture, we question the distribution of multiple histone PTMs on old versus new histones in synchronized human cells. We show that histone PTMs can be grouped into three categories according to their distributions. Most lysine mono-methylation and acetylation PTMs are either symmetrically distributed on old and new histones or are enriched on new histones. In contrast, most di- and tri-methylation PTMs are enriched on old histones, suggesting that the inheritance of different PTMs is regulated distinctly. Intriguingly, old and new histones are distinct in their phosphorylation status during early mitosis in the following three human cell types: HeLa, 293T, and human foreskin fibroblast cells. The mitotic hallmark H3S10ph is predominantly associated with old H3 at early mitosis and becomes symmetric with the progression of mitosis. This same distribution was observed with other mitotic phosphorylation marks, including H3T3/T6ph, H3.1/2S28ph, and H1.4S26ph but not S28/S31ph on the H3 variant H3.3. Although H3S10ph often associates with the neighboring Lys-9 di- or tri-methylations, they are not required for the asymmetric distribution of Ser-10 phosphorylation on the same H3 tail. Inhibition of the kinase Aurora B does not change the distribution despite significant reduction of H3S10ph levels. However, K9me2 abundance on the new H3 is significantly reduced after Aurora B inhibition, suggesting a cross-talk between H3S10ph and H3K9me2. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. An H1 histone gene from rainbow trout (Salmo gairdnerii).

    Science.gov (United States)

    Mezquita, J; Connor, W; Winkfein, R J; Dixon, G H

    A 1.7-kbp DNA region from the 10.2-kb cluster containing the five rainbow trout histone genes has been subcloned in pBR322 and completely sequenced. It contains a trout histone H1 gene together with its 5' and 3' flanking sequences. This H1 gene codes for a H1 variant different from the major trout testis H1 previously sequenced by Macleod et al. (1977). Northern blots of total RNA from trout testis, kidney, and liver indicate that this H1 gene is expressed in all three tissues but that the level of H1 mRNA is much higher in testis than in other tissues. The lack of heterogeneity in the sizes and 5' initiation sites of trout H1 mRNAs is surprising in view of the substantial heterogeneity of H1 variant proteins observed previously. The coding sequence of the H1 gene shows strong evidence of repeated partial duplications of a hexapeptide motif of the form Ala.Ala.Ala.Lys.Lys.Pro and of a pentapeptide phosphorylation-site sequence, Lys.Ser.Pro.Lys.Lys, during its evolution. Comparisons are drawn between this gene and the coding sequences of other vertebrate H1 genes from chicken and Xenopus, and a strong homology is seen in the region of amino acids 22-101, which form the hydrophobic "head" of the H1 molecule. The 5' and 3' regulatory signals in the trout H1 are also compared with those of H1 genes from other sequences.

  12. Structural Insights into Selective Histone H3 Recognition by the Human Polybromo bromodomain 2

    Energy Technology Data Exchange (ETDEWEB)

    Charlop-Powers, Z.; Zeng, L; Zhang, Q; Zhou, M

    2010-01-01

    The Polybromo (PB) protein functions as a key component of the human PBAF chromatin remodeling complex in regulation of gene transcription. PB is made up of modular domains including six bromodomains that are known as acetyl-lysine binding domains. However, histone-binding specificity of the bromodomains of PB has remained elusive. In this study, we report biochemical characterization of all six PB bromodomains' binding to a suite of lysine-acetylated peptides derived from known acetylation sites on human core histones. We demonstrate that bromodomain 2 of PB preferentially recognizes acetylated lysine 14 of histone H3 (H3K14ac), a post-translational mark known for gene transcriptional activation. We further describe the molecular basis of the selective H3K14ac recognition of bromodomain 2 by solving the protein structures in both the free and bound forms using X-ray crystallography and NMR, respectively.

  13. Identification and characterization of lysine-methylated sites on histones and non-histone proteins.

    Science.gov (United States)

    Lee, Tzong-Yi; Chang, Cheng-Wei; Lu, Cheng-Tzung; Cheng, Tzu-Hsiu; Chang, Tzu-Hao

    2014-06-01

    Protein methylation is a kind of post-translational modification (PTM), and typically takes place on lysine and arginine amino acid residues. Protein methylation is involved in many important biological processes, and most recent studies focused on lysine methylation of histones due to its critical roles in regulating transcriptional repression and activation. Histones possess highly conserved sequences and are homologous in most species. However, there is much less sequence conservation among non-histone proteins. Therefore, mechanisms for identifying lysine-methylated sites may greatly differ between histones and non-histone proteins. Nevertheless, this point of view was not considered in previous studies. Here we constructed two support vector machine (SVM) models by using lysine-methylated data from histones and non-histone proteins for predictions of lysine-methylated sites. Numerous features, such as the amino acid composition (AAC) and accessible surface area (ASA), were used in the SVM models, and the predictive performance was evaluated using five-fold cross-validations. For histones, the predictive sensitivity was 85.62% and specificity was 80.32%. For non-histone proteins, the predictive sensitivity was 69.1% and specificity was 88.72%. Results showed that our model significantly improved the predictive accuracy of histones compared to previous approaches. In addition, features of the flanking region of lysine-methylated sites on histones and non-histone proteins were also characterized and are discussed. A gene ontology functional analysis of lysine-methylated proteins and correlations of lysine-methylated sites with other PTMs in histones were also analyzed in detail. Finally, a web server, MethyK, was constructed to identify lysine-methylated sites. MethK now is available at http://csb.cse.yzu.edu.tw/MethK/.

  14. Structure, localization and histone binding properties of nuclear-associated nucleosome assembly protein from Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Maier Alexander G

    2010-04-01

    Full Text Available Abstract Background Nucleosome assembly proteins (NAPs are histone chaperones that are crucial for the shuttling and incorporation of histones into nucleosomes. NAPs participate in the assembly and disassembly of nucleosomes thus contributing to chromatin structure organization. The human malaria parasite Plasmodium falciparum contains two nucleosome assembly proteins termed PfNapL and PfNapS. Methods Three-dimensional crystal structure of PfNapS has been determined and analysed. Gene knockout and localization studies were also performed on PfNapS using transfection studies. Fluorescence spectroscopy was performed to identify histone-binding sites on PfNapS. Extensive sequence and structural comparisons were done with the crystal structures available for NAP/SET family of proteins. Results Crystal structure of PfNapS shares structural similarity with previous structures from NAP/SET family. Failed attempts to knock-out the gene for PfNapS from malaria parasite suggest essentiality in the parasite. GFP-fused PfNapS fusion protein targeting indicates cellular localization of PfNapS in the parasite nucleus. Fluorescence spectroscopy data suggest that PfNapS interacts with core histones (tetramer, octamer, H3, H4, H2A and H2B at a different site from its interaction with linker histone H1. This analysis illustrates two regions on the PfNapS dimer as the possible sites for histone recognition. Conclusions This work presents a thorough analysis of the structural, functional and regulatory attributes of PfNapS from P. falciparum with respect to previously studied histone chaperones.

  15. Histone demethylases in development and disease

    DEFF Research Database (Denmark)

    Pedersen, Marianne Terndrup; Helin, Kristian

    2010-01-01

    Histone modifications serve as regulatory marks that are instrumental for the control of transcription and chromatin architecture. Strict regulation of gene expression patterns is crucial during development and differentiation, where diverse cell types evolve from common predecessors. Since...... the first histone lysine demethylase was discovered in 2004, a number of demethylases have been identified and implicated in the control of gene expression programmes and cell fate decisions. Histone demethylases are now emerging as important players in developmental processes and have been linked to human...

  16. Dynamics of Histone Tails within Chromatin

    Science.gov (United States)

    Bernier, Morgan; North, Justin; Page, Michael; Jaroniec, Christopher; Hammel, Christopher; Poirier, Michael

    2012-02-01

    Genetic information in humans is encoded within DNA molecules that is wrapped around histone octamer proteins and compacted into a highly conserved structural polymer, chromatin. The physical and material properties of chromatin appear to influence gene expression by altering the accessibility of proteins to the DNA. The tails of the histones are flexible domains that are thought to play a role in regulating DNA accessibility and compaction; however the molecular mechanisms for these phenomena are not understood. I will present CW-EPR studies on site directed spin labeled nucleosomes that probe the structure and dynamics of these histone tails within nucleosomes.

  17. The impact of solubility and electrostatics on fibril formation by the H3 and H4 histones.

    Science.gov (United States)

    Topping, Traci B; Gloss, Lisa M

    2011-12-01

    The goal of this study was to examine fibril formation by the heterodimeric eukaryotic histones (H2A-H2B and H3-H4) and homodimeric archaeal histones (hMfB and hPyA1). The histone fold dimerization motif is an obligatorily domain-swapped structure comprised of two fused helix:β-loop:helix motifs. Domain swapping has been proposed as a mechanism for the evolution of protein oligomers as well as a means to form precursors in the formation of amyloid-like fibrils. Despite sharing a common fold, the eukaryotic histones of the core nucleosome and archaeal histones fold by kinetic mechanisms of differing complexity with transient population of partially folded monomeric and/or dimeric species. No relationship was apparent between fibrillation propensity and equilibrium stability or population of kinetic intermediates. Only H3 and H4, as isolated monomers and as a heterodimer, readily formed fibrils at room temperature, and this propensity correlates with the significantly lower solubility of these polypeptides. The fibrils were characterized by ThT fluorescence, FTIR, and far-UV CD spectroscopies and electron microscopy. The helical histone fold comprises the protease-resistant core of the fibrils, with little or no protease protection of the poorly structured N-terminal tails. The highly charged tails inhibit fibrillation through electrostatic repulsion. Kinetic studies indicate that H3 and H4 form a co-fibril, with simultaneous incorporation of both histones. The potential impact of H3 and H4 fibrillation on the cytotoxicity of extracellular histones and α-synuclein-mediated neurotoxicity and fibrillation is considered. Copyright © 2011 The Protein Society.

  18. Molecular mechanisms and potential functions of histone demethylases

    DEFF Research Database (Denmark)

    Kooistra, Susanne Marije; Helin, Kristian

    2012-01-01

    Histone modifications are thought to regulate chromatin structure, transcription and other nuclear processes. Histone methylation was originally believed to be an irreversible modification that could only be removed by histone eviction or by dilution during DNA replication. However, the isolation...... of two families of enzymes that can demethylate histones has changed this notion. The biochemical activities of these histone demethylases towards specific Lys residues on histones, and in some cases non-histone substrates, have highlighted their importance in developmental control, cell-fate decisions...

  19. Posttranslational modifications of the histone 3 tail and their impact on the activity of histone lysine demethylases in vitro.

    Directory of Open Access Journals (Sweden)

    Brian Lohse

    Full Text Available Posttranslational modifications (PTMs of the histone H3 tail such as methylation, acetylation and phosphorylation play important roles in epigenetic signaling. Here we study the effect of some of these PTMs on the demethylation rates of methylated lysine 9 in vitro using peptide substrates mimicking histone H3. Various combinations with other PTMs were employed to study possible cross-talk effects by comparing enzyme kinetic characteristics. We compared the kinetics of histone tail substrates for truncated histone lysine demethylases KDM4A and KDM4C containing only the catalytic core (cc and some combinations were characterized on full length (FL KDM4A and KDM4C. We found that the substrates combining trimethylated K4 and K9 resulted in a significant increase in the catalytic activity for FL-KDM4A. For the truncated versions of KDM4A and KDM4C a two-fold increase in the catalytic activity toward bis-trimethylated substrates could be observed. Furthermore, a significant difference in the catalytic activity between dimethylated and trimethylated substrates was found for full length demethylases in line with what has been reported previously for truncated demethylases. Histone peptide substrates phosphorylated at T11 could not be demethylated by neither truncated nor full length KDM4A and KDM4C, suggesting that phosphorylation of threonine 11 prevents demethylation of the H3K9me3 mark on the same peptide. Acetylation of K14 was also found to influence demethylation rates significantly. Thus, for truncated KDM4A, acetylation on K14 of the substrate leads to an increase in enzymatic catalytic efficiency (k cat/K m, while for truncated KDM4C it induces a decrease, primarily caused by changes in K m. This study demonstrates that demethylation activities towards trimethylated H3K9 are significantly influenced by other PTMs on the same peptide, and emphasizes the importance of studying these interactions at the peptide level to get a more detailed

  20. Transcription-Coupled Replacement of Histones: Degradation or Recycling?

    Institute of Scientific and Technical Information of China (English)

    Yu-Shan Chen; Xiao-Bo Qiu

    2012-01-01

    Histone modifications are proposed to constitute a “histone code” for epigenetic regulation of gene expression.However,recent studies demonstrate that histones have to be disassembled from chromatin during transcription.Recent evidence,though not conclusive,suggests that histones might be degradable after being removed from chromatin during transcription.Degradation of overexpressed excessive histones,instead of native histones,has been shown to be dependent on proteasomes and ubiquitination.Since the 26S proteasome usually recognizes polyubiquitinated substrates,it is critical to demonstrate whether degradation of histones is mediated by polyubiquitination.Unexpectedly,there is almost no evidence that any ubiquitin ligase can promote polyubiquitination-dependent degradation of constitutive histones.Meanwhile,acetylation and phosphorylation are also associated with histone degradation.This review attempts to summarize the current knowledge on the transcription-coupled degradation of histones and its regulation by posttranslational protein modifications.

  1. In vitro targeting reveals intrinsic histone tail specificity of the Sin3/histone deacetylase and N-CoR/SMRT corepressor complexes.

    NARCIS (Netherlands)

    Vermeulen, M.; Carrozza, M.J.; Lasonder, E.; Workman, J.L.; Logie, C.; Stunnenberg, H.G.

    2004-01-01

    The histone code is among others established via differential acetylation catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs). To unambiguously determine the histone tail specificity of HDAC-containing complexes, we have established an in vitro system consisting of

  2. Replication stress interferes with histone recycling and predeposition marking of new histones.

    Science.gov (United States)

    Jasencakova, Zuzana; Scharf, Annette N D; Ask, Katrine; Corpet, Armelle; Imhof, Axel; Almouzni, Geneviève; Groth, Anja

    2010-03-12

    To restore chromatin on new DNA during replication, recycling of histones evicted ahead of the fork is combined with new histone deposition. The Asf1 histone chaperone, which buffers excess histones under stress, is a key player in this process. Yet how histones handled by human Asf1 are modified remains unclear. Here we identify marks on histones H3-H4 bound to Asf1 and changes induced upon replication stress. In S phase, distinct cytosolic and nuclear Asf1b complexes show ubiquitous H4K5K12diAc and heterogeneous H3 marks, including K9me1, K14ac, K18ac, and K56ac. Upon acute replication arrest, the predeposition mark H3K9me1 and modifications typical of chromatin accumulate in Asf1 complexes. In parallel, ssDNA is generated at replication sites, consistent with evicted histones being trapped with Asf1. During recovery, histones stored with Asf1 are rapidly used as replication resumes. This shows that replication stress interferes with predeposition marking and histone recycling with potential impact on epigenetic stability.

  3. ADP-ribosylation of histones by ARTD1: an additional module of the histone code?

    Science.gov (United States)

    Hottiger, Michael O

    2011-06-01

    ADP-ribosylation is a covalent post-translational protein modification catalyzed by ADP-ribosyltransferases and is involved in important processes such as cell cycle regulation, DNA damage response, replication or transcription. Histones are ADP-ribosylated by ADP-ribosyltransferase diphtheria toxin-like 1 at specific amino acid residues, in particular lysines, of the histones tails. Specific ADP-ribosyl hydrolases and poly-ADP-ribose glucohydrolases degrade the ADP-ribose polymers. The ADP-ribose modification is read by zinc finger motifs or macrodomains, which then regulate chromatin structure and transcription. Thus, histone ADP-ribosylation may be considered an additional component of the histone code.

  4. Nucleosome Dancing at the Tempo of Histone Tail Acetylation

    Directory of Open Access Journals (Sweden)

    Angélique Galvani

    2015-07-01

    Full Text Available The impact of histone acetylation on transcription was revealed over 50 years ago by Allfrey and colleagues. However, it took decades for an understanding of the fine mechanism by which this posttranslational modification affects chromatin structure and promotes transcription. Here, we review breakthroughs linking histone tail acetylation, histone dynamics, and transcription. We also discuss the histone exchange during transcription and highlight the important function of a pool of non-chromatinized histones in chromatin dynamics.

  5. Histone acetylation in astrocytes suppresses GFAP and stimulates a reorganization of the intermediate filament network.

    Science.gov (United States)

    Kanski, Regina; Sneeboer, Marjolein A M; van Bodegraven, Emma J; Sluijs, Jacqueline A; Kropff, Wietske; Vermunt, Marit W; Creyghton, Menno P; De Filippis, Lidia; Vescovi, Angelo; Aronica, Eleonora; van Tijn, Paula; van Strien, Miriam E; Hol, Elly M

    2014-10-15

    Glial fibrillary acidic protein (GFAP) is the main intermediate filament in astrocytes and is regulated by epigenetic mechanisms during development. We demonstrate that histone acetylation also controls GFAP expression in mature astrocytes. Inhibition of histone deacetylases (HDACs) with trichostatin A or sodium butyrate reduced GFAP expression in primary human astrocytes and astrocytoma cells. Because splicing occurs co-transcriptionally, we investigated whether histone acetylation changes the ratio between the canonical isoform GFAPα and the alternative GFAPδ splice variant. We observed that decreased transcription of GFAP enhanced alternative isoform expression, as HDAC inhibition increased the GFAPδ∶GFAPα ratio. Expression of GFAPδ was dependent on the presence and binding of splicing factors of the SR protein family. Inhibition of HDAC activity also resulted in aggregation of the GFAP network, reminiscent of our previous findings of a GFAPδ-induced network collapse. Taken together, our data demonstrate that HDAC inhibition results in changes in transcription, splicing and organization of GFAP. These data imply that a tight regulation of histone acetylation in astrocytes is essential, because dysregulation of gene expression causes the aggregation of GFAP, a hallmark of human diseases like Alexander's disease.

  6. CFP1 Regulates Histone H3K4 Trimethylation and Developmental Potential in Mouse Oocytes

    Directory of Open Access Journals (Sweden)

    Chao Yu

    2017-08-01

    Full Text Available Trimethylation of histone H3 at lysine-4 (H3K4me3 is associated with eukaryotic gene promoters and poises their transcriptional activation during development. To examine the in vivo function of H3K4me3 in the absence of DNA replication, we deleted CXXC finger protein 1 (CFP1, the DNA-binding subunit of the SETD1 histone H3K4 methyltransferase, in developing oocytes. We find that CFP1 is required for H3K4me3 accumulation and the deposition of histone variants onto chromatin during oocyte maturation. Decreased H3K4me3 in oocytes caused global downregulation of transcription activity. Oocytes lacking CFP1 failed to complete maturation and were unable to gain developmental competence after fertilization, due to defects in cytoplasmic lattice formation, meiotic division, and maternal-zygotic transition. Our study highlights the importance of H3K4me3 in continuous histone replacement for transcriptional regulation, chromatin remodeling, and normal developmental progression in a non-replicative system.

  7. Point Mutations in Centromeric Histone Induce Post-zygotic Incompatibility and Uniparental Inheritance.

    Directory of Open Access Journals (Sweden)

    Sundaram Kuppu

    2015-09-01

    Full Text Available The centromeric histone 3 variant (CENH3, aka CENP-A is essential for the segregation of sister chromatids during mitosis and meiosis. To better define CENH3 functional constraints, we complemented a null allele in Arabidopsis with a variety of mutant alleles, each inducing a single amino acid change in conserved residues of the histone fold domain. Many of these transgenic missense lines displayed wild-type growth and fertility on self-pollination, but exhibited frequent post-zygotic death and uniparental inheritance when crossed with wild-type plants. The failure of centromeres marked by these missense mutation in the histone fold domain of CENH3 reproduces the genome elimination syndromes described with chimeric CENH3 and CENH3 from diverged species. Additionally, evidence that a single point mutation is sufficient to generate a haploid inducer provide a simple one-step method for the identification of non-transgenic haploid inducers in existing mutagenized collections of crop species. As proof of the extreme simplicity of this approach to create haploid-inducing lines, we performed an in silico search for previously identified point mutations in CENH3 and identified an Arabidopsis line carrying the A86V substitution within the histone fold domain. This A87V non-transgenic line, while fully fertile on self-pollination, produced postzygotic death and uniparental haploids when crossed to wild type.

  8. Point Mutations in Centromeric Histone Induce Post-zygotic Incompatibility and Uniparental Inheritance.

    Science.gov (United States)

    Kuppu, Sundaram; Tan, Ek Han; Nguyen, Hanh; Rodgers, Andrea; Comai, Luca; Chan, Simon W L; Britt, Anne B

    2015-09-01

    The centromeric histone 3 variant (CENH3, aka CENP-A) is essential for the segregation of sister chromatids during mitosis and meiosis. To better define CENH3 functional constraints, we complemented a null allele in Arabidopsis with a variety of mutant alleles, each inducing a single amino acid change in conserved residues of the histone fold domain. Many of these transgenic missense lines displayed wild-type growth and fertility on self-pollination, but exhibited frequent post-zygotic death and uniparental inheritance when crossed with wild-type plants. The failure of centromeres marked by these missense mutation in the histone fold domain of CENH3 reproduces the genome elimination syndromes described with chimeric CENH3 and CENH3 from diverged species. Additionally, evidence that a single point mutation is sufficient to generate a haploid inducer provide a simple one-step method for the identification of non-transgenic haploid inducers in existing mutagenized collections of crop species. As proof of the extreme simplicity of this approach to create haploid-inducing lines, we performed an in silico search for previously identified point mutations in CENH3 and identified an Arabidopsis line carrying the A86V substitution within the histone fold domain. This A87V non-transgenic line, while fully fertile on self-pollination, produced postzygotic death and uniparental haploids when crossed to wild type.

  9. Quantitative proteomic approaches to studying histone modifications.

    Science.gov (United States)

    Zee, Barry M; Young, Nicolas L; Garcia, Benjamin A

    2011-01-01

    Histone post-translational modifications (PTMs) positively and negatively regulate gene expression, and are consequently a vital influence on the genomic profile of all eukaryotic species. The study of histone PTMs using classical methods in molecular biology, such as immunofluorescence and Western blotting, is challenging given the technical issues of the approaches, and chemical diversity and combinatorial patterns of the modifications. In light of these many technical limitations, mass spectrometry (MS) is emerging as the most unbiased and rigorous experimental platform to identify and quantify histone PTMs in a high-throughput manner. This review covers the latest developments in mass spectrometry for the analysis of histone PTMs, with the hope of inspiring the continued integration of proteomic, genomic and epigenetic research.

  10. Behavioral variant of frontotemporal dementia mimicking Huntington's disease

    DEFF Research Database (Denmark)

    Nielsen, T Rune; Bruhn, Peter; Nielsen, Jørgen E

    2010-01-01

    Behavioral changes and cognitive decline are the core clinical manifestations in the behavioral variant of frontotemporal dementia (bv-FTD). The behavioral changes may include characteristic stereotypic movements. These movements, although without clear purpose, are not involuntary. Involuntary m...

  11. Histone Lysine Methylation in Diabetic Nephropathy

    Directory of Open Access Journals (Sweden)

    Guang-dong Sun

    2014-01-01

    Full Text Available Diabetic nephropathy (DN belongs to debilitating microvascular complications of diabetes and is the leading cause of end-stage renal diseases worldwide. Furthermore, outcomes from the DCCT/EDIC study showed that DN often persists and progresses despite intensive glucose control in many diabetes patients, possibly as a result of prior episode of hyperglycemia, which is called “metabolic memory.” The underlying mechanisms responsible for the development and progression of DN remain poorly understood. Activation of multiple signaling pathways and key transcription factors can lead to aberrant expression of DN-related pathologic genes in target renal cells. Increasing evidence suggests that epigenetic mechanisms in chromatin such as DNA methylation, histone acetylation, and methylation can influence the pathophysiology of DN and metabolic memory. Exciting researches from cell culture and experimental animals have shown that key histone methylation patterns and the related histone methyltransferases and histone demethylases can play important roles in the regulation of inflammatory and profibrotic genes in renal cells under diabetic conditions. Because histone methylation is dynamic and potentially reversible, it can provide a window of opportunity for the development of much-needed novel therapeutic potential for DN in the future. In this minireview, we discuss recent advances in the field of histone methylation and its roles in the pathogenesis and progression of DN.

  12. A brief histone in time: understanding the combinatorial functions of histone PTMs in the nucleosome context.

    Science.gov (United States)

    Ng, Marlee K; Cheung, Peter

    2016-02-01

    It has been over 50 years since Allfrey et al. proposed that histone acetylation regulates RNA synthesis, and the study of histone modifications has progressed at an extraordinary pace for the past two decades. In this review, we provide a perspective on some key events and advances in our understanding of histone modifications. We also highlight reagents and tools from past to present that facilitated progress in this research field. Using histone H3 phosphorylation as an underlying thread, we review the rationale that led to the proposal of the histone code hypothesis, as well as examples that illustrate the concepts of combinatorial histone modifications and cross-talk pathways. We further highlight the importance of investigating these mechanisms in the context of nucleosomes rather than just at the histone level and present current and developing approaches for such studies. Overall, research on histone modifications has yielded great mechanistic insights into the regulation of genomic functions, and extending these studies using nucleosomes will further elucidate the complexity of these pathways in a more physiologically relevant context.

  13. Neutron and x-ray scatter studies of the histone octamer and amino and carboxyl domain trimmed octamers.

    Science.gov (United States)

    Wood, M J; Yau, P; Imai, B S; Goldberg, M W; Lambert, S J; Fowler, A G; Baldwin, J P; Godfrey, J E; Moudrianakis, E N; Koch, M H

    1991-03-25

    The structure of the nucleosome has been under intense investigation using neutron crystallography, x-ray crystallography, and neutron solution scattering. However the dimension of the histone octamer inside the nucleosome is still a subject of controversy. The radius of gyration (Rg) of the octamer obtained from solution neutron scattering of core particles at 63% 2H2O, 37% 1H2O is 33 A, and x-ray crystallography study of isolated histone octamer gives a Rg of 32.5 A, while the reported values using x-ray crystallography of core particles from two individual studies are 29.7 and 30.4 A, respectively. We report here studies of isolated histone octamer and trypsin-limited digested octamer using both neutron solution scattering and small angle x-ray scattering. The Rg of the octamer obtained is 33 A, whereas that of the trimmed octamer is 29.8 A, similar to the structure obtained from the crystals of the core particles. The N-terminal domains of the core histones in the octamer have been shown by high resolution nuclear magnetic resonance (Schroth, G.P., Yau, P., Imai, B.S., Gatewood, J.M., and Bradbury, E.M. (1990) FEBS Lett. 268, 117-120) to be mobile and flexible; it is likely that these regions are disordered and "not seen" by x-ray crystallography.

  14. Hinge and chromoshadow of HP1α participate in recognition of K9 methylated histone H3 in nucleosomes.

    Science.gov (United States)

    Mishima, Yuichi; Watanabe, Makoto; Kawakami, Toru; Jayasinghe, Chanika D; Otani, Junji; Kikugawa, Yusuke; Shirakawa, Masahiro; Kimura, Hiroshi; Nishimura, Osamu; Aimoto, Saburo; Tajima, Shoji; Suetake, Isao

    2013-01-09

    The majority of the genome in eukaryotes is packaged into transcriptionally inactive chromatin. Heterochromatin protein 1 (HP1) is a major player in the establishment and maintenance of heterochromatin. HP1 specifically recognizes a methylated lysine residue at position 9 in histone H3 through its N-terminal chromo domain (CD). To elucidate the binding properties of HP1α to nucleosomes in vitro, we reconstituted nucleosomes containing histone H3 trimethylated at lysine 9. HP1α exhibited high-affinity binding to nucleosomes containing methylated histone H3 in a nucleosome core-number-dependent manner. The hinge region (HR) connecting the CD and C-terminal chromoshadow domain (CSD), and the CSD contributed to the selective binding of HP1α to histone H3 with trimethylated lysine 9 through weak DNA binding and by suppressing the DNA binding, respectively. We propose that not only the specific recognition of lysine 9 methylation of histone H3 by the CD but also the HR and the CSD cooperatively contribute to the selective binding of HP1α to histone H3 lysine 9 methylated nucleosomes.

  15. Cell-cycle-regulated control of VSG expression site silencing by histones and histone chaperones ASF1A and CAF-1b in Trypanosoma brucei.

    Science.gov (United States)

    Alsford, Sam; Horn, David

    2012-11-01

    Antigenic variation in African trypanosomes involves monoallelic expression and reversible silencing of variant surface glycoprotein (VSG) genes found adjacent to telomeres in polycistronic expression sites (ESs). We assessed the impact on ES silencing of five candidate essential chromatin-associated factors that emerged from a genome-wide RNA interference viability screen. Using this approach, we demonstrate roles in VSG ES silencing for two histone chaperones. Defects in S-phase progression in cells depleted for histone H3, or either chaperone, highlight in particular the link between chromatin assembly and DNA replication control. S-phase checkpoint arrest was incomplete, however, allowing G2/M-specific VSG ES derepression following knockdown of histone H3. In striking contrast, knockdown of anti-silencing factor 1A (ASF1A) allowed for derepression at all cell cycle stages, whereas knockdown of chromatin assembly factor 1b (CAF-1b) revealed derepression predominantly in S-phase and G2/M. Our results support a central role for chromatin in maintaining VSG ES silencing. ASF1A and CAF-1b appear to play constitutive and DNA replication-dependent roles, respectively, in the recycling and assembly of chromatin. Defects in these functions typically lead to arrest in S-phase but defective cells can also progress through the cell cycle leading to nucleosome depletion and derepression of telomeric VSG ESs.

  16. T rypanosoma brucei histone H1 inhibits RNA polymerase I transcription and is important for parasite fitness in vivo

    OpenAIRE

    Pena, Ana C.; Pimentel, Mafalda R; Manso, Helena; Vaz-Drago, Rita; Pinto-Neves, Daniel; Aresta-Branco, Francisco; Rijo-Ferreira, Filipa; Guegan, Fabien; Pedro Coelho, Luis; Carmo-Fonseca, Maria; Barbosa-Morais, Nuno L.; Figueiredo, Luisa M.

    2014-01-01

    T rypanosoma brucei is a unicellular parasite that causes sleeping sickness in humans. Most of its transcription is constitutive and driven by RNA polymerase II. RNA polymerase I (Pol I) transcribes not only ribosomal RNA genes, but also protein-encoding genes, including variant surface glycoproteins (VSGs) and procyclins. In T . brucei, histone H1 (H1) is required for VSG silencing and chromatin condensation. However, whether H1 has a genome-wide role in transcription is unknown. Here, using...

  17. The role of histone ubiquitination during spermatogenesis.

    Science.gov (United States)

    Sheng, Kai; Liang, Xiaotong; Huang, Sizhou; Xu, Wenming

    2014-01-01

    Protein ubiquitin-proteasome (ubiquitin-proteasome) system is the major mechanism responsible for protein degradation in eukaryotic cell. During spermatogenesis, the replacement of histone by protamine is vital for normal sperm formation, which is involved in ubiquitination enzymes expressed in testis. Recently, histone ubiquitin ligases have been shown to play critical roles in several aspects of spermatogenesis, such as meiotic sex chromosome inactivation (MSCI), DNA damage response, and spermiogenesis. In this review, we highlight recent progress in the discovery of several histone ubiquitin ligases and elaborate mechanisms of how these enzymes are involved in these processes through knockout mouse model. Using Huwe1, UBR2, and RNF8 as examples, we emphasized the diverse functions for each enzyme and the broad involvement of these enzymes in every stage, from spermatogonia differentiation and meiotic division to spermiogenesis; thus histone ubiquitin ligases represent a class of enzymes, which play important roles in spermatogenesis through targeting histone for ubiquitination and therefore are involved in transcription regulation, epigenetic modification, and other processes essential for normal gametes formation.

  18. The Role of Histone Ubiquitination during Spermatogenesis

    Directory of Open Access Journals (Sweden)

    Kai Sheng

    2014-01-01

    Full Text Available Protein ubiquitin-proteasome (ubiquitin-proteasome system is the major mechanism responsible for protein degradation in eukaryotic cell. During spermatogenesis, the replacement of histone by protamine is vital for normal sperm formation, which is involved in ubiquitination enzymes expressed in testis. Recently, histone ubiquitin ligases have been shown to play critical roles in several aspects of spermatogenesis, such as meiotic sex chromosome inactivation (MSCI, DNA damage response, and spermiogenesis. In this review, we highlight recent progress in the discovery of several histone ubiquitin ligases and elaborate mechanisms of how these enzymes are involved in these processes through knockout mouse model. Using Huwe1, UBR2, and RNF8 as examples, we emphasized the diverse functions for each enzyme and the broad involvement of these enzymes in every stage, from spermatogonia differentiation and meiotic division to spermiogenesis; thus histone ubiquitin ligases represent a class of enzymes, which play important roles in spermatogenesis through targeting histone for ubiquitination and therefore are involved in transcription regulation, epigenetic modification, and other processes essential for normal gametes formation.

  19. Histones bundle F-actin filaments and affect actin structure.

    Science.gov (United States)

    Blotnick, Edna; Sol, Asaf; Muhlrad, Andras

    2017-01-01

    Histones are small polycationic proteins complexed with DNA located in the cell nucleus. Upon apoptosis they are secreted from the cells and react with extracellular polyanionic compounds. Actin which is a polyanionic protein, is also secreted from necrotic cells and interacts with histones. We showed that both histone mixture (histone type III) and the recombinant H2A histone bundles F-actin, increases the viscosity of the F-actin containing solution and polymerizes G-actin. The histone-actin bundles are relatively insensitive to increase of ionic strength, unlike other polycation, histatin, lysozyme, spermine and LL-37 induced F-actin bundles. The histone-actin bundles dissociate completely only in the presence of 300-400 mM NaCl. DNA, which competes with F-actin for histones, disassembles histone induced actin bundles. DNase1, which depolymerizes F- to G-actin, actively unbundles the H2A histone induced but slightly affects the histone mixture induced actin bundles. Cofilin decreases the amount of F-actin sedimented by low speed centrifugation, increases light scattering and viscosity of F-actin-histone mixture containing solutions and forms star like superstructures by copolymerizing G-actin with H2A histone. The results indicate that histones are tightly attached to F-actin by strong electrostatic and hydrophobic forces. Since both histones and F-actin are present in the sputum of patients with cystic fibrosis, therefore, the formation of the stable histone-actin bundles can contribute to the pathology of this disease by increasing the viscosity of the sputum. The actin-histone interaction in the nucleus might affect gene expression.

  20. Small molecule inhibitors of histone deacetylases and acetyltransferases as potential therapeutics in oncology

    NARCIS (Netherlands)

    van den Bosch, Teatske; Leus, Niek; Timmerman, Tirza; Dekker, Frans

    2016-01-01

    Uncontrolled cell proliferation and resistance to apoptosis in cancer are, among others, regulated by post-translational modifications of histone proteins. The most investigated type of histone modification is lysine acetylation. Histone acetyltransferases (HATs), acetylate histone lysine residues,

  1. Acetylation of retinal histones in diabetes increases inflammatory proteins: effects of minocycline and manipulation of histone acetyltransferase (HAT) and histone deacetylase (HDAC).

    Science.gov (United States)

    Kadiyala, Chandra Sekhar Rao; Zheng, Ling; Du, Yunpeng; Yohannes, Elizabeth; Kao, Hung-Ying; Miyagi, Masaru; Kern, Timothy S

    2012-07-27

    Histone acetylation was significantly increased in retinas from diabetic rats, and this acetylation was inhibited in diabetics treated with minocycline, a drug known to inhibit early diabetic retinopathy in animals. Histone acetylation and expression of inflammatory proteins that have been implicated in the pathogenesis of diabetic retinopathy were increased likewise in cultured retinal Müller glia grown in a diabetes-like concentration of glucose. Both the acetylation and induction of the inflammatory proteins in elevated glucose levels were significantly inhibited by inhibitors of histone acetyltransferase (garcinol and antisense against the histone acetylase, p300) or activators of histone deacetylase (theophylline and resveratrol) and were increased by the histone deacetylase inhibitor, suberolylanilide hydroxamic acid. We conclude that hyperglycemia causes acetylation of retinal histones (and probably other proteins) and that the acetylation contributes to the hyperglycemia-induced up-regulation of proinflammatory proteins and thereby to the development of diabetic retinopathy.

  2. Histone H1 phosphorylation occurs site-specifically during interphase and mitosis: identification of a novel phosphorylation site on histone H1.

    Science.gov (United States)

    Sarg, Bettina; Helliger, Wilfried; Talasz, Heribert; Förg, Barbara; Lindner, Herbert H

    2006-03-10

    H1 histones, isolated from logarithmically growing and mitotically enriched human lymphoblastic T-cells (CCRF-CEM), were fractionated by reversed phase and hydrophilic interaction liquid chromatography, subjected to enzymatic digestion, and analyzed by amino acid sequencing and mass spectrometry. During interphase the four H1 subtypes present in these cells differ in their maximum phosphorylation levels: histone H1.5 is tri-, H1.4 di-, and H1.3 and H1.2, only monophosphorylated. The phosphorylation is site-specific and occurs exclusively on serine residues of SP(K/A)K motifs. The phosphorylation sites of histone H1.5 from mitotically enriched cells were also examined. In contrast to the situation in interphase, at mitosis there were additional phosphorylations, exclusively at threonine residues. Whereas the tetraphosphorylated H1.5 arises from the triphosphosphorylated form by phosphorylation of one of two TPKK motifs in the C-terminal domain, namely Thr137 and Thr154, the pentaphosphorylated H1.5 was the result of phosphorylation of one of the tetraphosphorylated forms at a novel nonconsensus motif at Thr10 in the N-terminal tail. Despite the fact that histone H1.5 has five (S/T)P(K/A)K motifs, all of these motifs were never found to be phosphorylated simultaneously. Our data suggest that phosphorylation of human H1 variants occurs nonrandomly during both interphase and mitosis and that distinct serine- or threonine-specific kinases are involved in different cell cycle phases. The order of increased phosphorylation and the position of modification might be necessary for regulated chromatin decondensation, thus facilitating processes of replication and transcription as well as of mitotic chromosome condensation.

  3. Antifungal properties of wheat histones (H1-H4) and purified wheat histone H1.

    Science.gov (United States)

    De Lucca, Anthony J; Heden, Lars-Olof; Ingber, Bruce; Bhatnagar, Deepak

    2011-07-13

    Wheat ( Triticum spp.) histones H1, H2, H3, and H4 were extracted, and H1 was further purified. The effect of these histones on specific fungi that may or may not be pathogenic to wheat was determined. These fungi included Aspergillus flavus , Aspergillus fumigatus , Aspergillus niger , Fusarium oxysporum , Fusarium verticillioides , Fusarium solani , Fusarium graminearum , Penicillium digitatum , Penicillium italicum , and Greeneria uvicola . Non-germinated and germinating conidia of these fungi were bioassayed separately. The non-germinated and germinating conidia of all Fusarium species were highly susceptible to the mixture (H1-H4) as well as pure H1, with viability losses of 99-100% found to be significant (p histone mixture and pure H1. F. graminearum was the most sensitive to histone activity. The histones were inactive against all of the non-germinated Penicillium spp. conidia. However, they significantly reduced the viability of the germinating conidia of the Penicillium spp. conidia, with 95% loss at 2.5 μM. Non-germinated and germinating conidia viability of the Aspergillus spp. and G. uvicola were unaffected when exposed to histones up to 10 μM. Results indicate that Fusarium spp. pathogenic to wheat are susceptible to wheat histones, indicating that these proteins may be a resistance mechanism in wheat against fungal infection.

  4. The FACT histone chaperone guides histone H4 into its nucleosomal conformation in Saccharomyces cerevisiae.

    Science.gov (United States)

    McCullough, Laura; Poe, Bryan; Connell, Zaily; Xin, Hua; Formosa, Tim

    2013-09-01

    The pob3-Q308K mutation alters the small subunit of the Saccharomyces cerevisiae histone/nucleosome chaperone Facilitates Chromatin Transactions (FACT), causing defects in both transcription and DNA replication. We describe histone mutations that suppress some of these defects, providing new insight into the mechanism of FACT activity in vivo. FACT is primarily known for its ability to promote reorganization of nucleosomes into a more open form, but neither the pob3-Q308K mutation nor the compensating histone mutations affect this activity. Instead, purified mutant FACT complexes fail to release from nucleosomes efficiently, and the histone mutations correct this flaw. We confirm that pob3-T252E also suppresses pob3-Q308K and show that combining two suppressor mutations can be detrimental, further demonstrating the importance of balance between association and dissociation for efficient FACT:nucleosome interactions. To explain our results, we propose that histone H4 can adopt multiple conformations, most of which are incompatible with nucleosome assembly. FACT guides H4 to adopt appropriate conformations, and this activity can be enhanced or diminished by mutations in Pob3 or histones. FACT can therefore destabilize nucleosomes by favoring the reorganized state, but it can also promote assembly by tethering histones and DNA together and maintaining them in conformations that promote canonical nucleosome formation.

  5. The emerging functions of histone demethylases

    DEFF Research Database (Denmark)

    Agger, Karl; Christensen, Jesper; Cloos, Paul Ac;

    2008-01-01

    Epigenetic information refers to heritable changes in gene function that are stable between cell divisions but which is not a result of changes in the DNA sequence. Part of the epigenetic mechanism has been ascribed to modifications of histones or DNA that affects the transcription of specific...... genes. In this context, post-translational modifications of histone tails, in particular methylation of lysines, are regarded as important for the storage of epigenetic information. Regulation of this information plays an important role during cellular differentiation where cells with different...... characteristic features evolve from the same ancestor, despite identical genomic material. The characterization of several enzymes catalyzing histone lysine methylation have supported this concept by showing the requirement of these enzymes for normal development and their involvement in diseases such as cancer...

  6. [Structure and function of histone chaperone FACT].

    Science.gov (United States)

    Bondarenko, M T; Maluchenko, N V; Valieva, M E; Gerasimova, N S; Kulaeva, O I; Georgiev, P G; Studitsky, V M

    2015-01-01

    FACT is heterodimer protein complex and histone chaperone that plays an important role in maintaining and modifying chromatin structure during various DNA-dependent processes. FACT is involved in nucleosome assembly de novo and in the preservation and recovery of the nucleosome structure during and after transcription, replication and repair of DNA. During transcript elongation FACT reduces the height of the nucleosome barrier and supports survival of the nucleosomes during and after passage of RNA polymerase II. In this process FACT interacts with histone H2A-H2B dimer in nucleosomes, thus facilitating uncoiling of nucleosomal DNA from the octamer of histones; it also facilitates subsequent recovery of the canonical structure of the nucleosome after transcription. FACT also plays an important role in transformation of human cells and in maintaining the viability of the tumor cells.

  7. H1 histones: current perspectives and challenges.

    Science.gov (United States)

    Harshman, Sean W; Young, Nicolas L; Parthun, Mark R; Freitas, Michael A

    2013-11-01

    H1 and related linker histones are important both for maintenance of higher-order chromatin structure and for the regulation of gene expression. The biology of the linker histones is complex, as they are evolutionarily variable, exist in multiple isoforms and undergo a large variety of posttranslational modifications in their long, unstructured, NH2- and COOH-terminal tails. We review recent progress in understanding the structure, genetics and posttranslational modifications of linker histones, with an emphasis on the dynamic interactions of these proteins with DNA and transcriptional regulators. We also discuss various experimental challenges to the study of H1 and related proteins, including limitations of immunological reagents and practical difficulties in the analysis of posttranslational modifications by mass spectrometry.

  8. Linker histones: History and current perspectives.

    Science.gov (United States)

    Crane-Robinson, C

    2016-03-01

    Although the overall structure of the fifth histone (linker histone, H1) is understood, its location on the nucleosome is only partially defined. Whilst it is clear that H1 helps condense the chromatin fibre, precisely how this is achieved remains to be determined. H1 is not a general gene repressor in that although it must be displaced from transcription start sites for activity to occur, there is only partial loss along the body of genes. How the deposition and removal of H1 occurs in particular need of further study. Linker histones are highly abundant nuclear proteins about which we know too little. Copyright © 2015. Published by Elsevier B.V.

  9. Chemical and semisynthesis of modified histones.

    Science.gov (United States)

    Maity, Suman Kumar; Jbara, Muhammad; Brik, Ashraf

    2016-05-01

    Post-translational modifications (PTMs) of histones play critical roles in the epigenetic regulation of eukaryotic genome by directly altering the biophysical properties of chromatin or by recruiting effector proteins. The large number of PTMs and the inherent complexity in their population and signaling processes make it highly challenging to understand epigenetics-related processes. To address these challenges, accesses to homogeneously modified histones are obligatory. Over the last decade, synthetic protein chemists have been devising novel synthetic tools and applying state-of-the-art chemoselective ligation strategies to prepare precious materials useful in answering fundamental questions in this area. In this short review, we cover some of the recent breakthroughs in these directions in particular the synthesis and semi-synthesis of modified histones and their use to unravel the mysteries of epigenetics. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  10. Proteomics Core

    Data.gov (United States)

    Federal Laboratory Consortium — Proteomics Core is the central resource for mass spectrometry based proteomics within the NHLBI. The Core staff help collaborators design proteomics experiments in a...

  11. Proteomics Core

    Data.gov (United States)

    Federal Laboratory Consortium — Proteomics Core is the central resource for mass spectrometry based proteomics within the NHLBI. The Core staff help collaborators design proteomics experiments in...

  12. Structure of histone-based chromatin in archaea

    National Research Council Canada - National Science Library

    Mattiroli, Francesca; Bhattacharyya, Sudipta; Dyer, Pamela N; White, Alison E; Sandman, Kathleen; Burkhart, Brett W; Byrne, Kyle R; Lee, Thomas; Aim, Natalie G; Santangelo, Thomas J; Reeve, John N; Luger, Karolin

    2017-01-01

    .... We report the crystal structure of an archaeal histone-DNA complex. DNA wraps around an extended polymer, formed by archaeal histone homodimers, in a quasi-continuous superhelix with the same geometry as DNA in the eukaryotic nucleosome...

  13. Histone H4 Lysine 20 methylation

    DEFF Research Database (Denmark)

    Jørgensen, Stine; Schotta, Gunnar; Sørensen, Claus Storgaard

    2013-01-01

    of histones have emerged as key regulators of genomic integrity. Intense research during the past few years has revealed histone H4 lysine 20 methylation (H4K20me) as critically important for the biological processes that ensure genome integrity, such as DNA damage repair, DNA replication and chromatin...... instability, demonstrating the important functions of H4K20 methylation in genome maintenance. In this review, we explain molecular mechanisms underlying these defects and discuss novel ideas for furthering our understanding of genome maintenance in higher eukaryotes....

  14. Acetylation of histone H3 at lysine 64 regulates nucleosome dynamics and facilitates transcription

    Science.gov (United States)

    Di Cerbo, Vincenzo; Mohn, Fabio; Ryan, Daniel P; Montellier, Emilie; Kacem, Salim; Tropberger, Philipp; Kallis, Eleni; Holzner, Monika; Hoerner, Leslie; Feldmann, Angelika; Richter, Florian Martin; Bannister, Andrew J; Mittler, Gerhard; Michaelis, Jens; Khochbin, Saadi; Feil, Robert; Schuebeler, Dirk; Owen-Hughes, Tom; Daujat, Sylvain; Schneider, Robert

    2014-01-01

    Post-translational modifications of proteins have emerged as a major mechanism for regulating gene expression. However, our understanding of how histone modifications directly affect chromatin function remains limited. In this study, we investigate acetylation of histone H3 at lysine 64 (H3K64ac), a previously uncharacterized acetylation on the lateral surface of the histone octamer. We show that H3K64ac regulates nucleosome stability and facilitates nucleosome eviction and hence gene expression in vivo. In line with this, we demonstrate that H3K64ac is enriched in vivo at the transcriptional start sites of active genes and it defines transcriptionally active chromatin. Moreover, we find that the p300 co-activator acetylates H3K64, and consistent with a transcriptional activation function, H3K64ac opposes its repressive counterpart H3K64me3. Our findings reveal an important role for a histone modification within the nucleosome core as a regulator of chromatin function and they demonstrate that lateral surface modifications can define functionally opposing chromatin states. DOI: http://dx.doi.org/10.7554/eLife.01632.001 PMID:24668167

  15. An update on histone lysine methylation in plants

    Institute of Scientific and Technical Information of China (English)

    Yu Yu; Zhongyuan Bu; Wen-Hui Shen; Aiwu Dong

    2009-01-01

    Histone methylation plays crucial roles in epigenetic regulation.The SET domain proteins are now recognized as generally having methyltransferase activity targeted to specific lysine residues of histones.The enzymes and their specific histone lysine methylation have enormous impacts on the regulation of chromatin structure and function.In this review,we discuss recent advances made on histone lysine methylations and their diverse functions in plant growth and development.

  16. Histones as mediators of host defense, inflammation and thrombosis

    NARCIS (Netherlands)

    Hoeksema, Marloes; Eijk, Martin van; Haagsman, Henk P; Hartshorn, Kevan L

    2016-01-01

    Histones are known for their ability to bind to and regulate expression of DNA. However, histones are also present in cytoplasm and extracellular fluids where they serve host defense functions and promote inflammatory responses. Histones are a major component of neutrophil extracellular traps that c

  17. Influence of chromatin structure, antibiotics, and endogenous histone methylation on phosphorylation of histones H1 and H3 in the presence of protein kinase A in rat liver nuclei in vitro.

    Science.gov (United States)

    Prusov, A N; Smirnova, T A; Kolomijtseva, G Ya

    2013-02-01

    In vitro phosphorylation of histones H1 and H3 by cAMP-dependent protein kinase A and endogenous phosphokinases in the presence of [γ-³²P]ATP was studied in isolated rat liver nuclei with different variants of chromatin structural organization: condensed (diameter of fibrils 100-200 nm; N-1) and partly decondensed (diameter of fibrils ~30 nm; N-2). In the N-1 state histone, H1 is phosphorylated approximately twice as much than histone H3. Upon the decondensation of the chromatin in the N-2 state, 1.5-fold decrease of total phosphorylation of H1 is observed, while that of H3 does not change, although the endogenous phosphorylation of both histones is reduced by half. Changes in histone phosphorylation in the presence of low or high concentrations of distamycin and chromomycin differ for H1 and H3 in N-1 and N-2. It was found that distamycin (DM) stimulates the phosphorylation of tightly bound H1 fraction, which is not extractable by polyglutamic acid (PG), especially in N-1. Chromomycin (CM) increases the phosphorylation of both histones in PG extracts and in the nuclear pellets, particularly in N-2. At the same time, in N-1 one can detect phosphorylation of a tightly bound fraction of histones H1 whose N-termini are located on AT-rich sites that become inaccessible for protein kinase in the process of chromatin decondensation in N-2. At the same time, in N-2 the accessibility for protein kinase A of tightly bound H1 fractions, whose N-termini are located on GC-rich sites, increases dramatically. High concentrations of both CM and DM in N-1 and N-2 stimulated phosphorylation of the non-extractable by PG fraction of H1 whose N-termini are located on sites where AT ≈ GC. CM at high concentration stimulated 4-7 times the phosphorylation of a small fraction of H3, which is extracted by PG from both types of nuclei. We detected an effect of endogenous methylation of histones H1 and H3 in the nuclei on their subsequent phosphorylation depending on the chromatin

  18. Inhibitors of DNA Methylation, Histone Deacetylation, and Histone Demethylation: A Perfect Combination for Cancer Therapy.

    Science.gov (United States)

    Zahnow, C A; Topper, M; Stone, M; Murray-Stewart, T; Li, H; Baylin, S B; Casero, R A

    2016-01-01

    Epigenetic silencing and inappropriate activation of gene expression are frequent events during the initiation and progression of cancer. These events involve a complex interplay between the hypermethylation of CpG dinucleotides within gene promoter and enhancer regions, the recruitment of transcriptional corepressors and the deacetylation and/or methylation of histone tails. These epigenetic regulators act in concert to block transcription or interfere with the maintenance of chromatin boundary regions. However, DNA/histone methylation and histone acetylation states are reversible, enzyme-mediated processes and as such, have emerged as promising targets for cancer therapy. This review will focus on the potential benefits and synergistic/additive effects of combining DNA-demethylating agents and histone deacetylase inhibitors or lysine-specific demethylase inhibitors together in epigenetic therapy for solid tumors and will highlight what is known regarding the mechanisms of action that contribute to the antitumor response.

  19. dKDM2 couples histone H2A ubiquitylation to histone H3 demethylation during Polycomb group silencing

    NARCIS (Netherlands)

    A. Lagarou (Anna); A.B. Mohd Sarip; Y.M. Moshkin (Yuri); G.E. Chalkley (Gillian); K. Bezstarosti (Karel); J.A.A. Demmers (Jeroen); C.P. Verrijzer (Peter)

    2008-01-01

    textabstractTranscription regulation involves enzyme-mediated changes in chromatin structure. Here, we describe a novel mode of histone crosstalk during gene silencing, in which histone H2A monoubiquitylation is coupled to the removal of histone H3 Lys 36 dimethylation (H3K36me2). This pathway was u

  20. Trypanosomal histone γH2A and the DNA damage response.

    Science.gov (United States)

    Glover, Lucy; Horn, David

    2012-05-01

    DNA damage and repair in trypanosomatids impacts virulence, drug resistance and antigenic variation but, currently, little is known about DNA damage responses or cell cycle checkpoints in these divergent protozoa. One of the earliest markers of DNA damage in eukaryotes is γH2A(X), a serine phosphorylated histone H2A (variant). Here, we report the identification and initial characterization of γH2A in Trypanosoma brucei. We identified Thr(130) within the replication-dependent histone H2A as a candidate phosphorylation site and found that the abundance of this trypanosomal γH2A increased in vivo in response to DNA damage. Nuclear γH2A foci mark the sites of putative natural replication fork stalling, sites of meganuclease-induced DNA double strand breaks and sites of methyl methanesulphonate-induced DNA damage. Naturally occurring and meganuclease-induced γH2A and RAD51 double-positive repair foci are typically found in S-phase or G(2) nuclei. The results link trypanosomal γH2A, with an unusual histone modification motif, to DNA damage sensing and mitotic checkpoint signaling.

  1. FACT plays a major role in histone dynamics affecting VSG expression site control in Trypanosoma brucei.

    Science.gov (United States)

    Denninger, Viola; Rudenko, Gloria

    2014-11-01

    Chromatin remodelling is involved in the transcriptional regulation of the RNA polymerase I transcribed variant surface glycoprotein (VSG) expression sites (ESs) of Trypanosoma brucei. We show that the T. brucei FACT complex contains the Pob3 and Spt16 subunits, and plays a key role in ES silencing. We see an inverse correlation between transcription and condensed chromatin, whereby FACT knockdown results in ES derepression and more open chromatin around silent ES promoters. Derepressed ESs show increased sensitivity to micrococcal nuclease (MNase) digestion, and a decrease in histones at silent ES promoters but not telomeres. In contrast, FACT knockdown results in more histones at the active ES, correlated with transcription shut-down. ES promoters are derepressed in cells stalled at the G2/M cell cycle stage after knockdown of FACT, but not in G2/M cells stalled after knockdown of cyclin 6. This argues that the observed ES derepression is a direct consequence of histone chaperone activity by FACT at the G2/M cell cycle stage which could affect transcription elongation, rather than an indirect consequence of a cell cycle checkpoint. These experiments highlight the role of the FACT complex in cell cycle-specific chromatin remodelling within VSG ESs. © 2014 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.

  2. Solution NMR structure and histone binding of the PHD domain of human MLL5.

    Directory of Open Access Journals (Sweden)

    Alexander Lemak

    Full Text Available Mixed Lineage Leukemia 5 (MLL5 is a histone methyltransferase that plays a key role in hematopoiesis, spermatogenesis and cell cycle progression. In addition to its catalytic domain, MLL5 contains a PHD finger domain, a protein module that is often involved in binding to the N-terminus of histone H3. Here we report the NMR solution structure of the MLL5 PHD domain showing a variant of the canonical PHD fold that combines conserved H3 binding features from several classes of other PHD domains (including an aromatic cage along with a novel C-terminal α-helix, not previously seen. We further demonstrate that the PHD domain binds with similar affinity to histone H3 tail peptides di- and tri-methylated at lysine 4 (H3K4me2 and H3K4me3, the former being the putative product of the MLL5 catalytic reaction. This work establishes the PHD domain of MLL5 as a bone fide 'reader' domain of H3K4 methyl marks suggesting that it may guide the spreading or further methylation of this site on chromatin.

  3. Highly selective inhibition of histone demethylases by de novo macrocyclic peptides.

    Science.gov (United States)

    Kawamura, Akane; Münzel, Martin; Kojima, Tatsuya; Yapp, Clarence; Bhushan, Bhaskar; Goto, Yuki; Tumber, Anthony; Katoh, Takayuki; King, Oliver N F; Passioura, Toby; Walport, Louise J; Hatch, Stephanie B; Madden, Sarah; Müller, Susanne; Brennan, Paul E; Chowdhury, Rasheduzzaman; Hopkinson, Richard J; Suga, Hiroaki; Schofield, Christopher J

    2017-04-06

    The JmjC histone demethylases (KDMs) are linked to tumour cell proliferation and are current cancer targets; however, very few highly selective inhibitors for these are available. Here we report cyclic peptide inhibitors of the KDM4A-C with selectivity over other KDMs/2OG oxygenases, including closely related KDM4D/E isoforms. Crystal structures and biochemical analyses of one of the inhibitors (CP2) with KDM4A reveals that CP2 binds differently to, but competes with, histone substrates in the active site. Substitution of the active site binding arginine of CP2 to N-ɛ-trimethyl-lysine or methylated arginine results in cyclic peptide substrates, indicating that KDM4s may act on non-histone substrates. Targeted modifications to CP2 based on crystallographic and mass spectrometry analyses results in variants with greater proteolytic robustness. Peptide dosing in cells manifests KDM4A target stabilization. Although further development is required to optimize cellular activity, the results reveal the feasibility of highly selective non-metal chelating, substrate-competitive inhibitors of the JmjC KDMs.

  4. Mutation Update: The Spectra of Nebulin Variants and Associated Myopathies

    Science.gov (United States)

    Lehtokari, Vilma-Lotta; Kiiski, Kirsi; Sandaradura, Sarah A.; Laporte, Jocelyn; Repo, Pauliina; Frey, Jennifer A.; Donner, Kati; Marttila, Minttu; Saunders, Carol; Barth, Peter G.; den Dunnen, Johan T.; Beggs, Alan H.; Clarke, Nigel F.; North, Kathryn N.; Laing, Nigel G.; Romero, Norma B.; Winder, Thomas L.; Pelin, Katarina; Wallgren-Pettersson, Carina

    2015-01-01

    A mutation update on the nebulin gene (NEB) is necessary because of recent developments in analysis methodology, the identification of increasing numbers and novel types of variants, and a widening in the spectrum of clinical and histological phenotypes associated with this gigantic, 183 exons containing gene. Recessive pathogenic variants in NEB are the major cause of nemaline myopathy (NM), one of the most common congenital myopathies. Moreover, pathogenic NEB variants have been identified in core-rod myopathy and in distal myopathies. In this update, we present the disease-causing variants in NEB in 159 families, 143 families with NM, and 16 families with NM-related myopathies. Eighty-eight families are presented here for the first time. We summarize 86 previously published and 126 unpublished variants identified in NEB. Furthermore, we have analyzed the NEB variants deposited in the Exome Variant Server (http://evs.gs.washington.edu/EVS/), identifying that pathogenic variants are a minor fraction of all coding variants (~7%). This indicates that nebulin tolerates substantial changes in its amino acid sequence, providing an explanation as to why variants in such a large gene result in relatively rare disorders. Lastly, we discuss the difficulties of drawing reliable genotype–phenotype correlations in NEB-associated disease. PMID:25205138

  5. An all-atom model of the chromatin fiber containing linker histones reveals a versatile structure tuned by the nucleosomal repeat length.

    Directory of Open Access Journals (Sweden)

    Hua Wong

    Full Text Available In the nucleus of eukaryotic cells, histone proteins organize the linear genome into a functional and hierarchical architecture. In this paper, we use the crystal structures of the nucleosome core particle, B-DNA and the globular domain of H5 linker histone to build the first all-atom model of compact chromatin fibers. In this 3D jigsaw puzzle, DNA bending is achieved by solving an inverse kinematics problem. Our model is based on recent electron microscopy measurements of reconstituted fiber dimensions. Strikingly, we find that the chromatin fiber containing linker histones is a polymorphic structure. We show that different fiber conformations are obtained by tuning the linker histone orientation at the nucleosomes entry/exit according to the nucleosomal repeat length. We propose that the observed in vivo quantization of nucleosomal repeat length could reflect nature's ability to use the DNA molecule's helical geometry in order to give chromatin versatile topological and mechanical properties.

  6. Regulation of replication fork progression through histone supply and demand

    DEFF Research Database (Denmark)

    Groth, Anja; Corpet, Armelle; Cook, Adam J L

    2007-01-01

    DNA replication in eukaryotes requires nucleosome disruption ahead of the replication fork and reassembly behind. An unresolved issue concerns how histone dynamics are coordinated with fork progression to maintain chromosomal stability. Here, we characterize a complex in which the human histone...... chaperone Asf1 and MCM2-7, the putative replicative helicase, are connected through a histone H3-H4 bridge. Depletion of Asf1 by RNA interference impedes DNA unwinding at replication sites, and similar defects arise from overproduction of new histone H3-H4 that compromises Asf1 function. These data link Asf......1 chaperone function, histone supply, and replicative unwinding of DNA in chromatin. We propose that Asf1, as a histone acceptor and donor, handles parental and new histones at the replication fork via an Asf1-(H3-H4)-MCM2-7 intermediate and thus provides a means to fine-tune replication fork...

  7. The histone H3 N-terminal tail: a computational analysis of the free energy landscape and kinetics.

    Science.gov (United States)

    Zheng, Yuqing; Cui, Qiang

    2015-05-28

    Histone tails are the short peptide protrusions outside of the nucleosome core particle and they play a critical role in regulating chromatin dynamics and gene activity. A histone H3 N-terminal tail, like other histone tails, can be covalently modified on different residues to activate or repress gene expression. Previous studies have indicated that, despite its intrinsically disordered nature, the histone H3 N-terminal tail has regions of notable secondary structural propensities. To further understand the structure-dynamics-function relationship in this system, we have carried out 75.6 μs long implicit solvent simulations and 29.3 μs long explicit solvent simulations. The extensive samplings allow us to better characterize not only the underlying free energy landscape but also kinetic properties through Markov state models (MSM). Dihedral principal component analysis (dPCA) and locally scaled diffusion map (LSDMap) analysis yield consistent results that indicate an overall flat free energy surface with several shallow basins that correspond to conformations with a high α-helical propensity in two regions of the peptide. Kinetic information extracted from Markov state models reveals rapid transitions between different metastable states with mean first passage times spanning from several hundreds of nanoseconds to hundreds of microseconds. These findings shed light on how the dynamical nature of the histone H3 N-terminal tail is related to its function. The complementary nature of dPCA, LSDMap and MSM for the analysis of biomolecules is also discussed.

  8. A role for nucleosome assembly protein 1 in the nuclear transport of histones H2A and H2B.

    Science.gov (United States)

    Mosammaparast, Nima; Ewart, Courtney S; Pemberton, Lucy F

    2002-12-02

    Import of core histones into the nucleus is a prerequisite for their deposition onto DNA and the assembly of chromatin. Here we demonstrate that nucleosome assembly protein 1 (Nap1p), a protein previously implicated in the deposition of histones H2A and H2B, is also involved in the transport of these two histones. We demonstrate that Nap1p can bind directly to Kap114p, the primary karyopherin/importin responsible for the nuclear import of H2A and H2B. Nap1p also serves as a bridge between Kap114p and the histone nuclear localization sequence (NLS). Nap1p acts cooperatively to increase the affinity of Kap114p for these NLSs. Nuclear accumulation of histone NLS-green fluorescent protein (GFP) reporters was decreased in deltanap1 cells. Furthermore, we demonstrate that Nap1p promotes the association of the H2A and H2B NLSs specifically with the karyopherin Kap114p. Localization studies demonstrate that Nap1p is a nucleocytoplasmic shuttling protein, and genetic experiments suggest that its shuttling is important for maintaining chromatin structure in vivo. We propose a model in which Nap1p links the nuclear transport of H2A and H2B to chromatin assembly.

  9. Med5(Nut1 and Med17(Srb4 are direct targets of mediator histone H4 tail interactions.

    Directory of Open Access Journals (Sweden)

    Zhongle Liu

    Full Text Available The Mediator complex transmits activation signals from DNA bound transcription factors to the core transcription machinery. In addition to its canonical role in transcriptional activation, recent studies have demonstrated that S. cerevisiae Mediator can interact directly with nucleosomes, and their histone tails. Mutations in Mediator subunits have shown that Mediator and certain chromatin structures mutually impact each other structurally and functionally in vivo. We have taken a UV photo cross-linking approach to further delineate the molecular basis of Mediator chromatin interactions and help determine whether the impact of certain Mediator mutants on chromatin is direct. Specifically, by using histone tail peptides substituted with an amino acid analog that is a UV activatible crosslinker, we have identified specific subunits within Mediator that participate in histone tail interactions. Using Mediator purified from mutant yeast strains we have evaluated the impact of these subunits on histone tail binding. This analysis has identified the Med5 subunit of Mediator as a target for histone tail interactions and suggests that the previously observed effect of med5 mutations on telomeric heterochromatin and silencing is direct.

  10. Histone H1 couples initiation and amplification of ubiquitin signalling after DNA damage

    DEFF Research Database (Denmark)

    Thorslund, Tina; Ripplinger, Anita; Hoffmann, Saskia;

    2015-01-01

    DNA double-strand breaks (DSBs) are highly cytotoxic DNA lesions that trigger non-proteolytic ubiquitylation of adjacent chromatin areas to generate binding sites for DNA repair factors. This depends on the sequential actions of the E3 ubiquitin ligases RNF8 and RNF168 (refs 1-6), and UBC13 (also...... ubiquitylation remain elusive. Here we elucidate how RNF8 and UBC13 promote recruitment of RNF168 and downstream factors to DSB sites in human cells. We establish that UBC13-dependent K63-linked ubiquitylation at DSB sites is predominantly mediated by RNF8 but not RNF168, and that H1-type linker histones......, but not core histones, represent major chromatin-associated targets of this modification. The RNF168 module (UDM1) recognizing RNF8-generated ubiquitylations is a high-affinity reader of K63-ubiquitylated H1, mechanistically explaining the essential roles of RNF8 and UBC13 in recruiting RNF168 to DSBs...

  11. Structure of the histone chaperone CIA/ASF1-double bromodomain complex linking histone modifications and site-specific histone eviction.

    Science.gov (United States)

    Akai, Yusuke; Adachi, Naruhiko; Hayashi, Yohei; Eitoku, Masamitsu; Sano, Norihiko; Natsume, Ryo; Kudo, Norio; Tanokura, Masaru; Senda, Toshiya; Horikoshi, Masami

    2010-05-04

    Nucleosomes around the promoter region are disassembled for transcription in response to various signals, such as acetylation and methylation of histones. Although the interactions between histone-acetylation-recognizing bromodomains and factors involved in nucleosome disassembly have been reported, no structural basis connecting histone modifications and nucleosome disassembly has been obtained. Here, we determined at 3.3 A resolution the crystal structure of histone chaperone cell cycle gene 1 (CCG1) interacting factor A/antisilencing function 1 (CIA/ASF1) in complex with the double bromodomain in the CCG1/TAF1/TAF(II)250 subunit of transcription factor IID. Structural, biochemical, and biological studies suggested that interaction between double bromodomain and CIA/ASF1 is required for their colocalization, histone eviction, and pol II entry at active promoter regions. Furthermore, the present crystal structure has characteristics that can connect histone acetylation and CIA/ASF1-mediated histone eviction. These findings suggest that the molecular complex between CIA/ASF1 and the double bromodomain plays a key role in site-specific histone eviction at active promoter regions. The model we propose here is the initial structure-based model of the biological signaling from histone modifications to structural change of the nucleosome (hi-MOST model).

  12. Condensed mitotic chromosome structure at nanometer resolution using PALM and EGFP- histones.

    Science.gov (United States)

    Matsuda, Atsushi; Shao, Lin; Boulanger, Jerome; Kervrann, Charles; Carlton, Peter M; Kner, Peter; Agard, David; Sedat, John W

    2010-09-15

    Photoactivated localization microscopy (PALM) and related fluorescent biological imaging methods are capable of providing very high spatial resolutions (up to 20 nm). Two major demands limit its widespread use on biological samples: requirements for photoactivatable/photoconvertible fluorescent molecules, which are sometimes difficult to incorporate, and high background signals from autofluorescence or fluorophores in adjacent focal planes in three-dimensional imaging which reduces PALM resolution significantly. We present here a high-resolution PALM method utilizing conventional EGFP as the photoconvertible fluorophore, improved algorithms to deal with high levels of biological background noise, and apply this to imaging higher order chromatin structure. We found that the emission wavelength of EGFP is efficiently converted from green to red when exposed to blue light in the presence of reduced riboflavin. The photon yield of red-converted EGFP using riboflavin is comparable to other bright photoconvertible fluorescent proteins that allow histone H2A variant, revealed filamentous components of ∼70 nm. This is the first observation of fine chromatin filaments specific for one histone variant at a resolution approximating that of conventional electron microscope images (10-30 nm). As demonstrated by modeling and experiments on a challenging specimen, the techniques described here facilitate super-resolution fluorescent imaging with common biological samples.

  13. The rapidly evolving centromere-specific histone has stringent functional requirements in Arabidopsis thaliana.

    Science.gov (United States)

    Ravi, Maruthachalam; Kwong, Pak N; Menorca, Ron M G; Valencia, Joel T; Ramahi, Joseph S; Stewart, Jodi L; Tran, Robert K; Sundaresan, Venkatesan; Comai, Luca; Chan, Simon W-L

    2010-10-01

    Centromeres control chromosome inheritance in eukaryotes, yet their DNA structure and primary sequence are hypervariable. Most animals and plants have megabases of tandem repeats at their centromeres, unlike yeast with unique centromere sequences. Centromere function requires the centromere-specific histone CENH3 (CENP-A in human), which replaces histone H3 in centromeric nucleosomes. CENH3 evolves rapidly, particularly in its N-terminal tail domain. A portion of the CENH3 histone-fold domain, the CENP-A targeting domain (CATD), has been previously shown to confer kinetochore localization and centromere function when swapped into human H3. Furthermore, CENP-A in human cells can be functionally replaced by CENH3 from distantly related organisms including Saccharomyces cerevisiae. We have used cenh3-1 (a null mutant in Arabidopsis thaliana) to replace endogenous CENH3 with GFP-tagged variants. A H3.3 tail domain-CENH3 histone-fold domain chimera rescued viability of cenh3-1, but CENH3's lacking a tail domain were nonfunctional. In contrast to human results, H3 containing the A. thaliana CATD cannot complement cenh3-1. GFP-CENH3 from the sister species A. arenosa functionally replaces A. thaliana CENH3. GFP-CENH3 from the close relative Brassica rapa was targeted to centromeres, but did not complement cenh3-1, indicating that kinetochore localization and centromere function can be uncoupled. We conclude that CENH3 function in A. thaliana, an organism with large tandem repeat centromeres, has stringent requirements for functional complementation in mitosis.

  14. Dynamic Regulation of Histone Modifications in Xenopus Oocytes through Histone Exchange

    Science.gov (United States)

    Stewart, M. David; Sommerville, John; Wong, Jiemin

    2006-01-01

    Histone H3 lysine 9 (H3K9) methylation has broad roles in transcriptional repression, gene silencing, maintenance of heterochromatin, and epigenetic inheritance of heterochromatin. Using Xenopus laevis oocytes, we have previously shown that targeting G9a, an H3K9 histone methyltransferase, to chromatin increases H3K9 methylation and consequently represses transcription. Here we report that treatment with trichostatin A induces histone acetylation and is sufficient to activate transcription repressed by G9a, and this activation is accompanied by a reduction in dimethyl H3K9 (H3K9me2). We tested the possibility that the reduction in H3K9me2 was due to the replacement of methylated H3 with unmethylated H3.3. Surprisingly, we found that both free H3 and H3.3 are continually exchanged with chromatin-associated histones. This dynamic exchange of chromatin-associated H3 with free H3/H3.3 was not affected by alterations in transcriptional activity, elongation, acetylation, H3K9 methylation, or DNA replication. In support of this continual histone exchange model, we show that maintenance of H3K9 methylation at a specific site requires the continual presence of an H3K9 histone methyltransferase. Upon dissociation of the methyltransferase, H3K9 methylation decreases. Taken together, our data suggest that chromatin-associated and non-chromatin-associated histones are continually exchanged in the Xenopus oocyte, creating a highly dynamic chromatin environment. PMID:16943430

  15. Ice cores

    DEFF Research Database (Denmark)

    Svensson, Anders

    2014-01-01

    Ice cores from Antarctica, from Greenland, and from a number of smaller glaciers around the world yield a wealth of information on past climates and environments. Ice cores offer unique records on past temperatures, atmospheric composition (including greenhouse gases), volcanism, solar activity......, dustiness, and biomass burning, among others. In Antarctica, ice cores extend back more than 800,000 years before present (Jouzel et al. 2007), whereas. Greenland ice cores cover the last 130,000 years...

  16. Ice cores

    DEFF Research Database (Denmark)

    Svensson, Anders

    2014-01-01

    Ice cores from Antarctica, from Greenland, and from a number of smaller glaciers around the world yield a wealth of information on past climates and environments. Ice cores offer unique records on past temperatures, atmospheric composition (including greenhouse gases), volcanism, solar activity......, dustiness, and biomass burning, among others. In Antarctica, ice cores extend back more than 800,000 years before present (Jouzel et al. 2007), whereas. Greenland ice cores cover the last 130,000 years...

  17. Desmoplastic variant of ameloblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Sohn, Jeong Ick; Kim, Dong Youn; Choi, Karp Shik [Dept. of Dental Radiology, College of Dentistry, Kyungpook National University, Daegu (Korea, Republic of)

    1995-02-15

    Desmoplastic variant of ameloblastoma is new and unusual variant of ameloblastoma with extensive stromal desmoplastic proliferation. The authors experienced a case of desmoplastic variant of amleloblastoma with moderate-defined radiolucency on the right maxillary anterior area in 62-year-old female. As a result of careful analysis of clinical, radiological examinations, we diagnosed it as desmoplastic variant of ameloblastoma. The following results were obtained; 1. Main clinical symptoms were nontender bony swelling with normal intact overlying mucosa on the right maxillary anterior area. 2. Radiographically, moderate-defined, multilocular radioluceney on the right maxillary anterior area were shown, and severe cortical bony thinning and expansion to labial and palatal sides were also observed. And this lesion was shown to be extended to the right nasal cavity. 3. Histopathologically, follicle-like epithelial islands with densely abundant collagenous stroma were morphologically compressed.

  18. Structure and function of histone acetyltransferase MOF.

    Science.gov (United States)

    Chen, Qiao Yi; Costa, Max; Sun, Hong

    2015-01-01

    MOF was first identified in Drosophila melanogaster as an important component of the dosage compensation complex. As a member of MYST family of histone acetyltransferase, MOF specifically deposits the acetyl groups to histone H4 lysine 16. Throughout evolution, MOF and its mammalian ortholog have retained highly conserved substrate specificity and similar enzymatic activities. MOF plays important roles in dosage compensation, ESC self-renewal, DNA damage and repair, cell survival, and gene expression regulation. Dysregulation of MOF has been implicated in tumor formation and progression of many types of human cancers. This review will discuss the structure and activity of mammalian hMOF as well as its function in H4K16 acetylation, DNA damage response, stem cell pluripotency, and carcinogenesis.

  19. Human histone acetyltransferase 1 (Hat1) acetylates lysine 5 of histone H2A in vivo.

    Science.gov (United States)

    Tafrova, Juliana I; Tafrov, Stefan T

    2014-07-01

    The primary structure of Histone Acetyltransferase 1 (Hat1) has been conserved throughout evolution; however, despite its ubiquity, its cellular function is not well characterized. To study its in vivo acetylation pattern and function, we utilized shRNAmir against Hat1 expressed in the well-substantiated HeLa (human cervical cancer) cell line. To reduce the interference by enzymes with similar HAT specificity, we used HeLa cells expressing histone acetyltransferase Tip60 with mutated acetyl-CoA binding site that abrogates its enzyme activity (mutant HeLa-tip60). Two shRNAmir were identified that reduced the expression of the cytoplasmic and nuclear forms of Hat1. Cytosolic protein preparations from these two clones showed decreased levels of acetylation of lysine 5 (K5) and K12 on histone H4, with the concomitant loss of the acetylation of histone H2A at K5. This pattern of decreased acetylation of H2AK5 was well defined in preparations of histone protein and insoluble nuclear-protein (INP) fractions as well. Abrogating the Hat1 expression caused a 74% decrease in colony-forming efficiency of mutant HeLa-tip60 cells, reduced the size of the colonies by 50%, and decreased the amounts of proteins with molecular weights below 35 kDa in the INP fractions.

  20. Bivalent histone modifications during tooth development

    Institute of Scientific and Technical Information of China (English)

    Li-Wei Zheng; Bin-Peng Zhang; Ruo-Shi Xu; Xin Xu; Ling Ye; Xue-Dong Zhou

    2014-01-01

    Histone methylation is one of the most widely studied post-transcriptional modifications. It is thought to be an important epigenetic event that is closely associated with cell fate determination and differentiation. To explore the spatiotemporal expression of histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 27 trimethylation (H3K27me3) epigenetic marks and methylation or demethylation transferases in tooth organ development, we measured the expression of SET7, EZH2, KDM5B and JMJD3 via immunohistochemistry and quantitative polymerase chain reaction (qPCR) analysis in the first molar of BALB/c mice embryos at E13.5, E15.5, E17.5, P0 and P3, respectively. We also measured the expression of H3K4me3 and H3K27me3 with immunofluorescence staining. During murine tooth germ development, methylation or demethylation transferases were expressed in a spatial–temporal manner. The bivalent modification characterized by H3K4me3 and H3K27me3 can be found during the tooth germ development, as shown by immunofluorescence. The expression of SET7, EZH2 as methylation transferases and KDM5B and JMJD3 as demethylation transferases indicated accordingly with the expression of H3K4me3 and H3K27me3 respectively to some extent. The bivalent histone may play a critical role in tooth organ development via the regulation of cell differentiation.

  1. Gene promoters dictate histone occupancy within genes.

    Science.gov (United States)

    Perales, Roberto; Erickson, Benjamin; Zhang, Lian; Kim, Hyunmin; Valiquett, Elan; Bentley, David

    2013-10-01

    Spt6 is a transcriptional elongation factor and histone chaperone that reassembles transcribed chromatin. Genome-wide H3 mapping showed that Spt6 preferentially maintains nucleosomes within the first 500 bases of genes and helps define nucleosome-depleted regions in 5' and 3' flanking sequences. In Spt6-depleted cells, H3 loss at 5' ends correlates with reduced pol II density suggesting enhanced transcription elongation. Consistent with its 'Suppressor of Ty' (Spt) phenotype, Spt6 inactivation caused localized H3 eviction over 1-2 nucleosomes at 5' ends of Ty elements. H3 displacement differed between genes driven by promoters with 'open'/DPN and 'closed'/OPN chromatin conformations with similar pol II densities. More eviction occurred on genes with 'closed' promoters, associated with 'noisy' transcription. Moreover, swapping of 'open' and 'closed' promoters showed that they can specify distinct downstream patterns of histone eviction/deposition. These observations suggest a novel function for promoters in dictating histone dynamics within genes possibly through effects on transcriptional bursting or elongation rate.

  2. Histone deacetylases and cardiovascular cell lineagecommitment

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    Cardiovascular diseases (CVDs), which include alldiseases of the heart and circulation system, arethe leading cause of deaths on the globally. Duringthe development of CVDs, choric inflammatory, lipidmetabolism disorder and endothelial dysfunction arewidely recognized risk factors. Recently, the newtreatment for CVDs that designed to regenerate thedamaged myocardium and injured vascular endotheliumand improve recovery by the use of stem cells, attractsmore and more public attention. Histone deacetylases(HDACs) are a family of enzymes that remove acetylgroups from lysine residues of histone proteinsallowing the histones to wrap the DNA more tightlyand commonly known as epigenetic regulators ofgene transcription. HDACs play indispensable roles innearly all biological processes, such as transcriptionalregulation, cell cycle progression and developmentalevents, and have originally shown to be involved incancer and neurological diseases. HDACs are alsofound to play crucial roles in cardiovascular diseases bymodulating vascular cell homeostasis (e.g. , proliferation,migration, and apoptosis of both ECs and SMCs). Thisreview focuses on the roles of different members ofHDACs and HDAC inhibitor on stem cell/ progenitor celldifferentiation toward vascular cell lineages (endothelialcells, smooth muscle cells and Cardiomyocytes) and itspotential therapeutics.

  3. Unique fluorophores in the dimeric archaeal histones hMfB and hPyA1 reveal the impact of nonnative structure in a monomeric kinetic intermediate.

    Science.gov (United States)

    Stump, Matthew R; Gloss, Lisa M

    2008-02-01

    Homodimeric archaeal histones and heterodimeric eukaryotic histones share a conserved structure but fold through different kinetic mechanisms, with a correlation between faster folding/association rates and the population of kinetic intermediates. Wild-type hMfB (from Methanothermus fervidus) has no intrinsic fluorophores; Met35, which is Tyr in hyperthermophilic archaeal histones such as hPyA1 (from Pyrococcus strain GB-3A), was mutated to Tyr and Trp. Two Tyr-to-Trp mutants of hPyA1 were also characterized. All fluorophores were introduced into the long, central alpha-helix of the histone fold. Far-UV circular dichroism (CD) indicated that the fluorophores did not significantly alter the helical content of the histones. The equilibrium unfolding transitions of the histone variants were two-state, reversible processes, with DeltaG degrees (H2O) values within 1 kcal/mol of the wild-type dimers. The hPyA1 Trp variants fold by two-state kinetic mechanisms like wild-type hPyA1, but with increased folding and unfolding rates, suggesting that the mutated residues (Tyr-32 and Tyr-36) contribute to transition state structure. Like wild-type hMfB, M35Y and M35W hMfB fold by a three-state mechanism, with a stopped-flow CD burst-phase monomeric intermediate. The M35 mutants populate monomeric intermediates with increased secondary structure and stability but exhibit decreased folding rates; this suggests that nonnative interactions occur from burial of the hydrophobic Tyr and Trp residues in this kinetic intermediate. These results implicate the long central helix as a key component of the structure in the kinetic monomeric intermediates of hMfB as well as the dimerization transition state in the folding of hPyA1.

  4. Theoretical framework for the histone modification network: modifications in the unstructured histone tails form a robust scale-free network.

    Science.gov (United States)

    Hayashi, Yohei; Senda, Toshiya; Sano, Norihiko; Horikoshi, Masami

    2009-07-01

    A rapid increase in research on the relationship between histone modifications and their subsequent reactions in the nucleus has revealed that the histone modification system is complex, and robust against point mutations. The prevailing theoretical framework (the histone code hypothesis) is inadequate to explain either the complexity or robustness, making the formulation of a new theoretical framework both necessary and desirable. Here, we develop a model of the regulatory network of histone modifications in which we encode histone modifications as nodes and regulatory interactions between histone modifications as links. This network has scale-free properties and subnetworks with a pseudo-mirror symmetry structure, which supports the robustness of the histone modification network. In addition, we show that the unstructured tail regions of histones are suitable for the acquisition of this scale-free property. Our model and related insights provide the first framework for an overall architecture of a histone modification network system, particularly with regard to the structural and functional roles of the unstructured histone tail region. In general, the post-translational "modification webs" of natively unfolded regions (proteins) may function as signal routers for the robust processing of the large amounts of signaling information.

  5. Replication-coupled chromatin assembly of newly synthesized histones: distinct functions for the histone tail domains.

    Science.gov (United States)

    Ejlassi-Lassallette, Aïda; Thiriet, Christophe

    2012-02-01

    The maintenance of the genome during replication requires the assembly of nucleosomes with newly synthesized histones. Achieving the deposition of newly synthesized histones in chromatin implies their transport from the cytoplasm to the nucleus at the replication sites. Several lines of evidence have revealed critical functions of the histone tail domains in these conserved cellular processes. In this review, we discuss the role of the amino termini of the nucleosome building blocks, H2A/H2B and H3/H4, in different model systems. The experimental data showed that H2A/H2B tails and H3/H4 tails display distinct functions in nuclear import and chromatin assembly. Furthermore, we describe recent studies exploiting the unique properties of the slime mold, Physarum polycephalum , that have advanced understanding of the function of the highly conserved replication-dependent diacetylation of H4.

  6. The Beauty of Being a Variant:H2A.Z and the SWR1 Complex in Plants

    Institute of Scientific and Technical Information of China (English)

    Rosana March-D(ī)az; Jose C.Reyes

    2009-01-01

    Numerous studies have shown that the nucleosome is a dynamic structure that strongly influences gene expression.Dynamism concerns different nucleosomal characteristics,including position,posttranslational modifications,and histone composition.Thus,within the nucleosome,canonical histones can be exchanged by histone variant proteins with specific functions-a process known as 'histone replacement'.The histone variant H2A.Z has an important function in transcription and,during the last few years,its role in plant development and immune response has become evident.Compiling genetic and biochemical studies from several laboratories has revealed that plants contain a multiprotein complex,similar to the SWR1/SRCAP complex from yeast and animals,involved in H2A.Z deposition.Despite intense research in different organisms,the mechanism by which H2A.Z influences transcription is still unknown.However,recent results from Arabidopsis have shown a strong inverse correlation between H2A.Z and DNA methylation,suggesting that H2A.Z might protect genes from silencing.

  7. Epigenetic histone marks of extended meta-polycentric centromeres of Lathyrus and Pisum chromosomes

    Directory of Open Access Journals (Sweden)

    Pavel eNeumann

    2016-03-01

    Full Text Available Species of the legume genera Lathyrus and Pisum possess chromosomes that exhibit a unique structure of their centromeric regions, which is clearly apparent during metaphase by the formation of extended primary constrictions which span up to a third of the length of the chromosome. In addition, these species express two different variants of the CenH3 protein which are co-localized in multiple domains along the poleward surface of the primary constrictions. Here we show that the constrictions represent a distinct type of chromatin differing from the chromosome arms. In metaphase, histone phosphorylation patterns including H3S10ph, H3S28ph and H3T3ph were observed along the entire constriction, in a way similar to holocentric chromosomes. On the other hand, distribution of phosphorylated H2AT120 was different from that previously reported from either, holocentric and monocentric chromosomes, occurring at chromatin surrounding but not overlapping CenH3 domains. Since some of these phosphorylations play a role in chromatid cohesion, it can be assumed that they facilitate correct chromosome segregation by ensuring that multiple separate CenH3 domains present on the same chromatid are oriented towards the same pole. The constrictions also displayed distinct patterns of histone methylation marks, being enriched in H3K9me2 and depleted in H3K4me3 and H3K27me2 compared to the chromosome arms. High resolution fluorescence microscopy revealed that although both CenH3 protein variants are present in all CenH3 domains detected on metaphase chromosomes, they are only partially co-localized while there are chromatin subdomains which are mostly made of only one CenH3 variant. Taken together, these data revealed specific features of extended primary constrictions of Lathyrus and Pisum and support the idea that they may represent an intermediate stage between monocentric and holocentric chromosomes.

  8. Meiosis-Specific Loading of the Centromere-Specific Histone CENH3 in Arabidopsis thaliana

    Science.gov (United States)

    Ravi, Maruthachalam; Shibata, Fukashi; Ramahi, Joseph S.; Nagaki, Kiyotaka; Chen, Changbin; Murata, Minoru; Chan, Simon W. L.

    2011-01-01

    Centromere behavior is specialized in meiosis I, so that sister chromatids of homologous chromosomes are pulled toward the same side of the spindle (through kinetochore mono-orientation) and chromosome number is reduced. Factors required for mono-orientation have been identified in yeast. However, comparatively little is known about how meiotic centromere behavior is specialized in animals and plants that typically have large tandem repeat centromeres. Kinetochores are nucleated by the centromere-specific histone CENH3. Unlike conventional histone H3s, CENH3 is rapidly evolving, particularly in its N-terminal tail domain. Here we describe chimeric variants of CENH3 with alterations in the N-terminal tail that are specifically defective in meiosis. Arabidopsis thaliana cenh3 mutants expressing a GFP-tagged chimeric protein containing the H3 N-terminal tail and the CENH3 C-terminus (termed GFP-tailswap) are sterile because of random meiotic chromosome segregation. These defects result from the specific depletion of GFP-tailswap protein from meiotic kinetochores, which contrasts with its normal localization in mitotic cells. Loss of the GFP-tailswap CENH3 variant in meiosis affects recruitment of the essential kinetochore protein MIS12. Our findings suggest that CENH3 loading dynamics might be regulated differently in mitosis and meiosis. As further support for our hypothesis, we show that GFP-tailswap protein is recruited back to centromeres in a subset of pollen grains in GFP-tailswap once they resume haploid mitosis. Meiotic recruitment of the GFP-tailswap CENH3 variant is not restored by removal of the meiosis-specific cohesin subunit REC8. Our results reveal the existence of a specialized loading pathway for CENH3 during meiosis that is likely to involve the hypervariable N-terminal tail. Meiosis-specific CENH3 dynamics may play a role in modulating meiotic centromere behavior. PMID:21695238

  9. NICKEL COMPOUNDS INDUCE HISTONE UBIQUITINATION BY INHIBITING HISTONE DEUBIQUITINATING ENZYME ACTIVITY

    OpenAIRE

    Ke, Qingdong; Ellen, Thomas P.; Costa, Max

    2007-01-01

    Nickel (Ni) compounds are known carcinogens but underlying mechanisms are not clear. Epigenetic changes are likely to play an important role in nickel ion carcinogenesis. Previous studies have shown epigenetic effects of nickel ions, including the loss of histone acetylation and a pronounced increase in dimethylated H3K9 in nickel-exposed cells. In this study, we demonstrated that both water-soluble and insoluble nickel compounds induce histone ubiquitination (uH2A and uH2B) in a variety of c...

  10. Linker histone H1 and protein-protein interactions

    OpenAIRE

    Kalashnikova, Anna A; Rogge, Ryan A.; Hansen, Jeffrey C

    2015-01-01

    Linker histones H1 are ubiquitous chromatin proteins that play important roles in chromatin compaction, transcription regulation, nucleosome spacing and chromosome spacing. H1 function in DNA and chromatin structure stabilization is well studied and established. The current paradigm of linker histone mode of function considers all other cellular roles of linker histones to be a consequence from H1 chromatin compaction and repression. Here we review the multiple processes regulated by linker h...

  11. Roles of histone ubiquitylation in DNA damage signaling

    Institute of Scientific and Technical Information of China (English)

    Sui-Sui DONG; Michael S. Y. HUEN

    2011-01-01

    Histone ubiquitylation has emerged as an important chromatin modification associated with DNA damage signaling and repair pathways.These histone marks,laid down by E3 ubiquitin ligases that include RNF8 and RNF168,decorate chromatin domains surrounding DNA double-strand breaks (DSBs).Recent work implicated ubiquitylated histones in orchestrating cell cycle checkpoints,DNA repair and gene transcription.Here we summarize recent advances that contribute to our current knowledge of the highly dynamic nature of DSB-associated histone ubiquitylation,and discuss major challenges ahead in understanding the versatility of ubiquitin conjugation in maintaining genome stability.

  12. Roles of histones and nucleosomes in gene transcription

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This article reviews the latest research developments in the field of eukaryotic gene regulation by the structural alterations of chromatin and nucleosomes. The following issues are briefly addressed: (ⅰ) nucleosome and histone modifications by both the ATP-dependent remodel- ing com-plexes and the histone acetyltransferases and their roles in gene activation; (ⅱ) competitive binding of histones and transcription factors on gene promoters, and transcription repression by nucleosomes; and (ⅲ) influences of linker histone H1 on gene regulation. Meanwhile, the significance and impact of these new research progresses, as well as issues worthwhile for further study are commented.

  13. Histone modifications and alcohol-induced liver disease: Are altered nutrients the missing link?

    Institute of Scientific and Technical Information of China (English)

    Akshata Moghe; Swati Joshi-Barve; Smita Ghare; Leila Gobejishvili; Irina Kirpich; Craig J McClain; Shirish Barve

    2011-01-01

    Alcoholism is a major health problem in the United States and worldwide, and alcohol remains the single most significant cause of liver-related diseases and deaths. Alcohol is known to influence nutritional status at many levels including nutrient intake, absorption, utilization, and excretion, and can lead to many nutritional disturbances and deficiencies. Nutrients can dramatically affect gene expression and alcohol-induced nutrient imbalance may be a major contributor to pathogenic gene expression in alcohol-induced liver disease (ALD). There is growing interest regarding epigenetic changes, including histone modifications that regulate gene expression during disease pathogenesis. Notably, modifications of core histones in the nucleosome regulate chromatin structure and DNA methylation, and control gene transcription. This review highlights the role of nutrient disturbances brought about during alcohol metabolism and their impact on epigenetic histone modifications that may contribute to ALD. The review is focused on four critical metabolites, namely, acetate, S-adenosylmethionine, nicotinamide adenine dinucleotide and zinc that are particularly relevant to alcohol metabolism and ALD.

  14. Histone Posttranslational Modifications of CD4+ T Cell in Autoimmune Diseases

    Directory of Open Access Journals (Sweden)

    Zijun Wang

    2016-09-01

    Full Text Available The complexity of immune system is tempered by precise regulation to maintain stabilization when exposed to various conditions. A subtle change in gene expression may be magnified when drastic changes are brought about in cellular development and function. Posttranslational modifications (PTMs timely alter the functional activity of immune system, and work proceeded in these years has begun to throw light upon it. Posttranslational modifications of histone tails have been mentioned in a large scale of biological developments and disease progression, thereby making them a central field to investigate. Conventional assessments of these changes are centered on the transcription factors and cytokines in T cells regulated by variable histone codes to achieve chromatin remodeling, as well as involved in many human diseases, especially autoimmune diseases. We here put forward an essential review of core posttranslational modulations that regulate T cell function and differentiation in the immune system, with a special emphasis on histone modifications in different T helper cell subsets as well as in autoimmune diseases.

  15. dbHiMo: a web-based epigenomics platform for histone-modifying enzymes.

    Science.gov (United States)

    Choi, Jaeyoung; Kim, Ki-Tae; Huh, Aram; Kwon, Seomun; Hong, Changyoung; Asiegbu, Fred O; Jeon, Junhyun; Lee, Yong-Hwan

    2015-01-01

    Over the past two decades, epigenetics has evolved into a key concept for understanding regulation of gene expression. Among many epigenetic mechanisms, covalent modifications such as acetylation and methylation of lysine residues on core histones emerged as a major mechanism in epigenetic regulation. Here, we present the database for histone-modifying enzymes (dbHiMo; http://hme.riceblast.snu.ac.kr/) aimed at facilitating functional and comparative analysis of histone-modifying enzymes (HMEs). HMEs were identified by applying a search pipeline built upon profile hidden Markov model (HMM) to proteomes. The database incorporates 11,576 HMEs identified from 603 proteomes including 483 fungal, 32 plants and 51 metazoan species. The dbHiMo provides users with web-based personalized data browsing and analysis tools, supporting comparative and evolutionary genomics. With comprehensive data entries and associated web-based tools, our database will be a valuable resource for future epigenetics/epigenomics studies. © The Author(s) 2015. Published by Oxford University Press.

  16. Comparison of peptide mass mapping and electron capture dissociation as assays for histone posttranslational modifications

    Science.gov (United States)

    Zhang, Liwen; Freitas, Michael A.

    2004-05-01

    Posttranslational modifications of core histones play a critical role in the structure of chromatin and the regulation of gene activities. Improved techniques for determining these modification sites may lead to a better understanding of histone regulation at the molecular level. LC-MS peptide mass mapping was performed on pepsin, trypsin and Glu-C digests of bovine thymus H4 using a QqTOF instrument. The well established modification sites of H4 (acetylation of K8, 12, 16 and methylation of K20) were observed in addition to several recently discovered modifications including: methylation of K31, 44, 59 and acetylation of K20, 77, 79. For comparison, electron capture dissociation (ECD) was performed on intact H4 along with several peptides from enzymatic digestion. The results from the ECD experiments of histone H4 indicated the acetylation of K5, 12, 16, 31, 91 and the methylation of K20 and 59 in good agreement with the result from peptide mapping. The work is dedicated to Alan G. Marshall on his 60th birthday. His endeavors in the advancement of FT-ICR facilitated experiments reported herein.

  17. N- and C-terminal domains determine differential nucleosomal binding geometry and affinity of linker histone isotypes H1(0) and H1c.

    Science.gov (United States)

    Vyas, Payal; Brown, David T

    2012-04-01

    Eukaryotic linker or H1 histones modulate DNA compaction and gene expression in vivo. In mammals, these proteins exist as multiple isotypes with distinct properties, suggesting a functional significance to the heterogeneity. Linker histones typically have a tripartite structure composed of a conserved central globular domain flanked by a highly variable short N-terminal domain and a longer highly basic C-terminal domain. We hypothesized that the variable terminal domains of individual subtypes contribute to their functional heterogeneity by influencing chromatin binding interactions. We developed a novel dual color fluorescence recovery after photobleaching assay system in which two H1 proteins fused to spectrally separable fluorescent proteins can be co-expressed and their independent binding kinetics simultaneously monitored in a single cell. This approach was combined with domain swap and point mutagenesis to determine the roles of the terminal domains in the differential binding characteristics of the linker histone isotypes, mouse H1(0) and H1c. Exchanging the N-terminal domains between H1(0) and H1c changed their overall binding affinity to that of the other variant. In contrast, switching the C-terminal domains altered the chromatin interaction surface of the globular domain. These results indicate that linker histone subtypes bind to chromatin in an intrinsically specific manner and that the highly variable terminal domains contribute to differences between subtypes. The methods developed in this study will have broad applications in studying dynamic properties of additional histone subtypes and other mobile proteins.

  18. Prediction of RNA Polymerase II recruitment, elongation and stalling from histone modification data

    DEFF Research Database (Denmark)

    Chen, Yun; Jørgensen, Mette; Kolde, Raivo

    2011-01-01

    ABSTRACT: BACKGROUND: Initiation and elongation of RNA polymerase II (RNAPII) transcription is regulated by both DNA sequence and chromatin signals. Recent breakthroughs make it possible to measure the chromatin state and activity of core promoters genome-wide, but dedicated computational...... in the gene body, the mRNA production originating from the promoter and finally also the stalling characteristics of RNAPII by considering both quantitative and spatial features of histone modifications around the transcription start site (TSS). As the model framework can also pinpoint the signals...

  19. Hemoglobin Variants in Mice

    Energy Technology Data Exchange (ETDEWEB)

    Popp, Raymond A.

    1965-04-22

    Variability among mammalian hemoglobins was observed many years ago (35). The chemical basis for differences among hemoglobins from different species of mammals has been studied by several investigators (5, 11, 18, 48). As well as interspecies differences, hemoglobin variants are frequently found within a species of mammals (2, 3, 7, 16) The inheritance of these intraspecies variants can be studied, and pedigrees indicate that the type of hemoglobin synthesized in an individual is genetically controlled (20). Several of the variant human hemoglobins are f'unctionally deficient (7, 16). Such hemoglobin anomalies are of basic interest to man because of the vital role of hemoglobin for transporting oxygen to all tissues of the body.

  20. Epigenetic regulation of GATA4 expression by histone modification in AFP-producing gastric adenocarcinoma.

    Science.gov (United States)

    Yamamura, Nobuhisa; Kishimoto, Takashi

    2012-08-01

    AFP-producing adenocarcinoma is a variant of adenocarcinoma with high malignancy. Production of AFP suggests enteroblastic or hepatoid differentiation of cancer cells. GATA4 is a key molecule involved in the prenatal development of the stomach and liver. GATA4 is epigenetically silenced by hypermethylation of primer region in many types of cancers including gastric cancer. The aim of this study is to investigate the expression and epigenetic regulation of GATA4 in AFP-producing adenocarcinoma. Immunohistochemical analysis revealed that GATA4 was positive in 3/8 cases of AFP-producing gastric adenocarcinomas and in 28/30 cases of common type adenocarcinomas. Epigenetic modification of GATA4 promoter region was investigated with 3 AFP-producing and 4 common-type gastric cancer cell lines. GATA4 mRNA was detected in 1/3 of AFP-producing and 2/4 of common-type gastric cancer cell lines by RT-PCR. Methylation-specific PCR revealed no GATA4 methylation in any of the AFP-producing gastric cancers, whereas methylation was consistent with GATA4 expression in the common-type gastric cancers. Chromatin immunoprecipitation assay for AFP-producing gastric cancers revealed that histones H3 and H4 were hypoacetylated in the GATA4-negative cells, while they were hyperacetylated in the GATA4-positive cells. Treatment with trichostain A, an inhibitor for histone deacetylase, induced acetylation of histones H3 and H4, and tri-methylation of lysine 4 of histone H3, which was associated with the active transcription of GATA4 in GATA4-negative AFP-producing cells. These results indicated that histone deacetylation is a silencing mechanism for GATA4 expression in AFP-producing gastric cancer cells. Differences between AFP-producing gastric cancer and common-type gastric cancer in terms of the mechanism of GATA4 regulation may be reflected in the phenotypic deviation of AFP-producing gastric cancer from common-type gastric cancer.

  1. Structure and function of the histone chaperone CIA/ASF1 complexed with histones H3 and H4.

    Science.gov (United States)

    Natsume, Ryo; Eitoku, Masamitsu; Akai, Yusuke; Sano, Norihiko; Horikoshi, Masami; Senda, Toshiya

    2007-03-15

    CIA (CCG1-interacting factor A)/ASF1, which is the most conserved histone chaperone among the eukaryotes, was genetically identified as a factor for an anti-silencing function (Asf1) by yeast genetic screening. Shortly after that, the CIA-histone-H3-H4 complex was isolated from Drosophila as a histone chaperone CAF-1 stimulator. Human CIA-I/II (ASF1a/b) was identified as a histone chaperone that interacts with the bromodomain-an acetylated-histone-recognizing domain-of CCG1, in the general transcription initiation factor TFIID. Intensive studies have revealed that CIA/ASF1 mediates nucleosome assembly by forming a complex with another histone chaperone in human cells and yeast, and is involved in DNA replication, transcription, DNA repair and silencing/anti-silencing in yeast. CIA/ASF1 was shown as a major storage chaperone for soluble histones in proliferating human cells. Despite all these biochemical and biological functional analyses, the structure-function relationship of the nucleosome assembly/disassembly activity of CIA/ASF1 has remained elusive. Here we report the crystal structure, at 2.7 A resolution, of CIA-I in complex with histones H3 and H4. The structure shows the histone H3-H4 dimer's mutually exclusive interactions with another histone H3-H4 dimer and CIA-I. The carboxy-terminal beta-strand of histone H4 changes its partner from the beta-strand in histone H2A to that of CIA-I through large conformational change. In vitro functional analysis demonstrated that CIA-I has a histone H3-H4 tetramer-disrupting activity. Mutants with weak histone H3-H4 dimer binding activity showed critical functional effects on cellular processes related to transcription. The histone H3-H4 tetramer-disrupting activity of CIA/ASF1 and the crystal structure of the CIA/ASF1-histone-H3-H4 dimer complex should give insights into mechanisms of both nucleosome assembly/disassembly and nucleosome semi-conservative replication.

  2. Transformer core

    NARCIS (Netherlands)

    Mehendale, A.; Hagedoorn, Wouter; Lötters, Joost Conrad

    2010-01-01

    A transformer core includes a stack of a plurality of planar core plates of a magnetically permeable material, which plates each consist of a first and a second sub-part that together enclose at least one opening. The sub-parts can be fitted together via contact faces that are located on either side

  3. Transformer core

    NARCIS (Netherlands)

    Mehendale, A.; Hagedoorn, Wouter; Lötters, Joost Conrad

    2008-01-01

    A transformer core includes a stack of a plurality of planar core plates of a magnetically permeable material, which plates each consist of a first and a second sub-part that together enclose at least one opening. The sub-parts can be fitted together via contact faces that are located on either side

  4. Histone lysine crotonylation during acute kidney injury in mice

    Directory of Open Access Journals (Sweden)

    Olga Ruiz-Andres

    2016-06-01

    Full Text Available Acute kidney injury (AKI is a potentially lethal condition for which no therapy is available beyond replacement of renal function. Post-translational histone modifications modulate gene expression and kidney injury. Histone crotonylation is a recently described post-translational modification. We hypothesized that histone crotonylation might modulate kidney injury. Histone crotonylation was studied in cultured murine proximal tubular cells and in kidneys from mice with AKI induced by folic acid or cisplatin. Histone lysine crotonylation was observed in tubular cells from healthy murine and human kidney tissue. Kidney tissue histone crotonylation increased during AKI. This was reproduced by exposure to the protein TWEAK in cultured tubular cells. Specifically, ChIP-seq revealed enrichment of histone crotonylation at the genes encoding the mitochondrial biogenesis regulator PGC-1α and the sirtuin-3 decrotonylase in both TWEAK-stimulated tubular cells and in AKI kidney tissue. To assess the role of crotonylation in kidney injury, crotonate was used to increase histone crotonylation in cultured tubular cells or in the kidneys in vivo. Crotonate increased the expression of PGC-1α and sirtuin-3, and decreased CCL2 expression in cultured tubular cells and healthy kidneys. Systemic crotonate administration protected from experimental AKI, preventing the decrease in renal function and in kidney PGC-1α and sirtuin-3 levels as well as the increase in CCL2 expression. For the first time, we have identified factors such as cell stress and crotonate availability that increase histone crotonylation in vivo. Overall, increasing histone crotonylation might have a beneficial effect on AKI. This is the first observation of the in vivo potential of the therapeutic manipulation of histone crotonylation in a disease state.

  5. Photoperiodic regulation of flowering time through periodic histone deacetylation of the florigen gene FT.

    Directory of Open Access Journals (Sweden)

    Xiaofeng Gu

    2013-09-01

    Full Text Available The developmental transition from a vegetative to a reproductive phase (i.e., flowering is timed by the seasonal cue day length or photoperiod in many plant species. Through the photoperiod pathway, inductive day lengths trigger the production of a systemic flowering signal, florigen, to provoke the floral transition. FLOWERING LOCUS T (FT, widely conserved in angiosperms, is a major component of the mobile florigen. In the long-day plant Arabidopsis, FT expression is rhythmically activated by the output of the photoperiod pathway CONSTANS (CO, specifically at the end of long days. How FT expression is modulated at an adequate level in response to the long-day cue to set a proper flowering time remains unknown. Here, we report a periodic histone deacetylation mechanism for the photoperiodic modulation of FT expression. We have identified a plant-unique core structural component of an Arabidopsis histone deacetylase (HDAC complex. In long days, this component accumulates at dusk, and is recruited by a MADS-domain transcription factor to the FT locus specifically at the end of the day, leading to periodic histone deacetylation of FT chromatin at dusk. Furthermore, we found that at the end of long days CO activity not only activates FT expression but also enables HDAC-activity recruitment to FT chromatin to dampen the level of FT expression, and so prevent precocious flowering in response to the inductive long-day cue. These results collectively reveal a periodic histone deacetylation mechanism for the day-length control of flowering time in higher plants.

  6. Photoperiodic regulation of flowering time through periodic histone deacetylation of the florigen gene FT.

    Science.gov (United States)

    Gu, Xiaofeng; Wang, Yizhong; He, Yuehui

    2013-09-01

    The developmental transition from a vegetative to a reproductive phase (i.e., flowering) is timed by the seasonal cue day length or photoperiod in many plant species. Through the photoperiod pathway, inductive day lengths trigger the production of a systemic flowering signal, florigen, to provoke the floral transition. FLOWERING LOCUS T (FT), widely conserved in angiosperms, is a major component of the mobile florigen. In the long-day plant Arabidopsis, FT expression is rhythmically activated by the output of the photoperiod pathway CONSTANS (CO), specifically at the end of long days. How FT expression is modulated at an adequate level in response to the long-day cue to set a proper flowering time remains unknown. Here, we report a periodic histone deacetylation mechanism for the photoperiodic modulation of FT expression. We have identified a plant-unique core structural component of an Arabidopsis histone deacetylase (HDAC) complex. In long days, this component accumulates at dusk, and is recruited by a MADS-domain transcription factor to the FT locus specifically at the end of the day, leading to periodic histone deacetylation of FT chromatin at dusk. Furthermore, we found that at the end of long days CO activity not only activates FT expression but also enables HDAC-activity recruitment to FT chromatin to dampen the level of FT expression, and so prevent precocious flowering in response to the inductive long-day cue. These results collectively reveal a periodic histone deacetylation mechanism for the day-length control of flowering time in higher plants.

  7. Secondary Structure Alterations of Histones H2A and H2B in X-Irradiated Human Cancer Cells: Altered Histones Persist in Cells for at Least 24 Hours.

    Science.gov (United States)

    Izumi, Yudai; Yamamoto, Satoshi; Fujii, Kentaro; Yokoya, Akinari

    2015-11-01

    We measured and compared the circular dichroism (CD) spectra and secondary structures of histone proteins H2A, H2B and their variants extracted from X-irradiated and unirradiated human HeLa cells. Compared to unirradiated cells, a relative increase in α-helix structure and decrease in other secondary structures was observed in X-irradiated cells. These structural alterations persisted for at least 24 h, which is substantially longer than the 2 h generally known to be required for DNA double-strand break repair.

  8. The histone chaperones Vps75 and Nap1 form ring-like, tetrameric structures in solution

    DEFF Research Database (Denmark)

    Bowman, Andrew; Hammond, Colin M; Stirling, Andrew

    2014-01-01

    NAP-1 fold histone chaperones play an important role in escorting histones to and from sites of nucleosome assembly and disassembly. The two NAP-1 fold histone chaperones in budding yeast, Vps75 and Nap1, have previously been crystalized in a characteristic homodimeric conformation. In this study...... fold histone chaperones. The tetramerisation of NAP-1 fold histone chaperones may act to shield acidic surfaces in the absence of histone cargo thus providing a 'self-chaperoning' type mechanism....

  9. Reorganization of Damaged Chromatin by the Exchange of Histone Variant H2A.Z-2

    Energy Technology Data Exchange (ETDEWEB)

    Nishibuchi, Ikuno [Department of Cellular Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima (Japan); Department of Radiation Oncology, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima (Japan); Department of Radiation Oncology, Hiroshima Prefectural Hospital, Hiroshima (Japan); Suzuki, Hidekazu; Kinomura, Aiko; Sun, Jiying; Liu, Ning-Ang [Department of Cellular Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima (Japan); Horikoshi, Yasunori [Department of Cellular Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima (Japan); Research Center for Mathematics of Chromatin Live Dynamics, Hiroshima University, Hiroshima (Japan); Shima, Hiroki [Department of Biochemistry, Graduate School of Medical Sciences, Tohoku University, Sendai (Japan); Kusakabe, Masayuki; Harata, Masahiko [Laboratory of Molecular Biology, Graduate School of Agricultural Science, Tohoku University, Sendai (Japan); Fukagawa, Tatsuo [Department of Molecular Genetics, National Institute of Genetics and The Graduate University for Advanced Studies, Mishima (Japan); Ikura, Tsuyoshi [Laboratory of Chromatin Regulatory Network, Department of Mutagenesis, Radiation Biology Center, Kyoto University, Kyoto (Japan); Ishida, Takafumi [Department of Cardiovascular Medicine, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima (Japan); Nagata, Yasushi [Department of Radiation Oncology, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima (Japan); Tashiro, Satoshi, E-mail: ktashiro@hiroshima-u.ac.jp [Department of Cellular Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima (Japan); Research Center for Mathematics of Chromatin Live Dynamics, Hiroshima University, Hiroshima (Japan)

    2014-07-15

    Purpose: The reorganization of damaged chromatin plays an important role in the regulation of the DNA damage response. A recent study revealed the presence of 2 vertebrate H2A.Z isoforms, H2A.Z-1 and H2A.Z-2. However, the roles of the vertebrate H2A.Z isoforms are still unclear. Thus, in this study we examined the roles of the vertebrate H2A.Z isoforms in chromatin reorganization after the induction of DNA double-strand breaks (DSBs). Methods and Materials: To examine the dynamics of H2A.Z isoforms at damaged sites, we constructed GM0637 cells stably expressing each of the green fluorescent protein (GFP)-labeled H2A.Z isoforms, and performed fluorescence recovery after photobleaching (FRAP) analysis and inverted FRAP analysis in combination with microirradiation. Immunofluorescence staining using an anti-RAD51 antibody was performed to study the kinetics of RAD51 foci formation after 2-Gy irradiation of wild-type (WT), H2A.Z-1- and H2A.Z-2-deficient DT40 cells. Colony-forming assays were also performed to compare the survival rates of WT, H2A.Z-1-, and H2A.Z-2-deficient DT40 cells with control, and H2A.Z-1- and H2A.Z-2-depleted U2OS cells after irradiation. Results: FRAP analysis revealed that H2A.Z-2 was incorporated into damaged chromatin just after the induction of DSBs, whereas H2A.Z-1 remained essentially unchanged. Inverted FRAP analysis showed that H2A.Z-2 was released from damaged chromatin. These findings indicated that H2A.Z-2 was exchanged at DSB sites immediately after the induction of DSBs. RAD51 focus formation after ionizing irradiation was disturbed in H2A.Z-2-deficient DT40 cells but not in H2A.Z-1-deficient cells. The survival rate of H2A.Z-2-deficient cells after irradiation was lower than those of WT and H2A.Z-1- DT40 cells. Similar to DT40 cells, H2A.Z-2-depleted U2OS cells were also radiation-sensitive compared to control and H2A.Z-1-depleted cells. Conclusions: We found that vertebrate H2A.Z-2 is involved in the regulation of the DNA damage response at a very early stage, via the damaged chromatin reorganization required for RAD51 focus formation.

  10. SIRT1-metabolite binding histone macroH2A1.1 protects hepatocytes against lipid accumulation

    OpenAIRE

    Pazienza, Valerio; Borghesan, Michela; Mazza, Tommaso; Sheedfar, Fareeba; Panebianco, Concetta; Williams, Roger; Mazzoccoli, Gianluigi; Andriulli, Angelo; Nakanishi, Tomoko; Vinciguerra, Manlio

    2014-01-01

    Non-alcoholic-fatty-liver-disease (NAFLD) encompasses conditions associated to fat deposition in the liver, which are generally deteriorated during the aging process. MacroH2A1, a variant of histone H2A, is a key transcriptional regulator involved in tumorigenic processes and cell senescence, and featuring two alternatively splicing isoforms, macroH2A1.1 and macroH2A1.2. MacroH2A1.1 binds with high affinity O-acetyl ADP ribose, a small metabolite produced by the reaction catalysed by NAD+-dep...

  11. Erasing the methyl mark: histone demethylases at the center of cellular differentiation and disease

    DEFF Research Database (Denmark)

    Cloos, Paul A C; Christensen, Jesper; Agger, Karl;

    2008-01-01

    The enzymes catalyzing lysine and arginine methylation of histones are essential for maintaining transcriptional programs and determining cell fate and identity. Until recently, histone methylation was regarded irreversible. However, within the last few years, several families of histone...... demethylases erasing methyl marks associated with gene repression or activation have been identified, underscoring the plasticity and dynamic nature of histone methylation. Recent discoveries have revealed that histone demethylases take part in large multiprotein complexes synergizing with histone deacetylases......, histone methyltransferases, and nuclear receptors to control developmental and transcriptional programs. Here we review the emerging biochemical and biological functions of the histone demethylases and discuss their potential involvement in human diseases, including cancer....

  12. Histone lysine demethylases as targets for anticancer therapy

    DEFF Research Database (Denmark)

    Højfeldt, Jonas W; Agger, Karl; Helin, Kristian

    2013-01-01

    interesting drug targets. The successful introduction of DNA methylation and histone deacetylase (HDAC) inhibitors for the treatment of specific subtypes of cancer has paved the way for the use of epigenetic therapy. Here, we highlight key biological findings demonstrating the roles of members of the histone...

  13. Enzyme kinetics and inhibition of histone acetyltransferase KAT8

    NARCIS (Netherlands)

    Wapenaar, Hannah; van der Wouden, Petra E; Groves, Matthew R; Rotili, Dante; Mai, Antonello; Dekker, Frank J

    2015-01-01

    Lysine acetyltransferase 8 (KAT8) is a histone acetyltransferase (HAT) responsible for acetylating lysine 16 on histone H4 (H4K16) and plays a role in cell cycle progression as well as acetylation of the tumor suppressor protein p53. Further studies on its biological function and drug discovery

  14. Interrogating the function of metazoan histones using engineered gene clusters.

    Science.gov (United States)

    McKay, Daniel J; Klusza, Stephen; Penke, Taylor J R; Meers, Michael P; Curry, Kaitlin P; McDaniel, Stephen L; Malek, Pamela Y; Cooper, Stephen W; Tatomer, Deirdre C; Lieb, Jason D; Strahl, Brian D; Duronio, Robert J; Matera, A Gregory

    2015-02-09

    Histones and their posttranslational modifications influence the regulation of many DNA-dependent processes. Although an essential role for histone-modifying enzymes in these processes is well established, defining the specific contribution of individual histone residues remains a challenge because many histone-modifying enzymes have nonhistone targets. This challenge is exacerbated by the paucity of suitable approaches to genetically engineer histone genes in metazoans. Here, we describe a platform in Drosophila for generating and analyzing any desired histone genotype, and we use it to test the in vivo function of three histone residues. We demonstrate that H4K20 is neither essential for DNA replication nor for completion of development, unlike inferences drawn from analyses of H4K20 methyltransferases. We also show that H3K36 is required for viability and H3K27 is essential for maintenance of cellular identity but not for gene activation. These findings highlight the power of engineering histones to interrogate genome structure and function in animals. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Structure and nucleosome interaction of the yeast NuA4 and Piccolo-NuA4 histone acetyltransferase complexes.

    Science.gov (United States)

    Chittuluru, Johnathan R; Chaban, Yuriy; Monnet-Saksouk, Julie; Carrozza, Michael J; Sapountzi, Vasileia; Selleck, William; Huang, Jiehuan; Utley, Rhea T; Cramet, Myriam; Allard, Stephane; Cai, Gang; Workman, Jerry L; Fried, Michael G; Tan, Song; Côté, Jacques; Asturias, Francisco J

    2011-10-09

    We have used EM and biochemistry to characterize the structure of NuA4, an essential yeast histone acetyltransferase (HAT) complex conserved throughout eukaryotes, and we have determined the interaction of NuA4 with the nucleosome core particle (NCP). The ATM-related Tra1 subunit, which is shared with the SAGA coactivator complex, forms a large domain joined to a second region that accommodates the catalytic subcomplex Piccolo and other NuA4 subunits. EM analysis of a NuA4-NCP complex shows the NCP bound at the periphery of NuA4. EM characterization of Piccolo and Piccolo-NCP provided further information about subunit organization and confirmed that histone acetylation requires minimal contact with the NCP. A small conserved region at the N terminus of Piccolo subunit enhancer of Polycomb-like 1 (Epl1) is essential for NCP interaction, whereas the subunit yeast homolog of mammalian Ing1 2 (Yng2) apparently positions Piccolo for efficient acetylation of histone H4 or histone H2A tails. Taken together, these results provide an understanding of the NuA4 subunit organization and the NuA4-NCP interactions.

  16. Histone deacetylase complexes promote trinucleotide repeat expansions.

    Directory of Open Access Journals (Sweden)

    Kim Debacker

    2012-02-01

    Full Text Available Expansions of DNA trinucleotide repeats cause at least 17 inherited neurodegenerative diseases, such as Huntington's disease. Expansions can occur at frequencies approaching 100% in affected families and in transgenic mice, suggesting that specific cellular proteins actively promote (favor expansions. The inference is that expansions arise due to the presence of these promoting proteins, not their absence, and that interfering with these proteins can suppress expansions. The goal of this study was to identify novel factors that promote expansions. We discovered that specific histone deacetylase complexes (HDACs promote CTG•CAG repeat expansions in budding yeast and human cells. Mutation or inhibition of yeast Rpd3L or Hda1 suppressed up to 90% of expansions. In cultured human astrocytes, expansions were suppressed by 75% upon inhibition or knockdown of HDAC3, whereas siRNA against the histone acetyltransferases CBP/p300 stimulated expansions. Genetic and molecular analysis both indicated that HDACs act at a distance from the triplet repeat to promote expansions. Expansion assays with nuclease mutants indicated that Sae2 is one of the relevant factors regulated by Rpd3L and Hda1. The causal relationship between HDACs and expansions indicates that HDACs can promote mutagenesis at some DNA sequences. This relationship further implies that HDAC3 inhibitors being tested for relief of expansion-associated gene silencing may also suppress somatic expansions that contribute to disease progression.

  17. Methylation of histone H3 lysine 9 occurs during translation

    Science.gov (United States)

    Rivera, Carlos; Saavedra, Francisco; Alvarez, Francisca; Díaz-Celis, César; Ugalde, Valentina; Li, Jianhua; Forné, Ignasi; Gurard-Levin, Zachary A.; Almouzni, Geneviève; Imhof, Axel; Loyola, Alejandra

    2015-01-01

    Histone post-translational modifications are key contributors to chromatin structure and function, and participate in the maintenance of genome stability. Understanding the establishment and maintenance of these marks, along with their misregulation in pathologies is thus a major focus in the field. While we have learned a great deal about the enzymes regulating histone modifications on nucleosomal histones, much less is known about the mechanisms establishing modifications on soluble newly synthesized histones. This includes methylation of lysine 9 on histone H3 (H3K9), a mark that primes the formation of heterochromatin, a critical chromatin landmark for genome stability. Here, we report that H3K9 mono- and dimethylation is imposed during translation by the methyltransferase SetDB1. We discuss the importance of these results in the context of heterochromatin establishment and maintenance and new therapeutic opportunities in pathologies where heterochromatin is perturbed. PMID:26405197

  18. New histone supply regulates replication fork speed and PCNA unloading

    DEFF Research Database (Denmark)

    Mejlvang, Jakob; Feng, Yunpeng; Alabert, Constance;

    2014-01-01

    Correct duplication of DNA sequence and its organization into chromatin is central to genome function and stability. However, it remains unclear how cells coordinate DNA synthesis with provision of new histones for chromatin assembly to ensure chromosomal stability. In this paper, we show...... that replication fork speed is dependent on new histone supply and efficient nucleosome assembly. Inhibition of canonical histone biosynthesis impaired replication fork progression and reduced nucleosome occupancy on newly synthesized DNA. Replication forks initially remained stable without activation...... of conventional checkpoints, although prolonged histone deficiency generated DNA damage. PCNA accumulated on newly synthesized DNA in cells lacking new histones, possibly to maintain opportunity for CAF-1 recruitment and nucleosome assembly. Consistent with this, in vitro and in vivo analysis showed that PCNA...

  19. Heterologous expression of human H1 histones in yeast.

    Science.gov (United States)

    Albig, W; Runge, D M; Kratzmeier, M; Doenecke, D

    1998-09-18

    The complete set of seven human H1 histone subtype genes was heterologously expressed in yeast. Since Saccharomyces cerevisiae lacks standard histone H1 we could isolate each recombinantly expressed human H1 subtype in pure form without contamination by endogenous H I histones. For isolation of the H1 histones in this expression system no tagging was needed and the isoforms could be extracted with the authentic primary structure by a single extraction step with 5%(0.74 M) perchloric acid. The isolated H1 histone proteins were used to assign the subtype genes to the corresponding protein spots or peaks after two-dimensional gel electrophoresis and capillary zone electrophoresis, respectively. This allowed us to correlate transcriptional data with protein data, which was barely possible until now.

  20. RNF8-dependent histone modifications regulate nucleosome removal during spermatogenesis.

    Science.gov (United States)

    Lu, Lin-Yu; Wu, Jiaxue; Ye, Lin; Gavrilina, Galina B; Saunders, Thomas L; Yu, Xiaochun

    2010-03-16

    During spermatogenesis, global nucleosome removal occurs where histones are initially replaced by transition proteins and subsequently by protamines. This chromatin reorganization is thought to facilitate the compaction of the paternal genome into the sperm head and to protect the DNA from damaging agents. Histone ubiquitination has been suggested to be important for sex chromosome inactivation during meiotic prophase and nucleosome removal at postmeiotic stages. However, the mechanisms regulating these ubiquitin-mediated processes are unknown. In this study, we investigate the role of the ubiquitin ligase RNF8 during spermatogenesis and find that RNF8-deficient mice are proficient in meiotic sex chromosome inactivation (MSCI) but deficient in global nucleosome removal. Moreover, we show that RNF8-dependent histone ubiquitination induces H4K16 acetylation, which may be an initial step in nucleosome removal. Thus, our results show that RNF8 plays an important role during spermatogenesis through histone ubiquitination, resulting in trans-histone acetylation and global nucleosome removal.

  1. Quantification of histone modification ChIP-seq enrichment for data mining and machine learning applications

    Directory of Open Access Journals (Sweden)

    Bekiranov Stefan

    2011-08-01

    Full Text Available Abstract Background The advent of ChIP-seq technology has made the investigation of epigenetic regulatory networks a computationally tractable problem. Several groups have applied statistical computing methods to ChIP-seq datasets to gain insight into the epigenetic regulation of transcription. However, methods for estimating enrichment levels in ChIP-seq data for these computational studies are understudied and variable. Since the conclusions drawn from these data mining and machine learning applications strongly depend on the enrichment level inputs, a comparison of estimation methods with respect to the performance of statistical models should be made. Results Various methods were used to estimate the gene-wise ChIP-seq enrichment levels for 20 histone methylations and the histone variant H2A.Z. The Multivariate Adaptive Regression Splines (MARS algorithm was applied for each estimation method using the estimation of enrichment levels as predictors and gene expression levels as responses. The methods used to estimate enrichment levels included tag counting and model-based methods that were applied to whole genes and specific gene regions. These methods were also applied to various sizes of estimation windows. The MARS model performance was assessed with the Generalized Cross-Validation Score (GCV. We determined that model-based methods of enrichment estimation that spatially weight enrichment based on average patterns provided an improvement over tag counting methods. Also, methods that included information across the entire gene body provided improvement over methods that focus on a specific sub-region of the gene (e.g., the 5' or 3' region. Conclusion The performance of data mining and machine learning methods when applied to histone modification ChIP-seq data can be improved by using data across the entire gene body, and incorporating the spatial distribution of enrichment. Refinement of enrichment estimation ultimately improved accuracy

  2. Ice Cores

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Records of past temperature, precipitation, atmospheric trace gases, and other aspects of climate and environment derived from ice cores drilled on glaciers and ice...

  3. Core BPEL

    DEFF Research Database (Denmark)

    Hallwyl, Tim; Højsgaard, Espen

    extensions. Combined with the fact that the language definition does not provide a formal semantics, it is an arduous task to work formally with the language (e.g. to give an implementation). In this paper we identify a core subset of the language, called Core BPEL, which has fewer and simpler constructs......, does not allow omissions, and does not contain ignorable elements. We do so by identifying syntactic sugar, including default values, and ignorable elements in WS-BPEL. The analysis results in a translation from the full language to the core subset. Thus, we reduce the effort needed for working...... formally with WS-BPEL, as one, without loss of generality, need only consider the much simpler Core BPEL. This report may also be viewed as an addendum to the WS-BPEL standard specification, which clarifies the WS-BPEL syntax and presents the essential elements of the language in a more concise way...

  4. Core BPEL

    DEFF Research Database (Denmark)

    Hallwyl, Tim; Højsgaard, Espen

    extensions. Combined with the fact that the language definition does not provide a formal semantics, it is an arduous task to work formally with the language (e.g. to give an implementation). In this paper we identify a core subset of the language, called Core BPEL, which has fewer and simpler constructs......, does not allow omissions, and does not contain ignorable elements. We do so by identifying syntactic sugar, including default values, and ignorable elements in WS-BPEL. The analysis results in a translation from the full language to the core subset. Thus, we reduce the effort needed for working...... formally with WS-BPEL, as one, without loss of generality, need only consider the much simpler Core BPEL. This report may also be viewed as an addendum to the WS-BPEL standard specification, which clarifies the WS-BPEL syntax and presents the essential elements of the language in a more concise way...

  5. Core benefits

    National Research Council Canada - National Science Library

    Keith, Brian W

    2010-01-01

    This SPEC Kit explores the core employment benefits of retirement, and life, health, and other insurance -benefits that are typically decided by the parent institution and often have significant governmental regulation...

  6. Differential regulation of the histone chaperone HIRA during muscle cell differentiation by a phosphorylation switch.

    Science.gov (United States)

    Yang, Jae-Hyun; Song, Tae-Yang; Jo, Chanhee; Park, Jinyoung; Lee, Han-Young; Song, Ilang; Hong, Suji; Jung, Kwan Young; Kim, Jaehoon; Han, Jeung-Whan; Youn, Hong-Duk; Cho, Eun-Jung

    2016-08-12

    Replication-independent incorporation of variant histone H3.3 has a profound impact on chromatin function and numerous cellular processes, including the differentiation of muscle cells. The histone chaperone HIRA and H3.3 have essential roles in MyoD regulation during myoblast differentiation. However, the precise mechanism that determines the onset of H3.3 deposition in response to differentiation signals is unclear. Here we show that HIRA is phosphorylated by Akt kinase, an important signaling modulator in muscle cells. By generating a phosphospecific antibody, we found that a significant amount of HIRA was phosphorylated in myoblasts. The phosphorylation level of HIRA and the occupancy of phosphorylated protein on muscle genes gradually decreased during cellular differentiation. Remarkably, the forced expression of the phosphomimic form of HIRA resulted in reduced H3.3 deposition and suppressed the activation of muscle genes in myotubes. Our data show that HIRA phosphorylation limits the expression of myogenic genes, while the dephosphorylation of HIRA is required for proficient H3.3 deposition and gene activation, demonstrating that the phosphorylation switch is exploited to modulate HIRA/H3.3-mediated muscle gene regulation during myogenesis.

  7. Natural Diversity in Pentose Fermentation Is Explained by Variations in Histone Deacetylases

    Directory of Open Access Journals (Sweden)

    Zvi Tamari

    2016-01-01

    Full Text Available The extent to which carbon flux is directed toward fermentation versus respiration differs between cell types and environmental conditions. Understanding the basic cellular processes governing carbon flux is challenged by the complexity of the metabolic and regulatory networks. To reveal the genetic basis for natural diversity in channeling carbon flux, we applied quantitative trait loci analysis by phenotyping and genotyping hundreds of individual F2 segregants of budding yeast that differ in their capacity to ferment the pentose sugar xylulose. Causal alleles were mapped to the RXT3 and PHO23 genes, two components of the large Rpd3 histone deacetylation complex. We show that these allelic variants modulate the expression of SNF1/AMPK-dependent respiratory genes. Our results suggest that over close evolutionary distances, diversification of carbon flow is driven by changes in global regulators, rather than adaptation of specific metabolic nodes. Such regulators may improve the ability to direct metabolic fluxes for biotechnological applications.

  8. Mislocalization of the Drosophila centromere-specific histone CIDpromotes formation of functional ectopic kinetochores

    Energy Technology Data Exchange (ETDEWEB)

    Heun, Patrick; Erhardt, Sylvia; Blower, Michael D.; Weiss,Samara; Skora, Andrew D.; Karpen, Gary H.

    2006-01-30

    The centromere-specific histone variant CENP-A (CID in Drosophila) is a structural and functional foundation for kinetochore formation and chromosome segregation. Here, we show that overexpressed CID is mislocalized into normally non-centromeric regions in Drosophila tissue culture cells and animals. Analysis of mitoses in living and fixed cells reveals that mitotic delays, anaphase bridges, chromosome fragmentation, and cell and organismal lethality are all direct consequences of CID mislocalization. In addition, proteins that are normally restricted to endogenous kinetochores assemble at a subset of ectopic CID incorporation regions. The presence of microtubule motors and binding proteins, spindle attachments, and aberrant chromosome morphologies demonstrate that these ectopic kinetochores are functional. We conclude that CID mislocalization promotes formation of ectopic centromeres and multicentric chromosomes, which causes chromosome missegregation, aneuploidy, and growth defects. Thus, CENP-A mislocalization is one possible mechanism for genome instability during cancer progression, as well as centromere plasticity during evolution.

  9. EXPRESSION AND FUNCTIONAL INTERACTION OF CHICKEN RbAp46 POLYPEPTIDE WITH HISTONES, HISTONE DEACETYLASE-1, AND HISTONE ACETYLTRANSFERASE-1

    Directory of Open Access Journals (Sweden)

    Ahyar Ahmad

    2013-08-01

    Full Text Available In this study, we cloned and sequenced cDNA encoding the chicken p46 polypeptide, RbAp46. The cDNA encoding a protein consists of 424 amino acids is a member of the WD protein family, with seven WD repeat motifs, and exhibits 90.3% identity to RbAp48, and 94.3% identity to the human RbAp46. The RbAp46 fusion protein were synthesized by in vitro translation system and in Escherichia coli under induction by 50 µM IPTG and single step purified with glutathione-Agarose beads, showed that GST-tagged protein of approximately 72 kDa. The in vitro experiment established that RbAp46 interacts with chicken histones, chHDAC-1, and chHAT-1. The in vitro immunoprecipitation experiment, involving truncated mutants of RbAp46, revealed not only that two regions comprising amino acids 33-179 and 375-404 are necessary for its binding to H2B, but also that two regions comprising amino acids 1-32 and 405-424 are necessary for its binding to H4. Furthermore, the GST pulldown affinity assay, involving truncated mutants of RbAp46, revealed that a region comprising amino acids 359-404 binds to chHAT-1 in vitro. Taken together, these results indicate not only that RbAp46 should participate differentially in a number of DNA-utilizing processes through interactions of its distinct regions with histones and chHAT-1, but also that the proper propeller structure of RbAp46 is not necessary for its interaction with chHAT-1.

  10. Recombinant thrombomodulin protects mice against histone-induced lethal thromboembolism.

    Directory of Open Access Journals (Sweden)

    Mayumi Nakahara

    Full Text Available INTRODUCTION: Recent studies have shown that histones, the chief protein component of chromatin, are released into the extracellular space during sepsis, trauma, and ischemia-reperfusion injury, and act as major mediators of the death of an organism. This study was designed to elucidate the cellular and molecular basis of histone-induced lethality and to assess the protective effects of recombinant thrombomodulin (rTM. rTM has been approved for the treatment of disseminated intravascular coagulation (DIC in Japan, and is currently undergoing a phase III clinical trial in the United States. METHODS: Histone H3 levels in plasma of healthy volunteers and patients with sepsis and DIC were measured using enzyme-linked immunosorbent assay. Male C57BL/6 mice were injected intravenously with purified histones, and pathological examinations were performed. The protective effects of rTM against histone toxicity were analyzed both in vitro and in mice. RESULTS: Histone H3 was not detectable in plasma of healthy volunteers, but significant levels were observed in patients with sepsis and DIC. These levels were higher in non-survivors than in survivors. Extracellular histones triggered platelet aggregation, leading to thrombotic occlusion of pulmonary capillaries and subsequent right-sided heart failure in mice. These mice displayed symptoms of DIC, including thrombocytopenia, prolonged prothrombin time, decreased fibrinogen, fibrin deposition in capillaries, and bleeding. Platelet depletion protected mice from histone-induced death in the first 30 minutes, suggesting that vessel occlusion by platelet-rich thrombi might be responsible for death during the early phase. Furthermore, rTM bound to extracellular histones, suppressed histone-induced platelet aggregation, thrombotic occlusion of pulmonary capillaries, and dilatation of the right ventricle, and rescued mice from lethal thromboembolism. CONCLUSIONS: Extracellular histones cause massive

  11. Histone deacetylase inhibitors (HDACIs: multitargeted anticancer agents

    Directory of Open Access Journals (Sweden)

    Ververis K

    2013-02-01

    Full Text Available Katherine Ververis,1 Alison Hiong,1 Tom C Karagiannis,1,* Paul V Licciardi2,*1Epigenomic Medicine, Alfred Medical Research and Education Precinct, 2Allergy and Immune Disorders, Murdoch Childrens Research Institute, Melbourne, VIC, Australia*These authors contributed equally to this workAbstract: Histone deacetylase (HDAC inhibitors are an emerging class of therapeutics with potential as anticancer drugs. The rationale for developing HDAC inhibitors (and other chromatin-modifying agents as anticancer therapies arose from the understanding that in addition to genetic mutations, epigenetic changes such as dysregulation of HDAC enzymes can alter phenotype and gene expression, disturb homeostasis, and contribute to neoplastic growth. The family of HDAC inhibitors is large and diverse. It includes a range of naturally occurring and synthetic compounds that differ in terms of structure, function, and specificity. HDAC inhibitors have multiple cell type-specific effects in vitro and in vivo, such as growth arrest, cell differentiation, and apoptosis in malignant cells. HDAC inhibitors have the potential to be used as monotherapies or in combination with other anticancer therapies. Currently, there are two HDAC inhibitors that have received approval from the US FDA for the treatment of cutaneous T-cell lymphoma: vorinostat (suberoylanilide hydroxamic acid, Zolinza and depsipeptide (romidepsin, Istodax. More recently, depsipeptide has also gained FDA approval for the treatment of peripheral T-cell lymphoma. Many more clinical trials assessing the effects of various HDAC inhibitors on hematological and solid malignancies are currently being conducted. Despite the proven anticancer effects of particular HDAC inhibitors against certain cancers, many aspects of HDAC enzymes and HDAC inhibitors are still not fully understood. Increasing our understanding of the effects of HDAC inhibitors, their targets and mechanisms of action will be critical for the

  12. Migraine Variants And Beyond

    Directory of Open Access Journals (Sweden)

    Chakravarty A

    2002-01-01

    Full Text Available The Classic presenting features of both migraine with and without aura have been clearly defined. Occasionally however migrainous headaches are accompanied by abrupt appearance of focal and ominous neurological signs. Such attacks can be labelled as migraine variants and the diagnosis in reality is one made by exclusion of other CNS diseases. Some but not all such conditions are mentioned in the International Headache Society (IHS classification under the general heading of migraine with aura. Rarely, the focal neurological deficit may outlast the migraine attack by days and occasionally with appearance of structural brain lesions on neuroimaging. Such attacks have been labelled as complicated Migraine by the IHS. The present review deal with the clinical, radiologic and pathophysiologic aspects of both these conditions - migraine variants and complicated migraine.

  13. The Clinical Significance of Unknown Sequence Variants in BRCA Genes

    Energy Technology Data Exchange (ETDEWEB)

    Calò, Valentina; Bruno, Loredana; Paglia, Laura La; Perez, Marco; Margarese, Naomi [Department of Surgery and Oncology, Regional Reference Center for the Biomolecular Characterization and Genetic Screening of Hereditary Tumors, University of Palermo, Via del Vespro 127, 90127 Palermo (Italy); Gaudio, Francesca Di [Department of Medical Biotechnologies and Legal Medicine, University of Palermo, Palermo (Italy); Russo, Antonio, E-mail: lab-oncobiologia@usa.net [Department of Surgery and Oncology, Regional Reference Center for the Biomolecular Characterization and Genetic Screening of Hereditary Tumors, University of Palermo, Via del Vespro 127, 90127 Palermo (Italy)

    2010-09-10

    Germline mutations in BRCA1/2 genes are responsible for a large proportion of hereditary breast and/or ovarian cancers. Many highly penetrant predisposition alleles have been identified and include frameshift or nonsense mutations that lead to the translation of a truncated protein. Other alleles contain missense mutations, which result in amino acid substitution and intronic variants with splicing effect. The discovery of variants of uncertain/unclassified significance (VUS) is a result that can complicate rather than improve the risk assessment process. VUSs are mainly missense mutations, but also include a number of intronic variants and in-frame deletions and insertions. Over 2,000 unique BRCA1 and BRCA2 missense variants have been identified, located throughout the whole gene (Breast Cancer Information Core Database (BIC database)). Up to 10–20% of the BRCA tests report the identification of a variant of uncertain significance. There are many methods to discriminate deleterious/high-risk from neutral/low-risk unclassified variants (i.e., analysis of the cosegregation in families of the VUS, measure of the influence of the VUSs on the wild-type protein activity, comparison of sequence conservation across multiple species), but only an integrated analysis of these methods can contribute to a real interpretation of the functional and clinical role of the discussed variants. The aim of our manuscript is to review the studies on BRCA VUS in order to clarify their clinical relevance.

  14. Variants of Uncertainty

    Science.gov (United States)

    1981-05-15

    Variants of Uncertainty Daniel Kahneman University of British Columbia Amos Tversky Stanford University DTI-C &%E-IECTE ~JUNO 1i 19 8 1j May 15, 1981... Dennett , 1979) in which different parts have ac- cess to different data, assign then different weights and hold different views of the situation...2robable and t..h1 provable. Oxford- Claredor Press, 1977. Dennett , D.C. Brainstorms. Hassocks: Harvester, 1979. Donchin, E., Ritter, W. & McCallum, W.C

  15. Variants of glycoside hydrolases

    Energy Technology Data Exchange (ETDEWEB)

    Teter, Sarah; Ward, Connie; Cherry, Joel; Jones, Aubrey; Harris, Paul; Yi, Jung

    2017-07-11

    The present invention relates to variants of a parent glycoside hydrolase, comprising a substitution at one or more positions corresponding to positions 21, 94, 157, 205, 206, 247, 337, 350, 373, 383, 438, 455, 467, and 486 of amino acids 1 to 513 of SEQ ID NO: 2, and optionally further comprising a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2 a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2, wherein the variants have glycoside hydrolase activity. The present invention also relates to nucleotide sequences encoding the variant glycoside hydrolases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  16. Variants of glycoside hydrolases

    Energy Technology Data Exchange (ETDEWEB)

    Teter, Sarah (Davis, CA); Ward, Connie (Hamilton, MT); Cherry, Joel (Davis, CA); Jones, Aubrey (Davis, CA); Harris, Paul (Carnation, WA); Yi, Jung (Sacramento, CA)

    2011-04-26

    The present invention relates to variants of a parent glycoside hydrolase, comprising a substitution at one or more positions corresponding to positions 21, 94, 157, 205, 206, 247, 337, 350, 373, 383, 438, 455, 467, and 486 of amino acids 1 to 513 of SEQ ID NO: 2, and optionally further comprising a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2 a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2, wherein the variants have glycoside hydrolase activity. The present invention also relates to nucleotide sequences encoding the variant glycoside hydrolases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  17. DNA methylation affects nuclear organization, histone modifications, and linker histone binding but not chromatin compaction.

    Science.gov (United States)

    Gilbert, Nick; Thomson, Inga; Boyle, Shelagh; Allan, James; Ramsahoye, Bernard; Bickmore, Wendy A

    2007-05-07

    DNA methylation has been implicated in chromatin condensation and nuclear organization, especially at sites of constitutive heterochromatin. How this is mediated has not been clear. In this study, using mutant mouse embryonic stem cells completely lacking in DNA methylation, we show that DNA methylation affects nuclear organization and nucleosome structure but not chromatin compaction. In the absence of DNA methylation, there is increased nuclear clustering of pericentric heterochromatin and extensive changes in primary chromatin structure. Global levels of histone H3 methylation and acetylation are altered, and there is a decrease in the mobility of linker histones. However, the compaction of both bulk chromatin and heterochromatin, as assayed by nuclease digestion and sucrose gradient sedimentation, is unaltered by the loss of DNA methylation. This study shows how the complete loss of a major epigenetic mark can have an impact on unexpected levels of chromatin structure and nuclear organization and provides evidence for a novel link between DNA methylation and linker histones in the regulation of chromatin structure.

  18. Dysregulation of Histone Acetyltransferases and Deacetylases in Cardiovascular Diseases

    Directory of Open Access Journals (Sweden)

    Yonggang Wang

    2014-01-01

    Full Text Available Cardiovascular disease (CVD remains a leading cause of mortality worldwide despite advances in its prevention and management. A comprehensive understanding of factors which contribute to CVD is required in order to develop more effective treatment options. Dysregulation of epigenetic posttranscriptional modifications of histones in chromatin is thought to be associated with the pathology of many disease models, including CVD. Histone acetyltransferases (HATs and deacetylases (HDACs are regulators of histone lysine acetylation. Recent studies have implicated a fundamental role of reversible protein acetylation in the regulation of CVDs such as hypertension, pulmonary hypertension, diabetic cardiomyopathy, coronary artery disease, arrhythmia, and heart failure. This reversible acetylation is governed by enzymes that HATs add or HDACs remove acetyl groups respectively. New evidence has revealed that histone acetylation regulators blunt cardiovascular and related disease states in certain cellular processes including myocyte hypertrophy, apoptosis, fibrosis, oxidative stress, and inflammation. The accumulating evidence of the detrimental role of histone acetylation in cardiac disease combined with the cardioprotective role of histone acetylation regulators suggests that the use of histone acetylation regulators may serve as a novel approach to treating the millions of patients afflicted by cardiac diseases worldwide.

  19. Histone Acylation beyond Acetylation: Terra Incognita in Chromatin Biology

    Directory of Open Access Journals (Sweden)

    Sophie Rousseaux

    2015-04-01

    Full Text Available Histone acetylation, one of the first and best studied histone post-translational modifications (PTMs, as well as the factors involved in its deposition (writers, binding (readers and removal (erasers, have been shown to act at the heart of regulatory circuits controlling essential cellular functions. The identification of a variety of competing histone lysine-modifying acyl groups including propionyl, butyryl, 2-hydroxyisobutyryl, crotonyl, malonyl, succinyl and glutaryl, raises numerous questions on their functional significance, the molecular systems that manage their establishment, removal and interplay with the well-known acetylation-based mechanisms. Detailed and large-scale investigations of two of these new histone PTMs, crotonylation and 2-hydroxyisobutyrylation, along with histone acetylation, in the context of male genome programming, where stage-specific gene expression programs are switched on and off in turn, have shed light on their functional contribution to the epigenome for the first time. These initial investigations fired many additional questions, which remain to be explored. This review surveys the major results taken from these two new histone acylations and discusses the new biology that is emerging based on the diversity of histone lysine acylations.

  20. PTEN Interacts with Histone H1 and Controls Chromatin Condensation

    Directory of Open Access Journals (Sweden)

    Zhu Hong Chen

    2014-09-01

    Full Text Available Chromatin organization and dynamics are integral to global gene transcription. Histone modification influences chromatin status and gene expression. PTEN plays multiple roles in tumor suppression, development, and metabolism. Here, we report on the interplay of PTEN, histone H1, and chromatin. We show that loss of PTEN leads to dissociation of histone H1 from chromatin and decondensation of chromatin. PTEN deletion also results in elevation of histone H4 acetylation at lysine 16, an epigenetic marker for chromatin activation. We found that PTEN and histone H1 physically interact through their C-terminal domains. Disruption of the PTEN C terminus promotes the chromatin association of MOF acetyltransferase and induces H4K16 acetylation. Hyperacetylation of H4K16 impairs the association of PTEN with histone H1, which constitutes regulatory feedback that may reduce chromatin stability. Our results demonstrate that PTEN controls chromatin condensation, thus influencing gene expression. We propose that PTEN regulates global gene transcription profiling through histones and chromatin remodeling.

  1. Nucleosomes, Linker DNA, and Linker Histone form a Unique Structural Motif that Directs the Higher-Order Folding and Compaction of Chromatin

    Science.gov (United States)

    Bednar, Jan; Horowitz, Rachel A.; Grigoryev, Sergei A.; Carruthers, Lenny M.; Hansen, Jeffrey C.; Koster, Abraham J.; Woodcock, Christopher L.

    1998-11-01

    The compaction level of arrays of nucleosomes may be understood in terms of the balance between the self-repulsion of DNA (principally linker DNA) and countering factors including the ionic strength and composition of the medium, the highly basic N termini of the core histones, and linker histones. However, the structural principles that come into play during the transition from a loose chain of nucleosomes to a compact 30-nm chromatin fiber have been difficult to establish, and the arrangement of nucleosomes and linker DNA in condensed chromatin fibers has never been fully resolved. Based on images of the solution conformation of native chromatin and fully defined chromatin arrays obtained by electron cryomicroscopy, we report a linker histone-dependent architectural motif beyond the level of the nucleosome core particle that takes the form of a stem-like organization of the entering and exiting linker DNA segments. DNA completes ≈ 1.7 turns on the histone octamer in the presence and absence of linker histone. When linker histone is present, the two linker DNA segments become juxtaposed ≈ 8 nm from the nucleosome center and remain apposed for 3-5 nm before diverging. We propose that this stem motif directs the arrangement of nucleosomes and linker DNA within the chromatin fiber, establishing a unique three-dimensional zigzag folding pattern that is conserved during compaction. Such an arrangement with peripherally arranged nucleosomes and internal linker DNA segments is fully consistent with observations in intact nuclei and also allows dramatic changes in compaction level to occur without a concomitant change in topology.

  2. Two distinct modes for propagation of histone PTMs across the cell cycle

    DEFF Research Database (Denmark)

    Alabert, Constance; Barth, Teresa K; Reverón-Gómez, Nazaret;

    2015-01-01

    of new histone deposition. Importantly, within one cell cycle, all PTMs are restored. In general, new histones are modified to mirror the parental histones. However, H3K9 trimethylation (H3K9me3) and H3K27me3 are propagated by continuous modification of parental and new histones because the establishment...

  3. Regulation of Insulin Gene Transcription by Multiple Histone Acetyltransferases

    OpenAIRE

    2012-01-01

    Glucose-stimulated insulin gene transcription is mainly regulated by a 340-bp promoter region upstream of the transcription start site by beta-cell-enriched transcription factors Pdx-1, MafA, and NeuroD1. Previous studies have shown that histone H4 hyperacetylation is important for acute up-regulation of insulin gene transcription. Until now, only the histone acetyltransferase (HAT) protein p300 has been shown to be involved in this histone H4 acetylation event. In this report we investigated...

  4. Protein-protein Förster resonance energy transfer analysis of nucleosome core particles containing H2A and H2A.Z.

    Science.gov (United States)

    Hoch, Duane A; Stratton, Jessica J; Gloss, Lisa M

    2007-08-24

    A protein-protein Förster resonance energy transfer (FRET) system, employing probes at multiple positions, was designed to specifically monitor the dissociation of the H2A-H2B dimer from the nucleosome core particle (NCP). Tryptophan donors and Cys-AEDANS acceptors were chosen because, compared to previous NCP FRET fluorophores, they: (1) are smaller and less hydrophobic, which should minimize perturbations of histone and NCP structure; and (2) have an R0 of 20 A, which is much less than the dimensions of the NCP (approximately 50 A width and approximately 100 A diameter). Equilibrium protein unfolding titrations indicate that the donor and acceptor moieties have minimal effects on the stability of the H2A-H2B dimer and (H3-H4)2 tetramer. NCPs containing the various FRET pairs were reconstituted with the 601 DNA positioning element. Equilibrium NaCl-induced dissociation of the modified NCPs showed that the 601 sequence stabilized the NCP to dimer dissociation relative to weaker positioning sequences. This finding implies a significant role for the H2A-H2B dimers in determining the DNA sequence dependence of NCP stability. The free energy of dissociation determined from reversible and well-defined sigmoidal transitions revealed two distinct phases reflecting the dissociation of individual H2A-H2B dimers, confirming cooperativity as suggested previously; these data allow quantitative description of the cooperativity. The FRET system was then used to study the effects of the histone variant H2A.Z on NCP stability; previous studies have reported both destabilizing and stabilizing effects. H2A.Z FRET NCP dissociation transitions suggest a slight increase in stability but a significant increase in cooperativity of the dimer dissociations. Thus, the utility of this protein-protein FRET system to monitor the effects of histone variants on NCP dynamics has been demonstrated, and the system appears equally well-suited for dissection of the kinetic processes of dimer

  5. Profiling post-translational modifications of histones in neural differentiation of embryonic stem cells using liquid chromatography-mass spectrometry.

    Science.gov (United States)

    Zheng, Shuzhen; Sun, Ming; Zhang, Kai; Gu, Junjie; Guo, Zhenchang; Tian, Shanshan; Zhai, Guijin; He, Xiwen; Jin, Ying; Zhang, Yukui

    2016-04-01

    The neural differentiation of embryonic stem cells (ESCs) is of great significance for understanding of the mechanism of diseases. Histone post-translational modifications (HPTMs) play a key role in the regulation of ESCs differentiation. Here, we combined the stable isotope chemical derivatization with nano-HPLC-mass spectrometry (MS) for comprehensive analysis and quantification of histone post-translational modifications (HPTMs) in mouse embryonic stem cells (mESCs) and neural progenitor cells (mNPCs) that was derived from ESCs. We identified 85 core HPTM sites in ESCs and 78HPTM sites in NPCs including some novel lysine modifications. Our quantitative analysis results further revealed the changes of HPTMs from ESCs to NPCs and suggested effect of combinational HPTMs in the differentiation. This study demonstrates that HPLC-MS-based quantitative proteomics has a considerable advantage on quantification of combinational PTMs and expands our understanding of HPTMs in the differentiation.

  6. Histone chaperone spt16 promotes redeposition of the original h3-h4 histones evicted by elongating RNA polymerase.

    Science.gov (United States)

    Jamai, Adil; Puglisi, Andrea; Strubin, Michel

    2009-08-14

    Nucleosomes are surprisingly dynamic structures in vivo, showing transcription-independent exchange of histones H2A-H2B genome-wide and exchange of H3-H4 mainly within the promoters of transcribed genes. In addition, nucleosomes are disrupted in front of and reassembled behind the elongating RNA polymerase. Here we show that inactivation of histone chaperone Spt16 in yeast results in rapid loss of H2B and H3 from transcribed genes but also from inactive genes. In all cases, histone loss is blocked by a transcription inhibitor, indicating a transcription-dependent event. Thus, nucleosomes are efficiently evicted by the polymerase but do not reform in the absence of Spt16. Yet exchange of nucleosomal H2B with free histones occurs normally, and, unexpectedly, incorporation of new H3 increases at all loci tested. This points to Spt16 restoring normal nucleosome structure by redepositing the displaced H3-H4 histones, thereby preventing incorporation of new histones and perhaps changes in histone modification patterns associated with ongoing transcription.

  7. The docking domain of histone H2A is required for H1 binding and RSC-mediated nucleosome remodeling.

    Science.gov (United States)

    Shukla, Manu Shubhdarshan; Syed, Sajad Hussain; Goutte-Gattat, Damien; Richard, John Lalith Charles; Montel, Fabien; Hamiche, Ali; Travers, Andrew; Faivre-Moskalenko, Cendrine; Bednar, Jan; Hayes, Jeffrey J; Angelov, Dimitar; Dimitrov, Stefan

    2011-04-01

    Histone variants within the H2A family show high divergences in their C-terminal regions. In this work, we have studied how these divergences and in particular, how a part of the H2A COOH-terminus, the docking domain, is implicated in both structural and functional properties of the nucleosome. Using biochemical methods in combination with Atomic Force Microscopy and Electron Cryo-Microscopy, we show that the H2A-docking domain is a key structural feature within the nucleosome. Deletion of this domain or replacement with the incomplete docking domain from the variant H2A.Bbd results in significant structural alterations in the nucleosome, including an increase in overall accessibility to nucleases, un-wrapping of ∼10 bp of DNA from each end of the nucleosome and associated changes in the entry/exit angle of DNA ends. These structural alterations are associated with a reduced ability of the chromatin remodeler RSC to both remodel and mobilize the nucleosomes. Linker histone H1 binding is also abrogated in nucleosomes containing the incomplete docking domain of H2A.Bbd. Our data illustrate the unique role of the H2A-docking domain in coordinating the structural-functional aspects of the nucleosome properties. Moreover, our data suggest that incorporation of a 'defective' docking domain may be a primary structural role of H2A.Bbd in chromatin.

  8. The HIRA complex that deposits the histone H3.3 is conserved in Arabidopsis and facilitates transcriptional dynamics

    Directory of Open Access Journals (Sweden)

    Xin Nie

    2014-08-01

    Full Text Available In animals, replication-independent incorporation of nucleosomes containing the histone variant H3.3 enables global reprogramming of histone modifications and transcriptional profiles. H3.3 enrichment over gene bodies correlates with gene transcription in animals and plants. In animals, H3.3 is deposited into chromatin by specific protein complexes, including the HIRA complex. H3.3 variants evolved independently and acquired similar properties in animals and plants, questioning how the H3.3 deposition machinery evolved in plants and what are its biological functions. We performed phylogenetic analyses in the plant kingdom and identified in Arabidopsis all orthologs of human genes encoding members of the HIRA complex. Genetic analyses, biochemical data and protein localisation suggest that these proteins form a complex able to interact with H3.3 in Arabidopsis in a manner similar to that described in mammals. In contrast to animals, where HIRA is required for fertilization and early development, loss of function of HIRA in Arabidopsis causes mild phenotypes in the adult plant and does not perturb sexual reproduction and embryogenesis. Rather, HIRA function is required for transcriptional reprogramming during dedifferentiation of plant cells that precedes vegetative propagation and for the appropriate transcription of genes responsive to biotic and abiotic factors. We conclude that the molecular function of the HIRA complex is conserved between plants and animals. Yet plants diversified HIRA functions to enable asexual reproduction and responsiveness to the environment in response to the plant sessile lifestyle.

  9. Features of the PHF8/KIAA1718 histone demethylase

    Institute of Scientific and Technical Information of China (English)

    Tamaki Suganuma; Jerry L Workman

    2010-01-01

    Gene regulation mechanisms in cellular events ranging from development to tumorigenesis target the fundamental unit of chromatin structure, the nucleosome, 146 bp of DNA wrapped around a histone octamer.

  10. Substrate Recognition of Histone H2B by DUBm

    Science.gov (United States)

    Henderson, Elizabeth; Berndsen, Christopher; Wolberger, Cynthia

    2011-03-01

    The SAGA complex is a transcriptional coactivator that regulates gene expression in eukaryotes via histone acetylation and deubiquitination, which are crucial for transcription. Our lab is investigating the SAGA-dependent deubiquitination of histone H2B. The deubiquitinating module (DUBm) of SAGA is comprised of a ubiquitin-specific protease, Ubp8, and three other proteins. It is known that Ubp8 cleaves ubiquitin from histone H2B, however, the specific way in which the enzyme binds to the substrate remains elusive. In order to unravel this mechanism, we attempted to determine the crystal structure of the substrate binding complex. We obtained this substrate by exploiting the techniques of intein chemistry to artificially ubiquitinate a histone H2B peptide, which we then co-crystallized with DUBm. Additionally, we synthesized Ub-K63R-linked chains and Ub-K48-linked chains and co-crystallized them with DUBm.

  11. Reshaping chromatin after DNA damage: the choreography of histone proteins.

    Science.gov (United States)

    Polo, Sophie E

    2015-02-13

    DNA damage signaling and repair machineries operate in a nuclear environment where DNA is wrapped around histone proteins and packaged into chromatin. Understanding how chromatin structure is restored together with the DNA sequence during DNA damage repair has been a topic of intense research. Indeed, chromatin integrity is central to cell functions and identity. However, chromatin shows remarkable plasticity in response to DNA damage. This review presents our current knowledge of chromatin dynamics in the mammalian cell nucleus in response to DNA double strand breaks and UV lesions. I provide an overview of the key players involved in regulating histone dynamics in damaged chromatin regions, focusing on histone chaperones and their concerted action with histone modifiers, chromatin remodelers and repair factors. I also discuss how these dynamics contribute to reshaping chromatin and, by altering the chromatin landscape, may affect the maintenance of epigenetic information.

  12. Interplay between histone H1 structure and function.

    Science.gov (United States)

    Roque, Alicia; Ponte, Inma; Suau, Pedro

    2016-03-01

    H1 linker histones are involved both in the maintenance of higher-order chromatin structure and in gene regulation. Histone H1 exists in multiple isoforms, is evolutionarily variable and undergoes a large variety of post-translational modifications. We review recent progress in the understanding of the folding and structure of histone H1 domains with an emphasis on the interactions with DNA. The importance of intrinsic disorder and hydrophobic interactions in the folding and function of the carboxy-terminal domain (CTD) is discussed. The induction of a molten globule-state in the CTD by macromolecular crowding is also considered. The effects of phosphorylation by cyclin-dependent kinases on the structure of the CTD, as well as on chromatin condensation and oligomerization, are described. We also address the extranuclear functions of histone H1, including the interaction with the β-amyloid peptide. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Histone modifications induced by a family of bacterial toxins.

    Science.gov (United States)

    Hamon, Mélanie Anne; Batsché, Eric; Régnault, Béatrice; Tham, To Nam; Seveau, Stéphanie; Muchardt, Christian; Cossart, Pascale

    2007-08-14

    Upon infection, pathogens reprogram host gene expression. In eukaryotic cells, genetic reprogramming is induced by the concerted activation/repression of transcription factors and various histone modifications that control DNA accessibility in chromatin. We report here that the bacterial pathogen Listeria monocytogenes induces a dramatic dephosphorylation of histone H3 as well as a deacetylation of histone H4 during early phases of infection. This effect is mediated by the major listerial toxin listeriolysin O in a pore-forming-independent manner. Strikingly, a similar effect also is observed with other toxins of the same family, such as Clostridium perfringens perfringolysin and Streptococcus pneumoniae pneumolysin. The decreased levels of histone modifications correlate with a reduced transcriptional activity of a subset of host genes, including key immunity genes. Thus, control of epigenetic regulation emerges here as an unsuspected function shared by several bacterial toxins, highlighting a common strategy used by intracellular and extracellular pathogens to modulate the host response early during infection.

  14. A histone demethylase is necessary for regeneration in zebrafish.

    Science.gov (United States)

    Stewart, Scott; Tsun, Zhi-Yang; Izpisua Belmonte, Juan Carlos

    2009-11-24

    Urodele amphibians and teleost fish regenerate amputated body parts via a process called epimorphic regeneration. A hallmark of this phenomenon is the reactivation of silenced developmental regulatory genes that previously functioned during embryonic patterning. We demonstrate that histone modifications silence promoters of numerous genes involved in zebrafish caudal fin regeneration. Silenced developmental regulatory genes contain bivalent me(3)K4/me(3)K27 H3 histone modifications created by the concerted action of Polycomb (PcG) and Trithorax histone methyltransferases. During regeneration, this silent, bivalent chromatin is converted to an active state by loss of repressive me(3)K27 H3 modifications, occurring at numerous genes that appear to function during regeneration. Loss-of-function studies demonstrate a requirement for a me(3)K27 H3 demethylase during fin regeneration. These results indicate that histone modifications at discreet genomic positions may serve as a crucial regulatory event in the initiation of fin regeneration.

  15. Targeting Histone Abnormality in Triple Negative Breast Cancer

    Science.gov (United States)

    2015-08-01

    respiration by 17β-estradiol through lactate dehydrogenase in MCF7 breast cancer cells. AACR Metabolism and Cancer , Baltimore, October 16-19, 2011. 20...1 AWARD NUMBER: W81XWH-14-1-0237 TITLE: Targeting Histone Abnormality in Triple-Negative Breast Cancer PRINCIPAL INVESTIGATOR: Yi...TITLE AND SUBTITLE 5a. CONTRACT NUMBER Targeting Histone Abnormality in Triple-Negative Breast Cancer 5b. GRANT NUMBER W81XWH-14-1-0237 5c

  16. The nonhistone, N-terminal tail of an essential, chimeric H2A variant regulates mitotic H3-S10 dephosphorylation.

    Science.gov (United States)

    Song, Xiaoyuan; Bowen, Josephine; Miao, Wei; Liu, Yifan; Gorovsky, Martin A

    2012-03-15

    H2A.Y is an essential, divergent Tetrahymena thermophila histone variant. It has a long nonhistone N terminus that contains leucine-rich repeats (LRR) and an LRR cap domain with similarity to Sds22p, a regulator of yeast protein phosphatase 1 (PP1) activity in the nucleus. In growing cells, H2A.Y is incorporated into micronuclei only during S phase, which occurs immediately after micronuclear mitosis. Depletion of H2A.Y causes prolonged retention of mitosis-associated histone H3-S10 phosphorylation and mitotic abnormalities that mimic S10E mutation. In cells where H2A.Y is depleted, an inducible chimeric gene, in which the H2A.Y N terminus is attached to H2A.X, is shown to regulate micronuclear H3-S10 phosphorylation. H2A.Y can also be specifically coimmunoprecipitated with a Tetrahymena PP1 ortholog (Ppo1p). Taken together, these results argue that the N terminus of H2A.Y functions to regulate H3-S10 dephosphorylation. This striking in vivo case of "cross-talk" between a H2A variant and a specific post-translational modification of another histone demonstrates a novel function for a histone variant.

  17. Eukaryotic Replisome Components Cooperate to Process Histones During Chromosome Replication

    Directory of Open Access Journals (Sweden)

    Magdalena Foltman

    2013-03-01

    Full Text Available DNA unwinding at eukaryotic replication forks displaces parental histones, which must be redeposited onto nascent DNA in order to preserve chromatin structure. By screening systematically for replisome components that pick up histones released from chromatin into a yeast cell extract, we found that the Mcm2 helicase subunit binds histones cooperatively with the FACT (facilitiates chromatin transcription complex, which helps to re-establish chromatin during transcription. FACT does not associate with the Mcm2-7 helicase at replication origins during G1 phase but is subsequently incorporated into the replisome progression complex independently of histone binding and uniquely among histone chaperones. The amino terminal tail of Mcm2 binds histones via a conserved motif that is dispensable for DNA synthesis per se but helps preserve subtelomeric chromatin, retain the 2 micron minichromosome, and support growth in the absence of Ctf18-RFC. Our data indicate that the eukaryotic replication and transcription machineries use analogous assemblies of multiple chaperones to preserve chromatin integrity.

  18. Mechanism of histone survival during transcription by RNA polymerase II.

    Science.gov (United States)

    Kulaeva, Olga I; Studitsky, Vasily M

    2010-01-01

    This work is related to and stems from our recent NSMB paper, "Mechanism of chromatin remodeling and recovery during passage of RNA polymerase II" (December 2009). Synopsis. Recent genomic studies from many laboratories have suggested that nucleosomes are not displaced from moderately transcribed genes. Furthermore, histones H3/H4 carrying the primary epigenetic marks are not displaced or exchanged (in contrast to H2A/H2B histones) during moderate transcription by RNA polymerase II (Pol II) in vivo. These exciting observations suggest that the large molecule of Pol II passes through chromatin structure without even transient displacement of H3/H4 histones. The most recent analysis of the RNA polymerase II (Pol II)-type mechanism of chromatin remodeling in vitro (described in our NSMB 2009 paper) suggests that nucleosome survival is tightly coupled with formation of a novel intermediate: a very small intranucleosomal DNA loop (Ø-loop) containing transcribing Pol II. In the submitted manuscript we critically evaluate one of the key predictions of this model: the lack of even transient displacement of histones H3/H4 during Pol II transcription in vitro. The data suggest that, indeed, histones H3/H4 are not displaced during Pol II transcription in vitro. These studies are directly connected with the observation in vivo on the lack of exchange of histones H3/H4 during Pol II transcription.

  19. Histone Methylation by Temozolomide; A Classic DNA Methylating Anticancer Drug

    Science.gov (United States)

    Pickard, Amanda J.; Diaz, Anthony Joseph; Mura, Hugo; Nyuwen, Lila; Coello, Daniel; Sheva, Saif; Maria, Nava; Gallo, James M.; Wang, Tieli

    2017-01-01

    Background/Aim The alkylating agent, temozolomide (TMZ), is considered the standard-of-care for high-grade astrocytomas –known as glioblastoma multiforme (GBM)– an aggressive type of tumor with poor prognosis. The therapeutic benefit of TMZ is attributed to formation of DNA adducts involving the methylation of purine bases in DNA. We investigated the effects of TMZ on arginine and lysine amino acids, histone H3 peptides and histone H3 proteins. Materials and Methods Chemical modification of amino acids, histone H3 peptide and protein by TMZ was performed in phosphate buffer at physiological pH. The reaction products were examined by mass spectrometry and western blot analysis. Results Our results showed that TMZ following conversion to a methylating cation, can methylate histone H3 peptide and histone H3 protein, suggesting that TMZ exerts its anticancer activity not only through its interaction with DNA, but also through alterations of protein post-translational modifications. Conclusion The possibility that TMZ can methylate histones involved with epigenetic regulation of protein indicates a potentially unique mechanism of action. The study will contribute to the understanding the anticancer activity of TMZ in order to develop novel targeted molecular strategies to advance the cancer treatment. PMID:27354585

  20. Structural Mechanisms of Nucleosome Recognition by Linker Histones.

    Science.gov (United States)

    Zhou, Bing-Rui; Jiang, Jiansheng; Feng, Hanqiao; Ghirlando, Rodolfo; Xiao, T Sam; Bai, Yawen

    2015-08-20

    Linker histones bind to the nucleosome and regulate the structure of chromatin and gene expression. Despite more than three decades of effort, the structural basis of nucleosome recognition by linker histones remains elusive. Here, we report the crystal structure of the globular domain of chicken linker histone H5 in complex with the nucleosome at 3.5 Å resolution, which is validated using nuclear magnetic resonance spectroscopy. The globular domain sits on the dyad of the nucleosome and interacts with both DNA linkers. Our structure integrates results from mutation analyses and previous cross-linking and fluorescence recovery after photobleach experiments, and it helps resolve the long debate on structural mechanisms of nucleosome recognition by linker histones. The on-dyad binding mode of the H5 globular domain is different from the recently reported off-dyad binding mode of Drosophila linker histone H1. We demonstrate that linker histones with different binding modes could fold chromatin to form distinct higher-order structures. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Eukaryotic replisome components cooperate to process histones during chromosome replication.

    Science.gov (United States)

    Foltman, Magdalena; Evrin, Cecile; De Piccoli, Giacomo; Jones, Richard C; Edmondson, Rick D; Katou, Yuki; Nakato, Ryuichiro; Shirahige, Katsuhiko; Labib, Karim

    2013-03-28

    DNA unwinding at eukaryotic replication forks displaces parental histones, which must be redeposited onto nascent DNA in order to preserve chromatin structure. By screening systematically for replisome components that pick up histones released from chromatin into a yeast cell extract, we found that the Mcm2 helicase subunit binds histones cooperatively with the FACT (facilitiates chromatin transcription) complex, which helps to re-establish chromatin during transcription. FACT does not associate with the Mcm2-7 helicase at replication origins during G1 phase but is subsequently incorporated into the replisome progression complex independently of histone binding and uniquely among histone chaperones. The amino terminal tail of Mcm2 binds histones via a conserved motif that is dispensable for DNA synthesis per se but helps preserve subtelomeric chromatin, retain the 2 micron minichromosome, and support growth in the absence of Ctf18-RFC. Our data indicate that the eukaryotic replication and transcription machineries use analogous assemblies of multiple chaperones to preserve chromatin integrity. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Histone lysine methylation: critical regulator of memory and behavior.

    Science.gov (United States)

    Jarome, Timothy J; Lubin, Farah D

    2013-01-01

    Histone lysine methylation is a well-established transcriptional mechanism for the regulation of gene expression changes in eukaryotic cells and is now believed to function in neurons of the central nervous system to mediate the process of memory formation and behavior. In mature neurons, methylation of histone proteins can serve to both activate and repress gene transcription. This is in stark contrast to other epigenetic modifications, including histone acetylation and DNA methylation, which have largely been associated with one transcriptional state in the brain. In this review, we discuss the evidence for histone methylation mechanisms in the coordination of complex cognitive processes such as long-term memory formation and storage. In addition, we address the current literature highlighting the role of histone methylation in intellectual disability, addiction, schizophrenia, autism, depression, and neurodegeneration. Further, we discuss histone methylation within the context of other epigenetic modifications and the potential advantages of exploring this newly identified mechanism of cognition, emphasizing the possibility that this molecular process may provide an alternative locus for intervention in long-term psychopathologies that cannot be clearly linked to genes or environment alone.

  3. Physical studies of chromatin. The recombination of histones with DNA.

    Science.gov (United States)

    Boseley, P G; Bradbury, E M; Butler-Browne, G S; Carpenter, B G; Stephens, R M

    1976-02-02

    Experiments have been carried out to define clearly which histone combinations can induce a higher order structure when combined with DNA. The criterion for a higher order structure being the series of low-angle X-ray diffraction maxima nominally at 5.5 nm, 3.7 nm, 2.7 nm and 2.2 nm. Such a pattern, with resolution similar to that of H1-depleted chromatin, is readily attainable by recombining histones H2A + H2B + H3 + H4 with DNA using a salt-gradient dialysis method. However, the use of urea in the recombination procedure is shown to be detrimental to the production of a higher order structure. Low-angle ring patterns are not obtained by recomgining DNA with single pure histones or any combination of histone pairs exept H3 + H4. The diffraction maxima from the latter are, however, weaker than those from chromatin and there are pronounced semi-equatorial arcs. The presence of a third histone, either H2A or H2B in the H3 + H4 recombination mixture tends to distort the recognised low-angle pattern. It is concluded that the histone pair H3 + H4 is essential for the formation of a regular higher order structure in chromatin, although for a complete structural development the presence of H2A + H2B is also required.

  4. Histone deacetylase inhibition abolishes stress-induced spatial memory impairment.

    Science.gov (United States)

    Vargas-López, Viviana; Lamprea, Marisol R; Múnera, Alejandro

    2016-10-01

    Acute stress induced before spatial training impairs memory consolidation. Although non-epigenetic underpinning of such effect has been described, the epigenetic mechanisms involved have not yet been studied. Since spatial training and intense stress have opposite effects on histone acetylation balance, it is conceivable that disruption of such balance may underlie acute stress-induced spatial memory consolidation impairment and that inhibiting histone deacetylases prevents such effect. Trichostatin-A (TSA, a histone deacetylase inhibitor) was used to test its effectiveness in preventing stress' deleterious effect on memory. Male Wistar rats were trained in a spatial task in the Barnes maze; 1-h movement restraint was applied to half of them before training. Immediately after training, stressed and non-stressed animals were randomly assigned to receive either TSA (1mg/kg) or vehicle intraperitoneal injection. Twenty-four hours after training, long-term spatial memory was tested; plasma and brain tissue were collected immediately after the memory test to evaluate corticosterone levels and histone H3 acetylation in several brain areas. Stressed animals receiving vehicle displayed memory impairment, increased plasma corticosterone levels and markedly reduced histone H3 acetylation in prelimbic cortex and hippocampus. Such effects did not occur in stressed animals treated with TSA. The aforementioned results support the hypothesis that acute stress induced-memory impairment is related to histone deacetylation. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Fine mapping of posttranslational modifications of the linker histone H1 from Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Ana Villar-Garea

    Full Text Available The linker histone H1 binds to the DNA in between adjacent nucleosomes and contributes to chromatin organization and transcriptional control. It is known that H1 carries diverse posttranslational modifications (PTMs, including phosphorylation, lysine methylation and ADP-ribosylation. Their biological functions, however, remain largely unclear. This is in part due to the fact that most of the studies have been performed in organisms that have several H1 variants, which complicates the analyses. We have chosen Drosophila melanogaster, a model organism, which has a single H1 variant, to approach the study of the role of H1 PTMs during embryonic development. Mass spectrometry mapping of the entire sequence of the protein showed phosphorylation only in the ten N-terminal amino acids, mostly at S10. For the first time, changes in the PTMs of a linker H1 during the development of a multicellular organism are reported. The abundance of H1 monophosphorylated at S10 decreases as the embryos age, which suggests that this PTM is related to cell cycle progression and/or cell differentiation. Additionally, we have found a polymorphism in the protein sequence that can be mistaken with lysine methylation if the analysis is not rigorous.

  6. PSG gene expression is up-regulated by lysine acetylation involving histone and nonhistone proteins.

    Directory of Open Access Journals (Sweden)

    Soledad A Camolotto

    Full Text Available BACKGROUND: Lysine acetylation is an important post-translational modification that plays a central role in eukaryotic transcriptional activation by modifying chromatin and transcription-related factors. Human pregnancy-specific glycoproteins (PSG are the major secreted placental proteins expressed by the syncytiotrophoblast at the end of pregnancy and represent early markers of cytotrophoblast differentiation. Low PSG levels are associated with complicated pregnancies, thus highlighting the importance of studying the mechanisms that control their expression. Despite several transcription factors having been implicated as key regulators of PSG gene family expression; the role of protein acetylation has not been explored. METHODOLOGY/PRINCIPAL FINDINGS: Here, we explored the role of acetylation on PSG gene expression in the human placental-derived JEG-3 cell line. Pharmacological inhibition of histone deacetylases (HDACs up-regulated PSG protein and mRNA expression levels, and augmented the amount of acetylated histone H3 associated with PSG 5'regulatory regions. Moreover, PSG5 promoter activation mediated by Sp1 and KLF6, via the core promoter element motif (CPE, -147/-140, was markedly enhanced in the presence of the HDAC inhibitor trichostatin A (TSA. This effect correlated with an increase in Sp1 acetylation and KLF6 nuclear localization as revealed by immunoprecipitation and subcellular fractionation assays. The co-activators PCAF, p300, and CBP enhanced Sp1-dependent PSG5 promoter activation through their histone acetylase (HAT function. Instead, p300 and CBP acetyltransferase domain was dispensable for sustaining co-activation of PSG5 promoter by KLF6. CONCLUSIONS/SIGNIFICANCE: Results are consistent with a regulatory role of lysine acetylation on PSG expression through a relaxed chromatin state and an increase in the transcriptional activity of Sp1 and KLF6 following an augmented Sp1 acetylation and KLF6 nuclear localization.

  7. Histone density is maintained during transcription mediated by the chromatin remodeler RSC and histone chaperone NAP1 in vitro.

    Science.gov (United States)

    Kuryan, Benjamin G; Kim, Jessica; Tran, Nancy Nga H; Lombardo, Sarah R; Venkatesh, Swaminathan; Workman, Jerry L; Carey, Michael

    2012-02-01

    ATPases and histone chaperones facilitate RNA polymerase II (pol II) elongation on chromatin. In vivo, the coordinated action of these enzymes is necessary to permit pol II passage through a nucleosome while restoring histone density afterward. We have developed a biochemical system recapitulating this basic process. Transcription through a nucleosome in vitro requires the ATPase remodels structure of chromatin (RSC) and the histone chaperone nucleosome assembly protein 1 (NAP1). In the presence of NAP1, RSC generates a hexasome. Despite the propensity of RSC to evict histones, NAP1 reprograms the reaction such that the hexasome is retained on the template during multiple rounds of transcription. This work has implications toward understanding the mechanism of pol II elongation on chromatin.

  8. Core Java

    CERN Document Server

    Horstmann, Cay S

    2013-01-01

    Fully updated to reflect Java SE 7 language changes, Core Java™, Volume I—Fundamentals, Ninth Edition, is the definitive guide to the Java platform. Designed for serious programmers, this reliable, unbiased, no-nonsense tutorial illuminates key Java language and library features with thoroughly tested code examples. As in previous editions, all code is easy to understand, reflects modern best practices, and is specifically designed to help jumpstart your projects. Volume I quickly brings you up-to-speed on Java SE 7 core language enhancements, including the diamond operator, improved resource handling, and catching of multiple exceptions. All of the code examples have been updated to reflect these enhancements, and complete descriptions of new SE 7 features are integrated with insightful explanations of fundamental Java concepts.

  9. Marked differences in the ability of distinct protamines to disassemble nucleosomal core particles in vitro.

    Science.gov (United States)

    Oliva, R; Mezquita, C

    1986-10-21

    In accordance with the results of classical experiments performed in vitro with calf thymus chromatin and the fish protamine salmine, we have observed that this highly basic, small molecular weight protamine cannot cause major displacement of histones from nucleosomal core particles at concentrations several times higher than physiological (arginine/nucleotide ratios 1-8) and that hyperacetylation of histones facilitates nucleosome disassembly. However, the avian protamine galline, with molecular weight and number of arginine residues almost twice those of common fish protamines, is able to displace the nucleosomal core histones from DNA in vitro at concentrations (arginine/nucleotide ratios 0.6-1.2) within the physiological range (0.8). Our results suggest that the binding of the avian protamine galline to chromatin could be directly involved in the rapid disassembly of nucleosomes that takes place during the nucleohistone nucleoprotamine transition in chicken spermiogenesis.

  10. HDAC8 substrates: Histones and beyond.

    Science.gov (United States)

    Wolfson, Noah A; Pitcairn, Carol Ann; Fierke, Carol A

    2013-02-01

    The lysine deacetylase family of enzymes (HDACs) was first demonstrated to catalyze deacetylation of acetyllysine residues on histones. In subsequent years, HDACs have been shown to recognize a large pool of acetylated nonhistone proteins as substrates. Recently, thousands of acetylated proteins have been discovered, yet in most cases, the HDAC that catalyzes deacetylation in vivo has not been identified. This gap has created the need for better in vivo, in vitro, and in silico approaches for determining HDAC substrates. While HDAC8 is the best kinetically and structurally characterized HDAC, few efficient substrates have yet been substantiated in vivo. In this review, we delineate factors that may be important for determining HDAC8 substrate recognition and catalytic activity, including structure, complex formation, and post-translational modifications. This summary provides insight into the challenges of identifying in vivo substrates for HDAC8, and provides a good vantage point for understanding the variables important for predicting HDAC substrate recognition. Copyright © 2012 Wiley Periodicals, Inc.

  11. Product Variant Master as a Means to Handle Variant Design

    DEFF Research Database (Denmark)

    Hildre, Hans Petter; Mortensen, Niels Henrik; Andreasen, Mogens Myrup

    1996-01-01

    The overall time requiered to design a new product variant relies on two factor: how good the methods to design the new variant are and how good these method are supported by computers.It has been estimated that 80% of all design tasks are variational in that the goal of the design is to adapt an...

  12. Post-Training Intrahippocampal Inhibition of Class I Histone Deacetylases Enhances Long-Term Object-Location Memory

    Science.gov (United States)

    Hawk, Joshua D.; Florian, Cedrick; Abel, Ted

    2011-01-01

    Long-term memory formation involves covalent modification of the histone proteins that package DNA. Reducing histone acetylation by mutating histone acetyltransferases impairs long-term memory, and enhancing histone acetylation by inhibiting histone deacetylases (HDACs) improves long-term memory. Previous studies using HDAC inhibitors to enhance…

  13. Post-Training Intrahippocampal Inhibition of Class I Histone Deacetylases Enhances Long-Term Object-Location Memory

    Science.gov (United States)

    Hawk, Joshua D.; Florian, Cedrick; Abel, Ted

    2011-01-01

    Long-term memory formation involves covalent modification of the histone proteins that package DNA. Reducing histone acetylation by mutating histone acetyltransferases impairs long-term memory, and enhancing histone acetylation by inhibiting histone deacetylases (HDACs) improves long-term memory. Previous studies using HDAC inhibitors to enhance…

  14. Variants of windmill nystagmus.

    Science.gov (United States)

    Choi, Kwang-Dong; Shin, Hae Kyung; Kim, Ji-Soo; Kim, Sung-Hee; Choi, Jae-Hwan; Kim, Hyo-Jung; Zee, David S

    2016-07-01

    Windmill nystagmus is characterized by a clock-like rotation of the beating direction of a jerk nystagmus suggesting separate horizontal and vertical oscillators, usually 90° out of phase. We report oculographic characteristics in three patients with variants of windmill nystagmus in whom the common denominator was profound visual loss due to retinal diseases. Two patients showed a clock-like pattern, while in the third, the nystagmus was largely diagonal (in phase or 180° out of phase) but also periodically changed direction by 180°. We hypothesize that windmill nystagmus is a unique manifestation of "eye movements of the blind." It emerges when the central structures, including the cerebellum, that normally keep eye movements calibrated and gaze steady can no longer perform their task, because they are deprived of the retinal image motion that signals a need for adaptive recalibration.

  15. Structural and Functional Profiling of the Human Histone Methyltransferase SMYD3

    Energy Technology Data Exchange (ETDEWEB)

    Foreman, Kenneth W.; Brown, Mark; Park, Frances; Emtage, Spencer; Harriss, June; Das, Chhaya; Zhu, Li; Crew, Andy; Arnold, Lee; Shaaban, Salam; Tucker, Philip (AltheaDx); (DiscoverEluc.); (Abbott Bioresearch); (OSI Pharm.); (Lilly); (Texas)

    2012-10-23

    The SET and MYND Domain (SMYD) proteins comprise a unique family of multi-domain SET histone methyltransferases that are implicated in human cancer progression. Here we report an analysis of the crystal structure of the full length human SMYD3 in a complex with an analog of the S-adenosyl methionine (SAM) methyl donor cofactor. The structure revealed an overall compact architecture in which the 'split-SET' domain adopts a canonical SET domain fold and closely assembles with a Zn-binding MYND domain and a C-terminal superhelical 9 ?-helical bundle similar to that observed for the mouse SMYD1 structure. Together, these structurally interlocked domains impose a highly confined binding pocket for histone substrates, suggesting a regulated mechanism for its enzymatic activity. Our mutational and biochemical analyses confirm regulatory roles of the unique structural elements both inside and outside the core SET domain and establish a previously undetected preference for trimethylation of H4K20.

  16. Asymmetric breathing motions of nucleosomal DNA and the role of histone tails

    Science.gov (United States)

    Chakraborty, Kaushik; Loverde, Sharon M.

    2017-08-01

    The most important packing unit of DNA in the eukaryotic cell is the nucleosome. It undergoes large-scale structural re-arrangements during different cell cycles. For example, the disassembly of the nucleosome is one of the key steps for DNA replication, whereas reassembly occurs after replication. Thus, conformational dynamics of the nucleosome is crucial for different DNA metabolic processes. We perform three different sets of atomistic molecular dynamics simulations of the nucleosome core particle at varying degrees of salt conditions for a total of 0.7 μs simulation time. We find that the conformational dynamics of the nucleosomal DNA tails are oppositely correlated from each other during the initial breathing motions. Furthermore, the strength of the interaction of the nucleosomal DNA tail with the neighboring H2A histone tail modulates the conformational state of the nucleosomal DNA tail. With increasing salt concentration, the degree of asymmetry in the conformation of the nucleosomal DNA tails decreases as both tails tend to unwrap. This direct correlation between the asymmetric breathing motions of the DNA tails and the H2A histone tails, and its decrease at higher salt concentrations, may play a significant role in the molecular pathway of unwrapping.

  17. Core promoter acetylation is not required for high transcription from the phosphoenolpyruvate carboxylase promoter in maize

    Directory of Open Access Journals (Sweden)

    Horst Ina

    2009-12-01

    Full Text Available Abstract Background Acetylation of promoter nucleosomes is tightly correlated and mechanistically linked to gene activity. However, transcription is not necessary for promoter acetylation. It seems, therefore, that external and endogenous stimuli control histone acetylation and by this contribute to gene regulation. Photosynthetic genes in plants are excellent models with which to study the connection between stimuli and chromatin modifications because these genes are strongly expressed and regulated by multiple stimuli that are easily manipulated. We have previously shown that acetylation of specific histone lysine residues on the photosynthetic phosphoenolpyruvate carboxylase (Pepc promoter in maize is controlled by light and is independent of other stimuli or gene activity. Acetylation of upstream promoter regions responds to a set of other stimuli which include the nutrient availability of the plant. Here, we have extended these studies by analysing histone acetylation during the diurnal and circadian rhythm of the plant. Results We show that histone acetylation of individual lysine residues is removed from the core promoter before the end of the illumination period which is an indication that light is not the only factor influencing core promoter acetylation. Deacetylation is accompanied by a decrease in gene activity. Pharmacological inhibition of histone deacetylation is not sufficient to prevent transcriptional repression, indicating that deacetylation is not controlling diurnal gene regulation. Variation of the Pepc promoter activity during the day is controlled by the circadian oscillator as it is maintained under constant illumination for at least 3 days. During this period, light-induced changes in histone acetylation are completely removed from the core promoter, although the light stimulus is continuously applied. However, acetylation of most sites on upstream promoter elements follows the circadian rhythm. Conclusion Our results

  18. PHF20 Readers Link Methylation of Histone H3K4 and p53 with H4K16 Acetylation

    Directory of Open Access Journals (Sweden)

    Brianna J. Klein

    2016-10-01

    Full Text Available PHF20 is a core component of the lysine acetyltransferase complex MOF (male absent on the first-NSL (non-specific lethal that generates the major epigenetic mark H4K16ac and is necessary for transcriptional regulation and DNA repair. The role of PHF20 in the complex remains elusive. Here, we report on functional coupling between methylation readers in PHF20. We show that the plant homeodomain (PHD finger of PHF20 recognizes dimethylated lysine 4 of histone H3 (H3K4me2 and represents an example of a native reader that selects for this modification. Biochemical and structural analyses help to explain this selectivity and the preference of Tudor2, another reader in PHF20, for dimethylated p53. Binding of the PHD finger to H3K4me2 is required for histone acetylation, accumulation of PHF20 at target genes, and transcriptional activation. Together, our findings establish a unique PHF20-mediated link between MOF histone acetyltransferase (HAT, p53, and H3K4me2, and suggest a model for rapid spreading of H4K16ac-enriched open chromatin.

  19. Regulation of genes related to immune signaling and detoxification in Apis mellifera by an inhibitor of histone deacetylation

    Science.gov (United States)

    Hu, Yee-Tung; Wu, Tsai-Chin; Yang, En-Cheng; Wu, Pei-Chi; Lin, Po-Tse; Wu, Yueh-Lung

    2017-01-01

    The western honeybee (Apis mellifera) is essential for the global economy due to its important role in ecosystems and agriculture as a pollinator of numerous flowering plants and crops. Pesticide abuse has greatly impacted honeybees and caused tremendous loss of honeybee colonies worldwide. The reasons for colony loss remain unclear, but involvement of pesticides and pathogen-pesticide interactions has been hypothesized. Histone deacetylase inhibitors (HDACis) inhibit the activity of histone acetylase, which causes the hyperacetylation of histone cores and influences gene expression. In this study, sodium butyrate, an HDACi, was used as a dietary supplement for honeybees; after treatment, gene expression profiles were analyzed using quantitative PCR. The results showed that sodium butyrate up-regulated genes involved in anti-pathogen and detoxification pathways. The bioassay results showed that honeybees treated with sodium butyrate were more tolerant to imidacloprid. Additionally, sodium butyrate strengthened the immune response of honeybees to invasions of Nosema ceranae and viral infections. We also performed a bioassay in which honeybees were exposed to pesticides and pathogens. Our results provide additional data regarding the mechanism by which honeybees react to stress and the potential application of HDACis in beekeeping. PMID:28112264

  20. The histone demethylases JMJD1A and JMJD2B are transcriptional targets of hypoxia-inducible factor HIF

    DEFF Research Database (Denmark)

    Beyer, Sophie; Kristensen, Malene Maag; Jensen, Kim Steen;

    2008-01-01

    Posttranslational histone modifications serve to store epigenetic information and control both nucleosome assembly and recruitment of non-histone proteins. Histone methylation occurs on arginine and lysine residues and is involved in the regulation of gene transcription. A dynamic control...

  1. Curcumin-induced Histone Acetylation in Malignant Hematologic Cells

    Institute of Scientific and Technical Information of China (English)

    Junbin HU; Yan WANG; Yan CHEN

    2009-01-01

    This study investigated the inhibitory effects of curcumin on proliferation of hemato-logical malignant cells in vitro and the anti-tumor mechanism at histone acetylation/histone deacety-lation levels.The effects of curcumin and histone deacetylase inhibitor trichostatin A (TSA) on the growth of Raji cells were tested by MTT assay.The expression of acetylated histone-3 (H3) in Raji,HL60 and K562 cells,and peripheral blood mononuclear cells (PBMCs) treated with curcumin or TSA was detected by immunohistochemistry and FACS.The results showed curcumin inhibited pro-liferation of Raji cells significantly in a time- and dose-dependent fashion,while exhibited low toxic-ity in PBMCs.Curcumin induced up-regulation of the expression of acetylated H3 dose-dependently in all malignant cell lines tested.In conclusion,curcumin inhibited proliferation of Raji cells selec-tively,enhanced the level of acetylated H3 in Raji,HL60,and K562 cells,which acted as a histone deacetylase inhibitor like TSA.Furthermore,up-regulation of H3 acetylation may play an important role in regulating the proliferation of Raji cells.

  2. The Functional Analysis of Histone Acetyltransferase MOF in Tumorigenesis.

    Science.gov (United States)

    Su, Jiaming; Wang, Fei; Cai, Yong; Jin, Jingji

    2016-01-14

    Changes in chromatin structure and heritably regulating the gene expression by epigenetic mechanisms, such as histone post-translational modification, are involved in most cellular biological processes. Thus, abnormal regulation of epigenetics is implicated in the occurrence of various diseases, including cancer. Human MOF (males absent on the first) is a member of the MYST (Moz-Ybf2/Sas3-Sas2-Tip60) family of histone acetyltransferases (HATs). As a catalytic subunit, MOF can form at least two distinct multiprotein complexes (MSL and NSL) in human cells. Both complexes can acetylate histone H4 at lysine 16 (H4K16); however, the NSL complex possesses broader substrate specificity and can also acetylate histone H4 at lysines 5 and 8 (H4K5 and H4K8), suggesting the complexity of the intracellular functions of MOF. Silencing of MOF in cells leads to genomic instability, inactivation of gene transcription, defective DNA damage repair and early embryonic lethality. Unbalanced MOF expression and its corresponding acetylation of H4K16 have been found in certain primary cancer tissues, including breast cancer, medulloblastoma, ovarian cancer, renal cell carcinoma, colorectal carcinoma, gastric cancer, as well as non-small cell lung cancer. In this review, we provide a brief overview of MOF and its corresponding histone acetylation, introduce recent research findings that link MOF functions to tumorigenesis and speculate on the potential role that may be relevant to tumorigenic pathways.

  3. The Functional Analysis of Histone Acetyltransferase MOF in Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Jiaming Su

    2016-01-01

    Full Text Available Changes in chromatin structure and heritably regulating the gene expression by epigenetic mechanisms, such as histone post-translational modification, are involved in most cellular biological processes. Thus, abnormal regulation of epigenetics is implicated in the occurrence of various diseases, including cancer. Human MOF (males absent on the first is a member of the MYST (Moz-Ybf2/Sas3-Sas2-Tip60 family of histone acetyltransferases (HATs. As a catalytic subunit, MOF can form at least two distinct multiprotein complexes (MSL and NSL in human cells. Both complexes can acetylate histone H4 at lysine 16 (H4K16; however, the NSL complex possesses broader substrate specificity and can also acetylate histone H4 at lysines 5 and 8 (H4K5 and H4K8, suggesting the complexity of the intracellular functions of MOF. Silencing of MOF in cells leads to genomic instability, inactivation of gene transcription, defective DNA damage repair and early embryonic lethality. Unbalanced MOF expression and its corresponding acetylation of H4K16 have been found in certain primary cancer tissues, including breast cancer, medulloblastoma, ovarian cancer, renal cell carcinoma, colorectal carcinoma, gastric cancer, as well as non-small cell lung cancer. In this review, we provide a brief overview of MOF and its corresponding histone acetylation, introduce recent research findings that link MOF functions to tumorigenesis and speculate on the potential role that may be relevant to tumorigenic pathways.

  4. Regulation of Histone Acetylation by Autophagy in Parkinson Disease.

    Science.gov (United States)

    Park, Goonho; Tan, Jieqiong; Garcia, Guillermina; Kang, Yunyi; Salvesen, Guy; Zhang, Zhuohua

    2016-02-12

    Parkinson disease (PD) is the most common age-dependent neurodegenerative movement disorder. Accumulated evidence indicates both environmental and genetic factors play important roles in PD pathogenesis, but the potential interaction between environment and genetics in PD etiology remains largely elusive. Here, we report that PD-related neurotoxins induce both expression and acetylation of multiple sites of histones in cultured human cells and mouse midbrain dopaminergic (DA) neurons. Consistently, levels of histone acetylation are markedly higher in midbrain DA neurons of PD patients compared to those of their matched control individuals. Further analysis reveals that multiple histone deacetylases (HDACs) are concurrently decreased in 1-methyl-4-phenylpyridinium (MPP(+))-treated cells and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mouse brains, as well as midbrain tissues of human PD patients. Finally, inhibition of histone acetyltransferase (HAT) protects, whereas inhibition of HDAC1 and HDAC2 potentiates, MPP(+)-induced cell death. Pharmacological and genetic inhibition of autophagy suppresses MPP(+)-induced HDACs degradation. The study reveals that PD environmental factors induce HDACs degradation and histone acetylation increase in DA neurons via autophagy and identifies an epigenetic mechanism in PD pathogenesis. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Chemical origins of isoform selectivity in histone deacetylase inhibitors.

    Science.gov (United States)

    Butler, Kyle V; Kozikowski, Alan P

    2008-01-01

    Histones undergo extensive posttranslational modifications that affect gene expression. Acetylation is a key histone modification that is primarily regulated by two enzymes, one of which is histone deacetylase (HDAC). The activity of HDAC causes transcriptional silencing of DNA. Eleven distinct zinc-dependent histone deacetylase isoforms have been identified in humans. Each isoform has a unique structure and function, and regulates a unique set of genes. HDAC is responsible for the regulation of many genes involved in cancer cell proliferation, and it has been implicated in the pathogenesis of many neurological conditions. HDAC inhibitors are known to be very effective anti-cancer agents, and research has shown them to be potential treatments for many other conditions. Histone deacetylase inhibitors modify the expression of many genes, and it is possible that inhibition of one isoform could cause epigenetic changes that are beneficial to treatment of a disease, while inhibition of another isoform could cause contradictory changes. Selective HDAC inhibitors will be better able to avoid these types of situations than non-specific inhibitors, and may also be less toxic than pan-HDAC inhibitors. Many potent pan-HDAC inhibitors have already been developed, leaving the development of selective inhibitors at the forefront of HDAC drug development. Certain structural moieties may be added to HDAC inhibitors to give isoform selectivity, and these will be discussed in this review. This review will focus on the applications of selective HDAC inhibitors, inhibitors reported to show selectivity, and the relationship between inhibitor structure and selectivity.

  6. The proteasome and epigenetics: zooming in on histone modifications.

    Science.gov (United States)

    Bach, Svitlana V; Hegde, Ashok N

    2016-08-01

    The proteasome is a structural complex of many proteins that degrades substrates marked by covalent linkage to ubiquitin. Many years of research has shown a role for ubiquitin-proteasome-mediated proteolysis in synaptic plasticity and memory mainly in degrading synaptic, cytoplasmic and nuclear proteins. Recent work indicates that the proteasome has wider proteolytic and non-proteolytic roles in processes such as histone modifications that affect synaptic plasticity and memory. In this review, we assess the evidence gathered from neuronal as well as non-neuronal cell types regarding the function of the proteasome in positive or negative regulation of posttranslational modifications of histones, such as acetylation, methylation and ubiquitination. We discuss the critical roles of the proteasome in clearing excess histone proteins in various cellular contexts and the possible non-proteolytic functions in regulating transcription of target genes. In addition, we summarize the current literature on diverse chromatin-remodeling machineries, such as histone acetyltransferases, deacetylates, methyltransferases and demethylases, as targets for proteasomal degradation across experimental models. Lastly, we provide a perspective on how proteasomal regulation of histone modifications may modulate synaptic plasticity in the nervous system.

  7. Single-nucleosome mapping of histone modifications in S. cerevisiae.

    Directory of Open Access Journals (Sweden)

    Chih Long Liu

    2005-10-01

    Full Text Available Covalent modification of histone proteins plays a role in virtually every process on eukaryotic DNA, from transcription to DNA repair. Many different residues can be covalently modified, and it has been suggested that these modifications occur in a great number of independent, meaningful combinations. Published low-resolution microarray studies on the combinatorial complexity of histone modification patterns suffer from confounding effects caused by the averaging of modification levels over multiple nucleosomes. To overcome this problem, we used a high-resolution tiled microarray with single-nucleosome resolution to investigate the occurrence of combinations of 12 histone modifications on thousands of nucleosomes in actively growing S. cerevisiae. We found that histone modifications do not occur independently; there are roughly two groups of co-occurring modifications. One group of lysine acetylations shows a sharply defined domain of two hypo-acetylated nucleosomes, adjacent to the transcriptional start site, whose occurrence does not correlate with transcription levels. The other group consists of modifications occurring in gradients through the coding regions of genes in a pattern associated with transcription. We found no evidence for a deterministic code of many discrete states, but instead we saw blended, continuous patterns that distinguish nucleosomes at one location (e.g., promoter nucleosomes from those at another location (e.g., over the 3' ends of coding regions. These results are consistent with the idea of a simple, redundant histone code, in which multiple modifications share the same role.

  8. Histone deacetylase inhibitors: pharmacotherapeutic implications as epigenetic modifier

    Directory of Open Access Journals (Sweden)

    Pinki Vishwakarma

    2014-02-01

    Full Text Available Epigenetic modifications such as acetylation and deacetylation of histone proteins play a decisive role in transcriptional alteration and expression of genes. Acetylation is catalysed by the histone acetyl transferases enzymes and activates expression of genes by converting chromatin into a less compact, transcriptionally active state. Histone deacetylases enzymes catalyze deacetylation that condenses chromatin into a closed structure .Consequently transcriptional factors are unable to access DNA and gene expression is suppressed. Balanced activity of HATs and HDACS is essential for normal gene expression. Increased HDAC activity can lead to imbalance in protein acetylation resulting in hypoacetylation, tight chromatin structure and suppression of various genes. This aberrant suppression of genes is the hallmark of several malignant and other diseases including neurodegenerative disorders. Histone Deacetylase Inhibitors (HDACIs have potential to restore the balance of histone acetylation that reverses the silencing of pathological genes. Thus HDACIs modify expression of genes without affecting sequence of DNA and act as epigenetic modifiers. Vorinostat and romidepsin are FDA approved HDACIs. Valproic acid, belinostat and many others are in different phases of clinical trials. This review article explores the target based epigenetic mechanisms as well as existing and potential therapeutic role of HDACIs in various malignant and non-malignant diseases. Data sources were articles published in medical journals and bibliographic database Medline. [Int J Basic Clin Pharmacol 2014; 3(1.000: 27-36

  9. Cellobiohydrolase variants and polynucleotides encoding same

    Energy Technology Data Exchange (ETDEWEB)

    Wogulis, Mark

    2017-04-04

    The present invention relates to variants of a parent cellobiohydrolase II. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

  10. Inter-α inhibitor protein and its associated glycosaminoglycans protect against histone-induced injury

    Science.gov (United States)

    Chaaban, Hala; Keshari, Ravi S.; Silasi-Mansat, Robert; Popescu, Narcis I.; Mehta-D’Souza, Padmaja; Lim, Yow-Pin

    2015-01-01

    Extracellular histones are mediators of tissue injury and organ dysfunction; therefore they constitute potential therapeutic targets in sepsis, inflammation, and thrombosis. Histone cytotoxicity in vitro decreases in the presence of plasma. Here, we demonstrate that plasma inter-α inhibitor protein (IAIP) neutralizes the cytotoxic effects of histones and decreases histone-induced platelet aggregation. These effects are mediated through the negatively charged glycosaminoglycans (GAGs) chondroitin sulfate and high-molecular-weight hyaluronan (HMW-HA) associated with IAIP. Cell surface anionic glycosaminoglycans heparan sulfate and HA protect the cells against histone-mediated damage in vitro. Surface plasmon resonance showed that both IAIP and HMW-HA directly bind to recombinant histone H4. In vivo neutralization of histones with IAIP and HMW-HA prevented histone-induced thrombocytopenia, bleeding, and lung microvascular thrombosis, decreased neutrophil activation, and averted histone-induced production of inflammatory cytokines and chemokines. IAIP and HMW-HA colocalized with histones in necrotic tissues and areas that displayed neutrophil extracellular traps. Increasing amounts of IAIP-histone complexes detected in the plasma of septic baboons correlated with increase in histones and/or nucleosomes and consumption of plasma IAIP. Our data suggest that IAIP, chondroitin sulfate, and HMW-HA are potential therapeutic agents to protect against histone-induced cytotoxicity, coagulopathy, systemic inflammation, and organ damage during inflammatory conditions such as sepsis and trauma. PMID:25631771

  11. Confirmed rare copy number variants implicate novel genes in schizophrenia.

    Science.gov (United States)

    Tam, Gloria W C; van de Lagemaat, Louie N; Redon, Richard; Strathdee, Karen E; Croning, Mike D R; Malloy, Mary P; Muir, Walter J; Pickard, Ben S; Deary, Ian J; Blackwood, Douglas H R; Carter, Nigel P; Grant, Seth G N

    2010-04-01

    Understanding how cognitive processes including learning, memory, decision making and ideation are encoded by the genome is a key question in biology. Identification of sets of genes underlying human mental disorders is a path towards this objective. Schizophrenia is a common disease with cognitive symptoms, high heritability and complex genetics. We have identified genes involved with schizophrenia by measuring differences in DNA copy number across the entire genome in 91 schizophrenia cases and 92 controls in the Scottish population. Our data reproduce rare and common variants observed in public domain data from >3000 schizophrenia cases, confirming known disease loci as well as identifying novel loci. We found copy number variants in PDE10A (phosphodiesterase 10A), CYFIP1 [cytoplasmic FMR1 (Fragile X mental retardation 1)-interacting protein 1], K(+) channel genes KCNE1 and KCNE2, the Down's syndrome critical region 1 gene RCAN1 (regulator of calcineurin 1), cell-recognition protein CHL1 (cell adhesion molecule with homology with L1CAM), the transcription factor SP4 (specificity protein 4) and histone deacetylase HDAC9, among others (see http://www.genes2cognition.org/SCZ-CNV). Integrating the function of these many genes into a coherent model of schizophrenia and cognition is a major unanswered challenge.

  12. Characterization of monoclonal antibodies to histone 2B. Localization of epitopes and analysis of binding to chromatin.

    Science.gov (United States)

    Whitfield, W G; Fellows, G; Turner, B M

    1986-06-16

    Two mouse monoclonal IgM antibodies have been isolated which bind to histone 2B (H2B), as shown by protein blotting and immunostaining and by solid-phase radioimmunoassay (RIA). One of these (HBC-7) was specific for H2B by both techniques whereas the other (2F8) cross-reacted with histone H1 by RIA. Both antibodies failed to recognize H2B limit peptides from trypsin-digested chromatin and did not bind to Drosophila H2B, which differs extensively from vertebrate H2B only in the N-terminal region. These findings indicate that both antibodies recognize epitopes within the trypsin-sensitive, N-terminal region comprising residues 1-20. Binding of antibody HBC-7 was inhibited by in vitro ADP-ribosylation of H2B at glutamic acid residue 2. This strongly suggests that the epitope recognized by HBC-7 is located at the N-terminus of H2B, probably between residues 1 and 8. We have used solid-phase radioimmunoassay to investigate factors which influence the accessibility of this epitope in chromatin. Removal of H1 ('stripping') from high-molecular-mass chromatin had no effect on HBC-7 binding, nor was any difference observed between binding to stripped chromatin and to 146-base-pair (bp) core particles derived from it by nuclease digestion. These results suggest that accessibility of the N-terminal region of H2B is not influenced by H1 itself or by the size or conformation of linker DNA. In contrast, binding of antibody HBC-7 to 146-bp core particles derived from unstripped chromatin was reduced by up to 70%. Binding was restored by exposure of these core particles to the conditions used for stripping. Analysis of the protein content of core particle preparations from stripped and unstripped chromatin suggests that these findings may be attributable to redistribution of non-histone proteins during nuclease digestion. Pre-treatment of high-molecular-mass chromatin or 146-bp core particles with the intercalating dye ethidium bromide resulted in a severalfold increase in binding

  13. The Role of Histone Protein Modifications and Mutations in Histone Modifiers in Pediatric B-Cell Progenitor Acute Lymphoblastic Leukemia

    Science.gov (United States)

    Janczar, Szymon; Janczar, Karolina; Pastorczak, Agata; Harb, Hani; Paige, Adam J. W.; Zalewska-Szewczyk, Beata; Danilewicz, Marian; Mlynarski, Wojciech

    2017-01-01

    While cancer has been long recognized as a disease of the genome, the importance of epigenetic mechanisms in neoplasia was acknowledged more recently. The most active epigenetic marks are DNA methylation and histone protein modifications and they are involved in basic biological phenomena in every cell. Their role in tumorigenesis is stressed by recent unbiased large-scale studies providing evidence that several epigenetic modifiers are recurrently mutated or frequently dysregulated in multiple cancers. The interest in epigenetic marks is especially due to the fact that they are potentially reversible and thus druggable. In B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) there is a relative paucity of reports on the role of histone protein modifications (acetylation, methylation, phosphorylation) as compared to acute myeloid leukemia, T-cell ALL, or other hematologic cancers, and in this setting chromatin modifications are relatively less well studied and reviewed than DNA methylation. In this paper, we discuss the biomarker associations and evidence for a driver role of dysregulated global and loci-specific histone marks, as well as mutations in epigenetic modifiers in BCP-ALL. Examples of chromatin modifiers recurrently mutated/disrupted in BCP-ALL and associated with disease outcomes include MLL1, CREBBP, NSD2, and SETD2. Altered histone marks and histone modifiers and readers may play a particular role in disease chemoresistance and relapse. We also suggest that epigenetic regulation of B-cell differentiation may have parallel roles in leukemogenesis. PMID:28054944

  14. Sliding and peeling of histone during chromatin remodelling

    CERN Document Server

    Garai, Ashok; Chowdhury, Debashish

    2011-01-01

    ATP-dependent chromatin remodeling enzymes (CRE) are bio-molecular motors in eukaryotic cells. These are driven by a chemical fuel, namely, adenosine triphosphate (ATP). CREs actively participate in many cellular processes that require accessibility of specific stretches of DNA which are packaged as chromatin. The basic unit of chromatin is a nucleosome where 146 bp $\\sim$ 50 nm of a double stranded DNA (dsDNA) is wrapped around a spool formed by histone proteins. We investigate the mechanism of peeling of the histone spool, and its complete detachment, from the dsDNA by a CRE. Our two-state model of a CRE captures effectively two distinct chemical (or conformational) states in the mechano-chemical cycle of each ATP-dependent CRE. We calculate the mean times for histone detachment. Our predictions on the ATP-dependence of the measurable quantities can be tested by carrying out {\\it in-vitro} experiments.

  15. Low Proteolytic Clipping of Histone H3 in Cervical Cancer

    Science.gov (United States)

    Sandoval-Basilio, Jorge; Serafín-Higuera, Nicolás; Reyes-Hernandez, Octavio D.; Serafín-Higuera, Idanya; Leija-Montoya, Gabriela; Blanco-Morales, Magali; Sierra-Martínez, Monica; Ramos-Mondragon, Roberto; García, Silvia; López-Hernández, Luz Berenice; Yocupicio-Monroy, Martha; Alcaraz-Estrada, Sofia L.

    2016-01-01

    Chromatin in cervical cancer (CC) undergoes chemical and structural changes that alter the expression pattern of genes. Recently, a potential mechanism, which regulates gene expression at transcriptional levels is the proteolytic clipping of histone H3. However, until now this process in CC has not been reported. Using HeLa cells as a model of CC and human samples from patients with CC, we identify that the H3 cleavage was lower in CC compared with control tissue. Additionally, the histone H3 clipping was performed by serine and aspartyl proteases in HeLa cells. These results suggest that histone H3 clipping operates as part of post-translational modification system in CC. PMID:27698925

  16. Histone modifications associated with cancer cell migration and invasion.

    Science.gov (United States)

    Hieda, Miki; Matsuura, Nariaki; Kimura, Hiroshi

    2015-01-01

    Genome-wide aberrant histone modifications are present in a wide range of cancers, and they are associated with carcinogenesis and cancer progression. Aberrant histone modification patterns affect transcriptional regulation, chromosome stability, chromatin structure, chromatin remodeling, and DNA methylation; furthermore, these patterns can predict clinical outcome in many types of cancer. The main cause of poor clinical outcome is metastasis, which is strongly associated with tissue invasion at the primary tumor site. Invasion of cancer cells into surrounding tissue and the vasculature is an important initial step in tumor metastasis, and cell migration is a critical requirement for metastasis. Here, we describe the advantages of detecting global histone modifications by immunohistochemical analysis and provide a collection of protocols for assaying cell migration, invasion, and cell-extracellular matrix adhesion in vitro.

  17. Histone modification as a drug resistance driver in brain tumors

    Institute of Scientific and Technical Information of China (English)

    Guifa Xi; Barbara Mania-Farnell; Ting Lei; Tadanori Tomita

    2016-01-01

    Patients with brain tumors, specificaly, malignant forms such as glioblastoma, meduloblas-toma and ependymoma, exhibit dismal survival rates despite advances in treatment strategies. Chemotherapeutics, the primary adjuvant treatment for human brain tumors folowing surgery, commonly lack eficacy due to either intrinsic or acquired drug resistance. New treatments tar-geting epigenetic factors are being explored. Post-translational histone modification provides a critical regulatory platform for processes such as chromosome condensation and segregation, apoptosis, gene transcription, and DNA replication and repair. This work reviews how aberrant histone modifications and alterations in histone-modifying enzymes can drive the acquisition of drug resistance in brain tumors. Elucidating these mechanisms should lead to new treatments for overcoming drug resistance.

  18. Involvement of Histone Modifications in Plant Abiotic Stress Responses

    Institute of Scientific and Technical Information of China (English)

    Lianyu Yuan; Xuncheng Liu; Ming Luo; Songguang Yang; Keqiang Wu

    2013-01-01

    As sessile organisms, plants encounter various environmental stimuli including abiotic stresses during their lifecycle. To survive under adverse conditions, plants have evolved intricate mechanisms to perceive external signals and respond accordingly. Responses to various stresses largely depend on the plant capacity to modulate the transcriptome rapidly and specifically. A number of studies have shown that the molecular mechanisms driving the responses of plants to environmental stresses often depend on nucleosome histone post-translational modifications including histone acetylation, methylation, ubiquitination, and phosphorylation. The combined effects of these modifications play an essential role in the regulation of stress responsive gene expression. In this review, we highlight our current understanding of the epigenetic mechanisms of histone modifications and their roles in plant abiotic stress response.

  19. Certain variants of multipermutohedron ideals

    Indian Academy of Sciences (India)

    AJAY KUMAR; CHANCHAL KUMAR

    2016-10-01

    Multipermutohedron ideals have rich combinatorial properties. An explicit combinatorial formula for the multigraded Betti numbers of a multipermutohedron ideal and their Alexander duals are known. Also, the dimension of the Artinian quotient of an Alexander dual of a multipermutohedron ideal is the number of generalized parking functions. In this paper, monomial ideals which are certain variants of multipermutohedron ideals are studied. Multigraded Betti numbers of these variant monomial ideals and their Alexander duals are obtained. Further, many interesting combinatorial properties of multipermutohedron ideals are extended to these variant monomial ideals.

  20. Schizophrenia copy number variants and associative learning

    Science.gov (United States)

    Clifton, N E; Pocklington, A J; Scholz, B; Rees, E; Walters, J T R; Kirov, G; O'Donovan, M C; Owen, M J; Wilkinson, L S; Thomas, K L; Hall, J

    2017-01-01

    Large-scale genomic studies have made major progress in identifying genetic risk variants for schizophrenia. A key finding from these studies is that there is an increased burden of genomic copy number variants (CNVs) in schizophrenia cases compared with controls. The mechanism through which these CNVs confer risk for the symptoms of schizophrenia, however, remains unclear. One possibility is that schizophrenia risk CNVs impact basic associative learning processes, abnormalities of which have long been associated with the disorder. To investigate whether genes in schizophrenia CNVs impact on specific phases of associative learning we combined human genetics with experimental gene expression studies in animals. In a sample of 11 917 schizophrenia cases and 16 416 controls, we investigated whether CNVs from patients with schizophrenia are enriched for genes expressed during the consolidation, retrieval or extinction of associative memories. We show that CNVs from cases are enriched for genes expressed during fear extinction in the hippocampus, but not genes expressed following consolidation or retrieval. These results suggest that CNVs act to impair inhibitory learning in schizophrenia, potentially contributing to the development of core symptoms of the disorder. PMID:27956746

  1. Effects of orally applied butyrate bolus on histone acetylation and cytochrome P450 enzyme activity in the liver of chicken – a randomized controlled trial

    Directory of Open Access Journals (Sweden)

    Mátis Gábor

    2013-01-01

    Full Text Available Abstract Background Butyrate is known as histone deacetylase inhibitor, inducing histone hyperacetylation in vitro and playing a predominant role in the epigenetic regulation of gene expression and cell function. We hypothesized that butyrate, endogenously produced by intestinal microbial fermentation or applied as a nutritional supplement, might cause similar in vivo modifications in the chromatin structure of the hepatocytes, influencing the expression of certain genes and therefore modifying the activity of hepatic microsomal drug-metabolizing cytochrome P450 (CYP enzymes. Methods An animal study was carried out in chicken as a model to investigate the molecular mechanisms of butyrate’s epigenetic actions in the liver. Broiler chicks in the early post-hatch period were treated once daily with orally administered bolus of butyrate following overnight starvation with two different doses (0.25 or 1.25 g/kg body weight per day for five days. After slaughtering, cell nucleus and microsomal fractions were separated by differential centrifugation from the livers. Histones were isolated from cell nuclei and acetylation of hepatic core histones was screened by western blotting. The activity of CYP2H and CYP3A37, enzymes involved in biotransformation in chicken, was detected by aminopyrine N-demethylation and aniline-hydroxylation assays from the microsomal suspensions. Results Orally added butyrate, applied in bolus, had a remarkable impact on nucleosome structure of hepatocytes: independently of the dose, butyrate caused hyperacetylation of histone H2A, but no changes were monitored in the acetylation state of H2B. Intensive hyperacetylation of H3 was induced by the higher administered dose, while the lower dose tended to increase acetylation ratio of H4. In spite of the observed modification in histone acetylation, no significant changes were observed in the hepatic microsomal CYP2H and CYP3A37 activity. Conclusion Orally added butyrate in bolus

  2. Structural insight into histone recognition by the ING PHD fingers.

    Science.gov (United States)

    Champagne, Karen S; Kutateladze, Tatiana G

    2009-05-01

    The Inhibitor of Growth (ING) tumor suppressors are implicated in oncogenesis, control of DNA damage repair, cellular senescence and apoptosis. All members of the ING family contain unique amino-terminal regions and a carboxy-terminal plant homeodomain (PHD) finger. While the amino-terminal domains associate with a number of protein effectors including distinct components of histone deacetylase (HDAC) and histone acetyltransferase (HAT) complexes, the PHD finger binds strongly and specifically to histone H3 trimethylated at lysine 4 (H3K4me3). In this review we describe the molecular mechanism of H3K4me3 recognition by the ING1-5 PHD fingers, analyze the determinants of the histone specificity and compare the biological activities and structures within subsets of PHD fingers. The atomic-resolution structures of the ING PHD fingers in complex with a H3K4me3 peptide reveal that the histone tail is bound in a large and deep binding site encompassing nearly one-third of the protein surface. An extensive network of intermolecular hydrogen bonds, hydrophobic and cation-pi contacts, and complementary surface interactions coordinate the first six residues of the H3K4me3 peptide. The trimethylated Lys4 occupies an elongated groove, formed by the highly conserved aromatic and hydrophobic residues of the PHD finger, whereas the adjacent groove accommodates Arg2. The two grooves are connected by a narrow channel, the small size of which defines the PHD finger's specificity, excluding interactions with other modified histone peptides. Binding of the ING PHD fingers to H3K4me3 plays a critical role in regulating chromatin acetylation. The ING proteins function as tethering molecules that physically link the HDAC and HAT enzymatic complexes to chromatin. In this review we also highlight progress recently made in understanding the molecular basis underlying biological and tumorigenic activities of the ING tumor suppressors.

  3. Substrate Specificity Profiling of Histone-Modifying Enzymes by Peptide Microarray

    Science.gov (United States)

    Cornett, E.M.; Dickson, B.M.; Vaughan, R.M.; Krishnan, S.; Trievel, R.C.; Strahl, B.D.; Rothbart, S.B.

    2017-01-01

    The dynamic addition and removal of covalent posttranslational modifications (PTMs) on histone proteins serves as a major mechanism regulating chromatin-templated biological processes in eukaryotic genomes. Histone PTMs and their combinations function by directly altering the physical structure of chromatin and as rheostats for effector protein interactions. In this chapter, we detail microarray-based methods for analyzing the substrate specificity of lysine methyltransferase and demethylase enzymes on immobilized synthetic histone peptides. Consistent with the “histone code” hypothesis, we reveal a strong influenceof adjacent and,surprisingly,distant histonePTMs onthe ability of histone-modifying enzymes to methylate or demethylate their substrates. This platform will greatly facilitate future investigations into histone substrate specificity and mechanisms of PTM signaling that regulate the catalytic properties of histone-modifying enzymes. PMID:27423856

  4. Histone acetylation in astrocytes suppresses GFAP and stimulates a reorganization of the intermediate filament network

    NARCIS (Netherlands)

    Kanski, Regina; Sneeboer, Marjolein A M; van Bodegraven, Emma J; Sluijs, Jacqueline A; Kropff, Wietske; Vermunt, Marit W.; Creyghton, Menno P; De Filippis, Lidia; Vescovi, Angelo; Aronica, Eleonora; van Tijn, P.; van Strien, Miriam E; Hol, Elly M

    2014-01-01

    Glial fibrillary acidic protein (GFAP) is the main intermediate filament in astrocytes and is regulated by epigenetic mechanisms during development. We demonstrate that histone acetylation also controls GFAP expression in mature astrocytes. Inhibition of histone deacetylases (HDACs) with trichostati

  5. Mechanisms of transcriptional repression by histone lysine methylation

    DEFF Research Database (Denmark)

    Hublitz, Philip; Albert, Mareike; Peters, Antoine H F M

    2009-01-01

    . In this report, we review the recent literature to deduce mechanisms underlying Polycomb and H3K9 methylation mediated repression, and describe the functional interplay with activating H3K4 methylation. We summarize recent data that indicate a close relationship between GC density of promoter sequences......, transcription factor binding and the antagonizing activities of distinct epigenetic regulators such as histone methyltransferases (HMTs) and histone demethylases (HDMs). Subsequently, we compare chromatin signatures associated with different types of transcriptional outcomes from stable repression to highly...... dynamic regulated genes, strongly suggesting that the interplay of different epigenetic pathways is essential in defining specific types of heritable chromatin and associated transcriptional states....

  6. Characterization of human papillomavirus type 16 pseudovirus containing histones.

    Science.gov (United States)

    Kim, Hyoung Jin; Kwag, Hye-Lim; Kim, Hong-Jin

    2016-08-27

    Pseudoviruses (PsVs) that encapsidate a reporter plasmid DNA have been used as surrogates for native human papillomavirus (HPV), whose continuous production is technically difficult. HPV PsVs have been designed to form capsids made up of the major capsid protein L1 and the minor capsid proteins L2. HPV PsVs have been produced in 293TT cells transfected with plasmid expressing L1 and L2 protein and plasmid containing the reporter gene. Several studies have suggested that naturally occurring HPV virions contain cellular histones, and histones have also been identified in mature HPV PsVs. However, the effect of the histones on the properties of the PsVs has not been investigated. Using heparin chromatography, we separated mature HPV type 16 PsVs into three fractions (I, II, and III) according to their heparin-binding affinities. The amounts of cellular histone and cellular nucleotides per PsV were found to increase in the order fraction I, II and III. It appeared that PsVs in fraction I contains just small amount of cellular histone in Western blot analysis. The proportions of the three fractions in PsV preparations were 83.4, 7.5, and 9.1 % for fraction I, II, and III PsVs, respectively. In the electron microscope PsVs in fraction I appeared to have a more condensed structure than those in fractions II and III. Under the electron microscope fraction II and III PsVs appeared to be covered by substantial amounts of cellular histone while there was no visible histone covering PsVs of fraction I. Also the levels of reporter gene expression in infections of fraction II and III PsVs to 293TT cells were significantly lower than those in infections of fraction I PsV, and fraction II and III particles had significantly reduced immunogenicity. Our findings suggest that the involvement of large amounts of cellular histones during PsV formation interferes with the structural integrity of the PsVs and affects their immunogenicity. The fraction I particle therefore has the most

  7. Histone Deacetylase Inhibitors: Synthesis of Tetrapeptide Analogue SAHA/TPX

    Directory of Open Access Journals (Sweden)

    Lynda Ekou

    2011-01-01

    Full Text Available The inhibition of HDAC (histone deacetylase activity by specific inhibitors induces growth arrest, differentiation and apoptosis of transformed or several cancer cells. Some of these inhibitors are in clinical trial at phase I or phase II. The discovery and development of specific HDAC inhibitors are helpful for cancer therapy. In this paper we describe the synthesis of simple inhibitor B hybrid analogue suberoylanilide hydroxamic acid (SAHA, trapoxin B (TPX B in as little as five steps. This compound is interesting lead for the design of potent inhibitors of histone deacetylase.

  8. Low Proteolytic Clipping of Histone H3 in Cervical Cancer

    OpenAIRE

    Sandoval-Basilio, Jorge; Serafín-Higuera, Nicolás; Reyes-Hernandez, Octavio D.; Serafín-Higuera, Idanya; Leija-Montoya, Gabriela; Blanco-Morales, Magali; Sierra-Martínez, Monica; Ramos-Mondragon, Roberto; García, Silvia; López-Hernández, Luz Berenice; Yocupicio-Monroy, Martha; Alcaraz-Estrada, Sofia L.

    2016-01-01

    Chromatin in cervical cancer (CC) undergoes chemical and structural changes that alter the expression pattern of genes. Recently, a potential mechanism, which regulates gene expression at transcriptional levels is the proteolytic clipping of histone H3. However, until now this process in CC has not been reported. Using HeLa cells as a model of CC and human samples from patients with CC, we identify that the H3 cleavage was lower in CC compared with control tissue. Additionally, the histone H3...

  9. Physarum polycephalum for Studying the Function of Histone Modifications In Vivo.

    Science.gov (United States)

    Menil-Philippot, Vanessa; Thiriet, Christophe

    2017-01-01

    Histone modifications have been widely correlated with genetic activities. However, how these posttranslational modifications affect the dynamics and the structure of chromatin is poorly understood. Here, we describe the incorporation of the exogenous histone proteins into the slime mold Physarum polycephalum, which has been revealed to be a valuable tool for examining different facets of the function histones in chromatin dynamics like replication-coupled chromatin assembly, histone exchange, and nucleosome turnover.

  10. Gene Variants Reduce Opioid Risks

    Science.gov (United States)

    ... Opioids Prescription Drugs & Cold Medicines Steroids (Anabolic) Synthetic Cannabinoids (K2/Spice) Synthetic Cathinones (Bath Salts) Tobacco/Nicotine ... variant of the gene for the μ-opioid receptor (OPRM1) with a decreased risk for addiction to ...

  11. Histone deacetylase 1, 2, 6 and acetylated histone H4 in B- and T-cell lymphomas

    DEFF Research Database (Denmark)

    Marquard, L.; Poulsen, C.B.; Gjerdrum, L.M.;

    2009-01-01

    AIMS: Histone deacetylase (HDAC) inhibitors are novel therapeutics in the treatment of peripheral T-cell lymphoma, unspecified (PTCL) and diffuse large B-cell lymphoma (DLBCL), where, for unknown reasons, T-cell malignancies appear to be more sensitive than B-cell malignancies. The aim was to det......AIMS: Histone deacetylase (HDAC) inhibitors are novel therapeutics in the treatment of peripheral T-cell lymphoma, unspecified (PTCL) and diffuse large B-cell lymphoma (DLBCL), where, for unknown reasons, T-cell malignancies appear to be more sensitive than B-cell malignancies. The aim...... was to determine HDAC expression in DLBCL and PTCL which has not previously been investigated. METHODS AND RESULTS: The expression of HDAC1, HDAC2, HDAC6 and acetylated histone H4 was examined immunohistochemically in 31 DLBCL and 45 PTCL. All four markers showed high expression in both DLBCL and PTCL compared...

  12. Quantitative analysis of histone modifications: formaldehyde is a source of pathological n(6)-formyllysine that is refractory to histone deacetylases.

    Science.gov (United States)

    Edrissi, Bahar; Taghizadeh, Koli; Dedon, Peter C

    2013-01-01

    Aberrant protein modifications play an important role in the pathophysiology of many human diseases, in terms of both dysfunction of physiological modifications and the formation of pathological modifications by reaction of proteins with endogenous electrophiles. Recent studies have identified a chemical homolog of lysine acetylation, N(6)-formyllysine, as an abundant modification of histone and chromatin proteins, one possible source of which is the reaction of lysine with 3'-formylphosphate residues from DNA oxidation. Using a new liquid chromatography-coupled to tandem mass spectrometry method to quantify all N(6)-methyl-, -acetyl- and -formyl-lysine modifications, we now report that endogenous formaldehyde is a major source of N(6)-formyllysine and that this adduct is widespread among cellular proteins in all compartments. N(6)-formyllysine was evenly distributed among different classes of histone proteins from human TK6 cells at 1-4 modifications per 10(4) lysines, which contrasted strongly with lysine acetylation and mono-, di-, and tri-methylation levels of 1.5-380, 5-870, 0-1400, and 0-390 per 10(4) lysines, respectively. While isotope labeling studies revealed that lysine demethylation is not a source of N(6)-formyllysine in histones, formaldehyde exposure was observed to cause a dose-dependent increase in N(6)-formyllysine, with use of [(13)C,(2)H2]-formaldehyde revealing unchanged levels of adducts derived from endogenous sources. Inhibitors of class I and class II histone deacetylases did not affect the levels of N(6)-formyllysine in TK6 cells, and the class III histone deacetylase, SIRT1, had minimal activity (removal by histone deacetylases, which supports the idea that this abundant protein modification could interfere with normal regulation of gene expression if it arises at conserved sites of physiological protein secondary modification.

  13. Quantitative analysis of histone modifications: formaldehyde is a source of pathological n(6-formyllysine that is refractory to histone deacetylases.

    Directory of Open Access Journals (Sweden)

    Bahar Edrissi

    Full Text Available Aberrant protein modifications play an important role in the pathophysiology of many human diseases, in terms of both dysfunction of physiological modifications and the formation of pathological modifications by reaction of proteins with endogenous electrophiles. Recent studies have identified a chemical homolog of lysine acetylation, N(6-formyllysine, as an abundant modification of histone and chromatin proteins, one possible source of which is the reaction of lysine with 3'-formylphosphate residues from DNA oxidation. Using a new liquid chromatography-coupled to tandem mass spectrometry method to quantify all N(6-methyl-, -acetyl- and -formyl-lysine modifications, we now report that endogenous formaldehyde is a major source of N(6-formyllysine and that this adduct is widespread among cellular proteins in all compartments. N(6-formyllysine was evenly distributed among different classes of histone proteins from human TK6 cells at 1-4 modifications per 10(4 lysines, which contrasted strongly with lysine acetylation and mono-, di-, and tri-methylation levels of 1.5-380, 5-870, 0-1400, and 0-390 per 10(4 lysines, respectively. While isotope labeling studies revealed that lysine demethylation is not a source of N(6-formyllysine in histones, formaldehyde exposure was observed to cause a dose-dependent increase in N(6-formyllysine, with use of [(13C,(2H2]-formaldehyde revealing unchanged levels of adducts derived from endogenous sources. Inhibitors of class I and class II histone deacetylases did not affect the levels of N(6-formyllysine in TK6 cells, and the class III histone deacetylase, SIRT1, had minimal activity (<10% with a peptide substrate containing the formyl adduct. These data suggest that N(6-formyllysine is refractory to removal by histone deacetylases, which supports the idea that this abundant protein modification could interfere with normal regulation of gene expression if it arises at conserved sites of physiological protein secondary

  14. Holocarboxylase synthetase interacts physically with euchromatic histone-lysine N-methyltransferase, linking histone biotinylation with methylation events.

    Science.gov (United States)

    Li, Yong; Hassan, Yousef I; Moriyama, Hideaki; Zempleni, Janos

    2013-08-01

    Holocarboxylase synthetase (HCS) catalyzes the binding of the vitamin biotin to histones H3 and H4, thereby creating rare histone biotinylation marks in the epigenome. These marks co-localize with K9-methylated histone H3 (H3K9me), an abundant gene repression mark. The abundance of H3K9me marks in transcriptionally competent loci decreases when HCS is knocked down and when cells are depleted of biotin. Here we tested the hypothesis that the creation of H3K9me marks is at least partially explained by physical interactions between HCS and histone-lysine N-methyltransferases. Using a novel in silico protocol, we predicted that HCS-interacting proteins contain a GGGG(K/R)G(I/M)R motif. This motif, with minor variations, is present in the histone-lysine N-methyltransferase EHMT1. Physical interactions between HCS and the N-terminal, ankyrin and SET domains in EHMT1 were confirmed using yeast-two-hybrid assays, limited proteolysis assays and co-immunoprecipitation. The interactions were stronger between HCS and the N-terminus in EHMT1 compared with the ankyrin and SET domains, consistent with the localization of the HCS-binding motif in the EHMT1 N-terminus. HCS has the catalytic activity to biotinylate K161 within the binding motif in EHMT1. Mutation of K161 weakened the physical interaction between EHMT1 and HCS, but it is unknown whether this effect was caused by loss of biotinylation or loss of the motif. Importantly, HCS knockdown decreased the abundance of H3K9me marks in repeats, suggesting that HCS plays a role in creating histone methylation marks in these loci. We conclude that physical interactions between HCS and EHMT1 mediate epigenomic synergies between biotinylation and methylation events. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Autoantibodies against citrullinated histone H3 in rheumatoid arthritis and periodontitis patients

    NARCIS (Netherlands)

    Janssen, Koen M. J.; de Smit, Menke J.; Withaar, Coenraad; Brouwer, Elisabeth; van Winkelhoff, Arie J.; Vissink, Arjan; Westra, Johanna

    Aim: To determine the presence of citrullinated histones in inflamed periodontal tissue and to determine the presence of anti-citrullinated histone autoantibodies in sera from patients with rheumatoid arthritis (RA) and periodontitis (PD) patients. Methods: The presence of citrullinated histone H3,

  16. Histone H3 Lysine Methylation in Cognition and Intellectual Disability Disorders

    Science.gov (United States)

    Parkel, Sven; Lopez-Atalaya, Jose P.; Barco, Angel

    2013-01-01

    Recent research indicates that epigenetic mechanisms and, in particular, the post-translational modification (PTM) of histones may contribute to memory encoding and storage. Among the dozens of possible histone PTMs, the methylation/demethylation of lysines in the N-terminal tail of histone H3 exhibits particularly strong links with cognitive…

  17. In vitro interactions of extracellular histones with LDL suggest a potential pro-atherogenic role.

    Directory of Open Access Journals (Sweden)

    Alan D Pemberton

    Full Text Available BACKGROUND: Nuclear histones have previously been shown to aggregate LDL in vitro, suggestive of a possible pro-atherogenic role. Recent studies indicate that histones are released during acute inflammation, and therefore might interact with circulating lipoproteins in vivo. In view of the associative link between inflammation and cardiovascular disease, the behaviour of histones was investigated using in vitro models of LDL retention and foam cell formation. METHODOLOGY/PRINCIPAL FINDINGS: Heparin agarose beads were used as a model of a matrix rich in sulphated glycosaminoglycans, to which histones bind strongly. Histone-modified beads were observed to pull down more LDL from solution than untreated beads, indicating that histones can function as bridging molecules, enhancing LDL retention. Furthermore, addition of heparin inhibited histone-induced aggregation of LDL. To model foam cell formation, murine RAW 264.7 macrophages were incubated for 24 h in the presence of LDL, histones, LDL plus histones or vehicle control. Cells incubated with LDL in the presence of histones accumulated significantly more intracellular lipid than with LDL or histone alone. CONCLUSIONS/SIGNIFICANCE: These results are consistent with a potential pro-atherogenic role for extracellular histones, which should be investigated further.

  18. Histone deacetylase inhibitors induced differentiation and accelerated mineralization of pulp-derived cells.

    LENUS (Irish Health Repository)

    Duncan, Henry F

    2012-03-01

    Histone deacetylase inhibitors (HDACis) alter the homeostatic balance between 2 groups of cellular enzymes, histone deacetylases (HDACs) and histone acetyltransferases (HATs), increasing transcription and influencing cell behavior. This study investigated the potential of 2 HDACis, valproic acid (VPA) and trichostatin A (TSA), to promote reparative processes in pulp cells as assayed by viability, cell cycle, and mineralization analyses.

  19. Autoantibodies against citrullinated histone H3 in rheumatoid arthritis and periodontitis patients

    NARCIS (Netherlands)

    Janssen, Koen M J; de Smit, Menke J; Withaar, Coenraad; Brouwer, Elisabeth; van Winkelhoff, Arie Jan; Vissink, Arjan; Westra, Johanna

    2017-01-01

    AIM: To determine presence of citrullinated histones in inflamed periodontal tissue, and to determine presence of anti-citrullinated histone autoantibodies in sera from rheumatoid arthritis (RA) and periodontitis (PD)-patients . METHODS: Presence of citrullinated histone H3, PAD4 and CD68 was determ

  20. Histones link inflammation and thrombosis through the induction of Weibel-Palade body exocytosis.

    Science.gov (United States)

    Michels, A; Albánez, S; Mewburn, J; Nesbitt, K; Gould, T J; Liaw, P C; James, P D; Swystun, L L; Lillicrap, D

    2016-11-01

    Essentials Dysregulated DNA and histone release can promote pathological immunothrombosis. Weibel-Palade bodies (WPBs) are sentinel-like organelles that respond to proinflammatory stimuli. Histones induce WPB exocytosis in a caspase, calcium and charge-dependent mechanism. A targetable axis may exist between DNA/histones and WPBs in inflammation and immunothrombosis.

  1. Novel variants in GNAI3 associated with auriculocondylar syndrome strengthen a common dominant negative effect

    OpenAIRE

    2014-01-01

    Auriculocondylar syndrome is a rare craniofacial disorder comprising core features of micrognathia, condyle dysplasia and question mark ear. Causative variants have been identified in PLCB4, GNAI3 and EDN1, which are predicted to function within the EDN1–EDNRA pathway during early pharyngeal arch patterning. To date, two GNAI3 variants in three families have been reported. Here we report three novel GNAI3 variants, one segregating with affected members in a family previously linked to 1p21.1-...

  2. Histone acetylation regulates osteodifferentiation of hDPSCs via DSPP.

    Science.gov (United States)

    Gu, Shensheng; Liang, Jingping; Wang, Jia; Liu, Bin

    2013-06-01

    Dental pulp stem cells (DPSCs) are a unique population of precursor cells isolated from postnatal human dental pulp, with the ability to regenerate a reparative dentin-like complex. We examined the regulation of odontoblast-like differentiation of DPSCs by histone acetylation. Western blot analysis showed that histone H3 acetylation was strongly induced in osteodifferentiation medium. Inhibition of histone acetyltransferase by garcinol reversed osteodifferentiation and mineral formation. Real-time polymerase chain reaction assay indicated that the dentin sialophosphoprotein (DSPP) gene, which is mainly expressed in odontoblasts and preameloblasts in teeth and plays an important role in tooth function, was also down-regulated in garcinol-treated cells. Moreover, lentivirus-mediated knockdown of DSPP in human DPSCs was associated with significant inhibition of mineral formation, but not osteoblast differentiation. In conclusion, the results of this study suggest that DSPP positively affects mineral formation, and that odontoblast-like differentiation and maturation of DPSCs can be regulated by histone acetylation of the DSPP gene.

  3. Small molecule modulators of histone acetyltransferase p300.

    Science.gov (United States)

    Balasubramanyam, Karanam; Swaminathan, V; Ranganathan, Anupama; Kundu, Tapas K

    2003-05-23

    Histone acetyltransferases (HATs) are a group of enzymes that play a significant role in the regulation of gene expression. These enzymes covalently modify the N-terminal lysine residues of histones by the addition of acetyl groups from acetyl-CoA. Dysfunction of these enzymes is often associated with the manifestation of several diseases, predominantly cancer. Here we report that anacardic acid from cashew nut shell liquid is a potent inhibitor of p300 and p300/CBP-associated factor histone acetyltranferase activities. Although it does not affect DNA transcription, HAT-dependent transcription from a chromatin template was strongly inhibited by anacardic acid. Furthermore, we describe the design and synthesis of an amide derivative N-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-6-pentadecyl-benzamide (CTPB) using anacardic acid as a synthon, which remarkably activates p300 HAT activity but not that of p300/CBP-associated factor. Although CTPB does not affect DNA transcription, it enhances the p300 HAT-dependent transcriptional activation from in vitro assembled chromatin template. However, it has no effect on histone deacetylase activity. These compounds would be useful as biological switching molecules for probing into the role of p300 in transcriptional studies and may also be useful as new chemical entities for the development of anticancer drugs.

  4. Mechanistic stochastic model of histone modification pattern formation

    NARCIS (Netherlands)

    L.C.M. Anink-Groenen; T.R. Maarleveld; P.J. Verschure; F.J. Bruggeman

    2014-01-01

    BACKGROUND: The activity of a single gene is influenced by the composition of the chromatin in which it is embedded. Nucleosome turnover, conformational dynamics, and covalent histone modifications each induce changes in the structure of chromatin and its affinity for regulatory proteins. The dynami

  5. Innovative Strategies for Selective Inhibition of Histone Deacetylases

    DEFF Research Database (Denmark)

    Maolanon, Alex Ramalak; Madsen, Andreas Stahl; Olsen, Christian Adam

    2016-01-01

    Histone deacetylases (HDAC) are a family of closely related enzymes involved in epigenetic and posttranscriptional regulation of numerous genes and proteins. Their deregulation is associated with a number of diseases, and a handful of HDAC inhibitors have been approved for cancer treatment. None...

  6. Cell shape regulates global histone acetylation in human mammaryepithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Le Beyec, Johanne; Xu, Ren; Lee, Sun-Young; Nelson, Celeste M.; Rizki, Aylin; Alcaraz, Jordi; Bissell, Mina J.

    2007-02-28

    Extracellular matrix (ECM) regulates cell morphology and gene expression in vivo; these relationships are maintained in three-dimensional (3D) cultures of mammary epithelial cells. In the presence of laminin-rich ECM (lrECM), mammary epithelial cells round up and undergo global histone deacetylation, a process critical for their functional differentiation. However, it remains unclear whether lrECM-dependent cell rounding and global histone deacetylation are indeed part of a common physical-biochemical pathway. Using 3D cultures as well as nonadhesive and micropatterned substrata, here we showed that the cell 'rounding' caused by lrECM was sufficient to induce deacetylation of histones H3 and H4 in the absence of biochemical cues. Microarray and confocal analysis demonstrated that this deacetylation in 3D culture is associated with a global increase in chromatin condensation and a reduction in gene expression. Whereas cells cultured on plastic substrata formed prominent stress fibers, cells grown in 3D lrECM or on micropatterns lacked these structures. Disruption of the actin cytoskeleton with cytochalasin D phenocopied the lrECM-induced cell rounding and histone deacetylation. These results reveal a novel link between ECM-controlled cell shape and chromatin structure, and suggest that this link is mediated by changes in the actin cytoskeleton.

  7. Histone 3 s10 phosphorylation: "caught in the R loop!".

    Science.gov (United States)

    Skourti-Stathaki, Konstantina; Proudfoot, Nicholas J

    2013-11-21

    In this issue of Molecular Cell, Castellano-Pozo et al. (2013) describe a connection between R loop structures and histone 3 S10 phosphorylation (H3S10P), a mark of chromatin compaction. Their results constitute a significant advance in our understanding of the role of R loops in genomic instability.

  8. Functional characterization of Candida albicans Hos2 histone deacetylase.

    Science.gov (United States)

    Karthikeyan, G; Paul-Satyaseela, Maneesh; Dhatchana Moorthy, Nachiappan; Gopalaswamy, Radha; Narayanan, Shridhar

    2013-01-01

    Candida albicans is a mucosal commensal organism capable of causing superficial (oral and vaginal thrush) infections in immune normal hosts, but is a major pathogen causing systemic and mucosal infections in immunocompromised individuals. Azoles have been very effective anti-fungal agents and the mainstay in treating opportunistic mold and yeast infections. Azole resistant strains have emerged compromising the utility of this class of drugs. It has been shown that azole resistance can be reversed by the co-administration of a histone deacetylase (HDAC) inhibitor, suggesting that resistance is mediated by epigenetic mechanisms possibly involving Hos2, a fungal deacetylase. We report here the cloning and functional characterization of  HOS2 (High Osmolarity  Sensitive) , a gene coding for fungal histone deacetylase from  C. albicans. Inhibition studies showed that Hos2 is susceptible to pan inhibitors such as trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA), but is not inhibited by class I inhibitors such as MS-275. This  in  vitro enzymatic assay, which is amenable to high throughput could be used for screening potent fungal Hos2 inhibitors that could be a potential anti-fungal adjuvant. Purified Hos2 protein consistently deacetylated tubulins, rather than histones from TSA-treated cells. Hos2 has been reported to be a putative NAD+ dependent histone deacetylase, a feature of sirtuins. We assayed for sirtuin activation with resveratrol and purified Hos2 protein and did not find any sirtuin activity.

  9. Sepsis and ARDS: The Dark Side of Histones

    Directory of Open Access Journals (Sweden)

    Zhiheng Xu

    2015-01-01

    Full Text Available Despite advances in management over the last several decades, sepsis and acute respiratory distress syndrome (ARDS still remain major clinical challenges and the leading causes of death for patients in intensive care units (ICUs due to insufficient understanding of the pathophysiological mechanisms of these diseases. However, recent studies have shown that histones, also known as chromatin-basic structure proteins, could be released into the extracellular space during severe stress and physical challenges to the body (e.g., sepsis and ARDS. Due to their cytotoxic and proinflammatory effects, extracellular histones can lead to excessive and overwhelming cell damage and death, thus contributing to the pathogenesis of both sepsis and ARDS. In addition, antihistone-based treatments (e.g., neutralizing antibodies, activated protein C, and heparin have shown protective effects and have significantly improved the outcomes of mice suffering from sepsis and ARDS. Here, we review researches related to the pathological role of histone in context of sepsis and ARDS and evaluate the potential value of histones as biomarkers and therapeutic targets of these diseases.

  10. Sepsis and ARDS: The Dark Side of Histones.

    Science.gov (United States)

    Xu, Zhiheng; Huang, Yongbo; Mao, Pu; Zhang, Jianrong; Li, Yimin

    2015-01-01

    Despite advances in management over the last several decades, sepsis and acute respiratory distress syndrome (ARDS) still remain major clinical challenges and the leading causes of death for patients in intensive care units (ICUs) due to insufficient understanding of the pathophysiological mechanisms of these diseases. However, recent studies have shown that histones, also known as chromatin-basic structure proteins, could be released into the extracellular space during severe stress and physical challenges to the body (e.g., sepsis and ARDS). Due to their cytotoxic and proinflammatory effects, extracellular histones can lead to excessive and overwhelming cell damage and death, thus contributing to the pathogenesis of both sepsis and ARDS. In addition, antihistone-based treatments (e.g., neutralizing antibodies, activated protein C, and heparin) have shown protective effects and have significantly improved the outcomes of mice suffering from sepsis and ARDS. Here, we review researches related to the pathological role of histone in context of sepsis and ARDS and evaluate the potential value of histones as biomarkers and therapeutic targets of these diseases.

  11. Myc localizes to histone locus bodies during replication in Drosophila.

    Directory of Open Access Journals (Sweden)

    Kaveh Daneshvar

    Full Text Available Myc is an important protein at the center of multiple pathways required for growth and proliferation in animals. The absence of Myc is lethal in flies and mice, and its over-production is a potent inducer of over-proliferation and cancer. Myc protein is localized to the nucleus where it executes its many functions, however the specific sub-nuclear localization of Myc has rarely been reported. The work we describe here began with an observation of unexpected, punctate spots of Myc protein in certain regions of Drosophila embryos. We investigated the identity of these puncta and demonstrate that Myc is co-localized with coilin, a marker for sub-nuclear organelles known as Cajal Bodies (CBs, in embryos, larvae and ovaries. Using antibodies specific for U7 snRNP component Lsm11, we show that the majority of Myc and coilin co-localization occurs in Histone Locus Bodies (HLBs, the sites of histone mRNA transcription and processing. Furthermore, Myc localizes to HLBs only during replication in mitotic and endocycling cells, suggesting that its role there relates to replication-dependent canonical histone gene transcription. These results provide evidence that sub-nuclear localization of Myc is cell-cycle dependent and potentially important for histone mRNA production and processing.

  12. Histone deacetylase-mediated morphological transition in Candida albicans.

    Science.gov (United States)

    Kim, Jueun; Lee, Ji-Eun; Lee, Jung-Shin

    2015-12-01

    Candida albicans is the most common opportunistic fungal pathogen, which switches its morphology from single-cell yeast to filament through the various signaling pathways responding to diverse environmental cues. Various transcriptional factors such as Nrg1, Efg1, Brg1, Ssn6, and Tup1 are the key components of these signaling pathways. Since C. albicans can regulate its transcriptional gene expressions using common eukaryotic regulatory systems, its morphological transition by these signaling pathways could be linked to the epigenetic regulation by chromatin structure modifiers. Histone proteins, which are critical components of eukaryotic chromatin structure, can regulate the eukaryotic chromatin structure through their own modifications such as acetylation, methylation, phosphorylation and ubiquitylation. Recent studies revealed that various histone modifications, especially histone acetylation and deacetylation, participate in morphological transition of C. albicans collaborating with well-known transcription factors in the signaling pathways. Here, we review recent studies about chromatin-mediated morphological transition of C. albicans focusing on the interaction between transcription factors in the signaling pathways and histone deacetylases.

  13. Histone Acetylation Inhibitors Promote Axon Growth in Adult DRG neurons

    Science.gov (United States)

    Lin, Shen; Nazif, Kutaiba; Smith, Alexander; Baas, Peter W; Smith, George M

    2015-01-01

    Intrinsic mechanisms that guide damaged axons to regenerate following spinal cord injury remain poorly understood. Manipulation of posttranslational modifications of key proteins in mature neurons could re-invigorate growth machinery after injury. One such modification is acetylation, a reversible process controlled by two enzyme families acting in opposition, the Histone Deacetylases (HDACs) and the Histone Acetyl Transferases (HATs). While acetylated histones in the nucleus is associated with upregulation of growth promoting genes, de-acetylated tubulin in the axoplasm is associated with more labile microtubules, conducive to axon growth. In this study we investigated the effects of HAT inhibitors and HDAC inhibitors on cultured adult dorsal root ganglia (DRG) neurons. We found that inhibition of HATs, using Anacardic Acid or CPTH2, improved axon outgrowth, while inhibition of HDACs using TSA or Tubacin, inhibited axon growth. Furthermore, Anacardic Acid increased the number of axons able to cross an inhibitory chondroitin sulfate proteoglycan (CSPG) border. Histone acetylation, but not tubulin acetylation levels, was affected by HAT inhibitors, whereas tubulin acetylation levels were increased in the presence of HDAC inhibitor Tubacin. Although microtubule stabilizing drug taxol did not have an effect on the lengths of DRG axons, nocodazole decreased axon lengths. While the mechanistic basis will require future studies, our data show that inhibitors of HAT can augment axon growth in adult DRG neurons, with the potential of aiding axon growth over inhibitory substrates produced by the glial scar. PMID:25702820

  14. Relationship of histone acetylation to DNA topology and transcription.

    Science.gov (United States)

    Krajewski, W A; Luchnik, A N

    1991-12-01

    An autonomously replicating plasmid constructed from bovine papiloma virus (BPV) and pBR322 was stably maintained as a nuclear episome in a mouse cell culture. Addition to a cell culture of sodium butyrate (5 mM) induced an increase in plasmid DNA supercoiling of 3-5 turns, an increase in acetylation of cellular histones, and a decrease in plasmid transcription by 2- to 4-fold. After withdrawal of butyrate, DNA supercoiling began to fluctuate in a wave-like manner with an amplitude of up to 3 turns and a period of 3-4 h. These waves gradually faded by 24 h. The transcription of the plasmid and acetylation of cellular histones also oscillated with the same period. The wave-like alterations were not correlated with the cell cycle, for there was no resumption of DNA replication after butyrate withdrawal for at least 24 h. In vitro chemical acetylation of histones with acetyl adenylate also led to an increase in the superhelical density of plasmid DNA. The parallel changes in transcription, histone acetylation, and DNA supercoiling in vivo may indicate a functional innerconnection. Also, the observed in vivo variation in the level of DNA supercoiling directly indicates the possibility of its natural regulation in eukaryotic cells.

  15. Depletion of the histone chaperone tNASP inhibits proliferation and induces apoptosis in prostate cancer PC-3 cells

    Directory of Open Access Journals (Sweden)

    Tsuruta James K

    2011-04-01

    Full Text Available Abstract Background NASP (Nuclear Autoantigenic Sperm Protein is a histone chaperone that is present in all dividing cells. NASP has two splice variants: tNASP and sNASP. Only cancer, germ, transformed, and embryonic cells have a high level of expression of the tNASP splice variant. We examined the consequences of tNASP depletion for prostate cancer PC-3 cells. Methods tNASP was depleted from prostate cancer PC-3 cells, cervical cancer HeLa cells, and prostate epithelial PWR-1E cells using lentivirus expression of tNASP shRNA. Cell cycle changes were studied by proliferation assay with CFSE labeling and double thymidine synchronization. Gene expression profiles were detected using RT2Profiler PCR Array, Western and Northern blotting. Results PC-3 and HeLa cells showed inhibited proliferation, increased levels of cyclin-dependant kinase inhibitor p21 protein and apoptosis, whereas non-tumorigenic PWR-1E cells did not. All three cell types showed decreased levels of HSPA2. Supporting in vitro experiments demonstrated that tNASP, but not sNASP is required for activation of HSPA2. Conclusions Our results demonstrate that PC-3 and HeLa cancer cells require tNASP to maintain high levels of HSPA2 activity and therefore viability, while PWR-1E cells are unaffected by tNASP depletion. These different cellular responses most likely arise from changes in the interaction between tNASP and HSPA2 and disturbed tNASP chaperoning of linker histones. This study has demonstrated that tNASP is critical for the survival of prostate cancer cells and suggests that targeting tNASP expression can lead to a new approach for prostate cancer treatment.

  16. Comprehensive histone phosphorylation analysis and identification of Pf14-3-3 protein as a histone H3 phosphorylation reader in malaria parasites.

    Directory of Open Access Journals (Sweden)

    Eeshita G Dastidar

    Full Text Available The important role of histone posttranslational modifications, particularly methylation and acetylation, in Plasmodium falciparum gene regulation has been established. However, the role of histone phosphorylation remains understudied. Here, we investigate histone phosphorylation utilizing liquid chromatography and tandem mass spectrometry to analyze histones extracted from asexual blood stages using two improved protocols to enhance preservation of PTMs. Enrichment for phosphopeptides lead to the detection of 14 histone phospho-modifications in P. falciparum. The majority of phosphorylation sites were observed at the N-terminal regions of various histones and were frequently observed adjacent to acetylated lysines. We also report the identification of one novel member of the P. falciparum histone phosphosite binding protein repertoire, Pf14-3-3I. Recombinant Pf14-3-3I protein bound to purified parasite histones. In silico structural analysis of Pf14-3-3 proteins revealed that residues responsible for binding to histone H3 S10ph and/or S28ph are conserved at the primary and the tertiary structure levels. Using a battery of H3 specific phosphopeptides, we demonstrate that Pf14-3-3I preferentially binds to H3S28ph over H3S10ph, independent of modification of neighbouring residues like H3S10phK14ac and H3S28phS32ph. Our data provide key insight into histone phosphorylation sites. The identification of a second member of the histone modification reading machinery suggests a widespread use of histone phosphorylation in the control of various nuclear processes in malaria parasites.

  17. Comprehensive histone phosphorylation analysis and identification of Pf14-3-3 protein as a histone H3 phosphorylation reader in malaria parasites.

    Science.gov (United States)

    Dastidar, Eeshita G; Dzeyk, Kristina; Krijgsveld, Jeroen; Malmquist, Nicholas A; Doerig, Christian; Scherf, Artur; Lopez-Rubio, Jose-Juan

    2013-01-01

    The important role of histone posttranslational modifications, particularly methylation and acetylation, in Plasmodium falciparum gene regulation has been established. However, the role of histone phosphorylation remains understudied. Here, we investigate histone phosphorylation utilizing liquid chromatography and tandem mass spectrometry to analyze histones extracted from asexual blood stages using two improved protocols to enhance preservation of PTMs. Enrichment for phosphopeptides lead to the detection of 14 histone phospho-modifications in P. falciparum. The majority of phosphorylation sites were observed at the N-terminal regions of various h