Genome-wide patterns of histone modifications in fission yeast

We have used oligonucleotide tiling arrays to construct genome-wide high-resolution histone acetylation maps for fission yeast. The maps are corrected for nucleosome density and reveal surprisingly uniform patterns of modifications for five different histone acetylation sites. We found that histone acetylation and methylation patterns are generally polar, i.e. they change as a function of distance from the ATG codon. A typical fission yeast gene shows a distinct peak of histone acetylation around the ATG and gradually decreased acetylation levels in the coding region. The patterns are independent of gene length but dependent on the gene expression levels. H3K9Ac shows a stronger peak near the ATG and is more reduced in the coding regions of genes with high expression compared with genes with low expression levels. H4K16Ac is strongly reduced in coding regions of highly expressed genes. A second microarray platform was used to confirm the 5′ to 3′ polarity effects observed with tiling microarrays. By comparing coding region histone acetylation data in HDAC mutants and wild type, we found that hos2 affects primarily the 5′ regions, sir2 and clr6 affect middle regions, and clr6 affects 3′ regions. Thus, mechanisms involving different HDACs modulate histone acetylation levels to maintain a 5′ to 3′ polarity within the coding regions. Electronic Supplementary Material Supplementary material is available for this article at and accessible for authorised users.     

Fission yeast CENP-B homologs nucleate centromeric heterochromatin by promoting heterochromatin-specific histone tail modifications

Heterochromatin is a functionally important chromosomal component, especially at centromeres. In fission yeast, conserved heterochromatin-specific modifications of the histone H3 tail, involving deacetylation of Lys 9 and Lys 14 and subsequent methylation of Lys 9, promote the recruitment of a heterochromatin protein, Swi6, a homolog of the Drosophila heterochromatin protein 1. However, the primary determinants of the positioning of heterochromatin are still unclear. The fission yeast proteins Abp1, Cbh1, and Cbh2 are homologs of the human protein CENP-B that bind to centromeric alpha-satellite DNA and associate with centromeric heterochromatin. We show that the CENP-B homologs are functionally redundant at centromeres, and that Abp1 binds specifically to centromeric heterochromatin. In the absence of Abp1 or Cbh1, the centromeric association of Swi6 is diminished, resulting in a decrease in silencing of the region. CENP-B-homolog double disruptants show a synergistic reduction of Swi6 at centromeric heterochromatin, indicating that the three proteins are functionally redundant in the recruitment of Swi6. Furthermore, using chromatin immunoprecipitation assays, we show that disruption of CENP-B homologs causes a decrease in heterochromatin-specific modifications of histone H3. These results indicate that the CENP-B homologs act as site-specific nucleation factors for the formation of centromeric heterochromatin by heterochromatin-specific modifications of histone tails.     

Broad chromosomal domains of histone modification patterns in C. elegans

Chromatin immunoprecipitation identifies specific interactions between genomic DNA and proteins, advancing our understanding of gene-level and chromosome-level regulation. Based on chromatin immunoprecipitation experiments using validated antibodies, we define the genome-wide distributions of 19 histone modifications, one histone variant, and eight chromatin-associated proteins in Caenorhabditis elegans embryos and L3 larvae. Cluster analysis identified five groups of chromatin marks with shared features: Two groups correlate with gene repression, two with gene activation, and one with the X chromosome. The X chromosome displays numerous unique properties, including enrichment of monomethylated H4K20 and H3K27, which correlate with the different repressive mechanisms that operate in somatic tissues and germ cells, respectively. The data also revealed striking differences in chromatin composition between the autosomes and between chromosome arms and centers. Chromosomes I and III are globally enriched for marks of active genes, consistent with containing more highly expressed genes, compared to chromosomes II, IV, and especially V. Consistent with the absence of cytological heterochromatin and the holocentric nature of C. elegans chromosomes, markers of heterochromatin such as H3K9 methylation are not concentrated at a single region on each chromosome. Instead, H3K9 methylation is enriched on chromosome arms, coincident with zones of elevated meiotic recombination. Active genes in chromosome arms and centers have very similar histone mark distributions, suggesting that active domains in the arms are interspersed with heterochromatin-like structure. These data, which confirm and extend previous studies, allow for in-depth analysis of the organization and deployment of the C. elegans genome during development.     

Drosophila GAGA factor directs histone H3.3 replacement that prevents the heterochromatin spreading

Epigenetic maintenance of the expression state of the genome is critical for development. Drosophila GAGA factor interacts with FACT and modulates chromatin structure for the maintenance of gene expression. Here we show that the GAGA factor-FACT complex and its binding site just downstream from the white gene are crucial for position effect variegation. Interestingly there is a dip of histone H3 Lys 9 methylation and a peak of H3 Lys 4 methylation at this site. The GAGA factor and FACT direct replacement of histone H3 by H3.3 through association of HIRA at this site, and maintain white expression under the heterochromatin environment. Based on these findings we propose that the GAGA factor and FACT-dependent replacement of Lys 9-methylated histone H3 by H3.3 counteracts the spreading of silent chromatin.     

Plasticity of Social Behavior in Drosophila

This article presents results obtained from studies of the plasticity of changes in social behavior in Drosophila (interactions between individuals in groups) in conditions of homo- and heterogeneous environments. This is the first report of data illustrating self-starting acquisition by female Drosophila of a classical conditioned reflex to contextual factors signaling possible threats from other individuals and blocking the initiation of activity. A previously described operant conditioned reflex also helped flies avoid aggression from other individuals and make more efficient use of food resources by decreasing the initially high level of activity. Classical conditioning had the effect that the fly did not need to repeat acquisition of the conditioned reflex each time: when placed into an analogous situation, the fly's activity automatically decreased as a result of exposure to the conditioned stimulus, i.e., contextual factors.     

A histone code in meiosis: the histone kinase, NHK-1, is required for proper chromosomal architecture in Drosophila oocytes

To promote faithful propagation of the genetic material during sexual reproduction, meiotic chromosomes undergo specialized morphological changes that ensure accurate segregation of homologous chromosomes. The molecular mechanisms that establish the meiotic chromosomal structures are largely unknown. We describe a mutation in a recently identified Histone H2A kinase, nhk-1, in Drosophila that leads to female sterility due to defects in the formation of the meiotic chromosomal structures. The metaphase I arrest and the karyosome, a critical prophase I chromosomal structure, require nucleosomal histone kinase-1 (NHK-1) function. The defects are a result of failure to disassemble the synaptonemal complex and to load condensin onto the mutant chromosomes. Embryos laid by nhk-1-/- mutant females arrest with aberrant polar bodies and mitotic spindles, revealing that mitosis is affected as well. We analyzed the role of Histone H2A phosphorylation with respect to the histone code hypothesis and found that it is required for acetylation of Histone H3 and Histone H4 in meiosis. These studies reveal a critical role for histone modifications in chromosome dynamics in meiosis and mitosis.     

Inheritance of Polycomb-dependent chromosomal interactions in Drosophila

Maintenance of cell identity is a complex task that involves multiple layers of regulation, acting at all levels of chromatin packaging, from nucleosomes to folding of chromosomal domains in the cell nucleus. Polycomb-group (PcG) and trithorax-group (trxG) proteins maintain memory of chromatin states through binding at cis-regulatory elements named PcG response elements or cellular memory modules. Fab-7 is a well-defined cellular memory module involved in regulation of the homeotic gene Abdominal-B (Abd-B). In addition to its action in cis, we show here by three-dimensional FISH that the Fab-7 element leads to association of transgenes with each other or with the endogenous Fab-7, even when inserted in different chromosomes. These long-distance interactions enhance PcG-mediated silencing. They depend on PcG proteins, on DNA sequence homology, and on developmental progression. Once long-distance pairing is abolished by removal of the endogenous Fab-7, the derepressed chromatin state induced at the transgene locus can be transmitted through meiosis into a large fraction of the progeny, even after reintroduction of the endogenous Fab-7. Strikingly, meiotic inheritance of the derepressed state involves loss of pairing between endogenous and transgenic Fab-7. This suggests that transmission of nuclear architecture through cell division might contribute to inheritance of chromatin states in eukaryotes.     

Genome-wide analysis of histone modifications in human pancreatic islets

The global diabetes epidemic poses a major challenge. Epigenetic events contribute to the etiology of diabetes; however, the lack of epigenomic analysis has limited the elucidation of the mechanistic basis for this link. To determine the epigenetic architecture of human pancreatic islets we mapped the genome-wide locations of four histone marks: three associated with gene activation—H3K4me1, H3K4me2, and H3K4me3—and one associated with gene repression, H3K27me3. Interestingly, the promoters of the highly transcribed insulin and glucagon genes are occupied only sparsely by H3K4me2 and H3K4me3. Globally, we identified important relationships between promoter structure, histone modification, and gene expression. We demonstrated co-occurrences of histone modifications including bivalent marks in mature islets. Furthermore, we found a set of promoters that is differentially modified between islets and other cell types. We also use our histone marks to determine which of the known diabetes-associated single-nucleotide polymorphisms are likely to be part of regulatory elements. Our global map of histone marks will serve as an important resource for understanding the epigenetic basis of type 2 diabetes.Genetic and epigenetic factors determine cell fate and function. Recent breakthroughs in genotyping technology have led to the identification of more than 20 loci associated with the risk of type 2 diabetes (Sambuy 2007; Zhao et al. 2009). However, all together these loci explain <5% of the genetic risk for diabetes. Epigenetic events have been implicated as contributing factors for metabolic diseases (Barker 1988; Kaput et al. 2007). Unhealthy diet and a sedentary lifestyle likely lead to epigenetic changes that can, in turn, contribute to the onset of diabetes (Kaput et al. 2007). At present, the underlying molecular mechanisms for disease progression remain to be elucidated.Epigenetic modifications encompass both DNA methylation and histone modifications (Cedar and Bergman 2009). In recent years, genome-wide maps of epigenetic marks have been generated for yeast (Pokholok et al. 2005) and several cell types in mice and humans (Bernstein et al. 2005; Roh et al. 2006; Barski et al. 2007; Mikkelsen et al. 2007; Pan et al. 2007; Zhao et al. 2007). However, no genome-wide map of histone modifications has been reported for the human pancreatic islet, a key player in the etiology of diabetes. In the present study, we have used chromatin immunoprecipitation with massively parallel sequencing (ChIP-seq) technology to create a genome-wide map of four histone modifications associated with gene activation or repression in human pancreatic islets.     

The roles of histone modifications and small RNA in centromere function

Here, epigenetic regulation of centromeric chromatin in fission yeast (Schizosaccharomyces pombe) is reviewed, focussing on the role of histone modifications and the link to RNA interference (RNAi). Fission yeast centromeres are organized into two structurally and functionally distinct domains, both of which are required for centromere function. The central core domain anchors the kinetochore structure while the flanking heterochromatin domain is important for sister centromere cohesion. The chromatin structure of both domains is regulated epigenetically. In the central core domain, the histone H3 variant Cnp1(CENP-A) plays a key role. In the flanking heterochromatin domain, histones are kept underacetylated by the histone deacetylases (HDACs) Clr3, Clr6 and Sir2, and methylated by Clr4 methyltransferase (HMTase) to create a specific binding site for the Swi6 protein. Swi6 then directly mediates cohesin binding to the centromeric heterochromatin. Recently, a surprising link was made between heterochromatin formation and RNAi.     

Post-translational modifications of Trypanosoma cruzi histone H4

Histone tails provide sites for a variety of post-translational modifications implicated in the control of gene expression and chromatin assembly. As both histones and control of gene expression in trypanosomes are highly divergent compared to most eukaryotes, post-translational modifications of Trypanosoma cruzi histones were investigated. After in vivo incubation of live parasites with radiolabeled precursors, histone H4 mainly incorporates [³H]-acetyl, and to a lesser extent [³H]-methyl residues. In contrast, histone H3 preferentially incorporates [³H]-methyl residues. The modifications of histone H4 were further characterized by mass spectrometry. MALDI-TOF–TOF-MS analysis revealed that peptides from histone H4 amino-terminus, obtained by either endoproteinase Glu-C or endoproteinase Arg-C digestion, contain isoforms with 14 and 42 Da additions, suggesting the presence of simultaneous acetylations and/or methylations. Tandem mass spectrometry analysis demonstrated that the N-terminal alanine is methylated, and lysine residues at positions 4, 10, 14 and 57 are acetylated; lysine at position 18 is mono-methylated, while arginine at position 53 is dimethylated. Immunoblotting analyses using specific antibodies raised against synthetic and acetylated peptides of T. cruzi histone H4 indicate that lysine 4 is acetylated in the majority of histone H4, while other acetylations at the N-terminus portion of histone H4 are less abundant.     

Age-dependent chromosomal distribution of male-biased genes in Drosophila

We investigated the correlation between the chromosomal location and age distribution of new male-biased genes formed by duplications via DNA intermediates (DNA-level) or by de novo origination in Drosophila. Our genome-wide analysis revealed an excess of young X-linked male-biased genes. The proportion of X-linked male-biased genes then diminishes through time, leading to an autosomal excess of male-biased genes. The switch between X-linked and autosomal enrichment of male-biased genes was also present in the distribution of both protein-coding genes on the D. pseudoobscura neo-X chromosome and microRNA genes of D. melanogaster. These observations revealed that the evolution of male-biased genes is more complicated than the previously detected one-step X→A gene traffic and the enrichment of the male-biased genes on autosomes. The pattern we detected suggests that the interaction of various evolutionary forces such as the meiotic sex chromosome inactivation (MSCI), faster-X effect, and sexual antagonism in the male germline might have shaped the chromosomal distribution of male-biased genes on different evolutionary time scales.It has been observed that male-biased genes in Drosophila are overrepresented on autosomes (Parisi et al. 2003; Ranz et al. 2003). Consistent with this result, a dynamic process that can explain the nonrandom autosomal distribution has also been observed, in which autosomal new genes with X-linked parental genes are often male-biased. Specifically, a significant excess of autosomal testis-expressed retrogenes were identified as RNA-duplicates of X-linked parental genes (Betran et al. 2002). Recently, similar X→A gene traffic was observed in the DNA-level duplication and relocation data set of the Drosophila genus (Vibranovski et al. 2009b), and was further confirmed for DNA-level duplications in the D. pseudoobscura neo-X chromosome (Meisel et al. 2009). In addition, selective extinction of neo-X linked male-biased genes also occurred in D. pseudoobscura (Sturgill et al. 2007). These three lines of genome-wide investigation support a common pattern of out-of-X traffic for male-biased genes, resulting in an enrichment of these genes on autosomes in the long term.Various hypotheses have been proposed to explain the chromosomal distribution of sex-biased genes. The sexual antagonism hypothesis (Rice 1984) predicts an increase of X-linked sexually antagonistic genes in a population over a wide range of parameters (e.g., recessive mutations advantageous for males and disadvantageous for females). With the assumption of a modifier gene that may restrict expression to the advantageous sex, X-linked antagonistic genes would be more likely to be involved in the evolution of sexually dimorphic traits (Rice 1984; Vicoso and Charlesworth 2006). In autosomal inheritance, when the advantageous effect on one sex is greater than the disadvantageous effect on the other sex, sexually antagonistic genes can spread in the population (Rice 1984). Although sexual antagonism has often been used to explain the evolution of sex-biased genes (Betran et al. 2002; Parisi et al. 2003; Ranz et al. 2003; Meisel et al. 2009), recent reports questioned the association between sex-biased expression and sexual antagonism (Vicoso and Charlesworth 2009; Innocenti and Morrow 2010). On the other hand, meiotic sex chromosome inactivation (MSCI) in spermatogenesis predicts X demasculinization via selection favoring autosomal male-biased genes, if the X chromosome is inactive during meiosis (Lifschytz and Lindsley 1972; Betran et al. 2002; Hense et al. 2007; Vibranovski et al. 2009a, 2010). Thus, if sex-biased expression were a proxy for sexual antagonism, sexually antagonistic selection is compatible with a paucity of X-linked somatic male-biased genes (Parisi et al. 2003). Additionally, the existence of MSCI-based selection is supported by the recent finding that autosomal genes retroposed from the X chromosome frequently demonstrate complementary expression in meiosis compared to their X-linked parental genes (Vibranovski et al. 2009a).However, a number of X-linked evolutionarily young genes have been identified recently (Arguello et al. 2006; Levine et al. 2006; Chen et al. 2007), all of which are male-biased. In order to understand whether these cases represent a general pattern, we examined the chromosomal distribution of male-biased genes of different evolutionary ages. Unexpectedly, we observed that the X chromosome has undergone an initial enrichment of young male-biased genes through intrachromosomal origination. However, as gene age increases, this excess gradually diminishes and is finally reversed, resulting in an overrepresentation of male-biased genes on the autosomes. This dynamic suggests a significant impact of the evolutionary time scale on the different mechanisms responsible for the evolution of male-biased genes. Here, we discuss how different evolutionary forces may impact the chromosomal distribution of male-biased genes with different ages.     

DNA甲基化与组蛋白修饰在自身免疫性疾病中的研究进展

自身免疫性疾病(AID)是一类由异常免疫反应引起的对健康组织和细胞造成严重损害的慢性炎症性疾病.包括系统系红斑狼疮(SLE)、类风湿性关节炎(RA)、系统性硬化病(SSc)以及急性冠脉综合征(ACS)等.其发病机制尚未明确,近年来研究表明,表观遗传在AID的发病机制中起重要作用.因此,进一步了解DNA甲基化、组蛋白修饰与AID的关系,为AID的预防和治疗提供了新线索.