Histone deacetylation directs DNA methylation in survivin gene silencing

DNA methylation and histone acetylation are major epigenetic modifications in gene silencing. In our previous research, we found that the methylated oligonucleotide (SurKex) complementary to a region of promoter of survivin could induce DNA methylation in a site-specific manner leading to survivin silencing. Here, we further studied the role of histone acetylation in survivin silencing and the relationship between histone acetylation and DNA methylation.First we observed the levels of histone H4 and H4K16 acetylation that were decreased after SurKex treatment by using the chromatin immunoprecipitation (ChIP) assay. Next, we investigated the roles of histone acetylation and DNA methylation in survivin silencing after blockade of histone deacetylation with Trichostatin A (TSA). We assessed survivin mRNA expression by RT-PCR, measured survivin promoter methylation by bisulfite sequencing and examined the level of histone acetylation by the ChIP assay. The results showed that histone deacetylation blocked by TSA reversed the effects of SurKex on inhibiting the expression of survivin mRNA, inducing a site-specific methylation on survivin promoter and decreasing the level of histone acetylation. Finally, we examined the role of histone acetylation in the expression of DNA methyltransferase 1 (DNMT1) mRNA. The results showed that histone deacetylation blocked by TSA reversed the increasing effect of histone deacetylation on the expression of survivin mRNA. This study suggests that histone deacetylation guides SurKex-induced DNA methylation in survivin silencing possibly through increasing the expression of DNMT1 mRNA.     

Core histone acetylation during lymphocyte activation

Histone acetylation has been followed in cultures of human lymphocytes, in PHA-stimulated lymphocytes and in mixed lymphocytes obtained from identical twins and from unrelated donors. A computer assisted analysis of two-dimensional gels and autoradiograms revealed that in cultured lymphocytes only H3 and H4 core histones incorporate labeled acetate and that two H3 variants greatly differ in their rate of acetate uptake.     

Histone acetylation in gene regulation.

Genetic information is packaged in the highly dynamic nucleoprotein structure called chromatin. Many biological processes are regulated via post-translational modifications of key proteins. Acetylation of lysine residues at the N-terminal histone tails is one of the most studied covalent modifications influencing gene regulation in eukaryotic cells. This review focuses on the role of enzymes involved in controlling both histone and non-histone proteins acetylation levels in the cell, with particular emphasis on their effects on cancer.     

Histone acetylation facilitates association of nucleosomes with SET domain of ALL-1 methyltransferase in vitro

The inheritable methylation pattern of gene activity, created upon cell differentiation, is further maintained by the “SET” (methyltransferase)-domain proteins. However, it is still not clear how SET-proteins can decide on the required gene activity state and the way their chromatin association is maintained. Here we have found that high levels of histone acetylation - the hallmarks of active chromosome regions in vivo - can increase the affinity of reconstituted nucleosomes to the SET domain of ALL-1 histone methyltransferase in a defined system in vitro.     

Changes in histone acetylation during mouse oocyte meiosis

We examined global changes in the acetylation of histones in mouse oocytes during meiosis. Immunocytochemistry with specific antibodies against various acetylated lysine residues on histones H3 and H4 showed that acetylation of all the lysines decreased to undetectable or negligible levels in the oocytes during meiosis, whereas most of these lysines were acetylated during mitosis in preimplantation embryos and somatic cells. When the somatic cell nuclei were transferred into enucleated oocytes, the acetylation of lysines decreased markedly. This type of deacetylation was inhibited by trichostatin A, a specific inhibitor of histone deacetylase (HDAC), thereby indicating that HDAC is able to deacetylate histones during meiosis but not during mitosis. Meiosis-specific deacetylation may be a consequence of the accessibility of HDAC1 to the chromosome, because HDAC1 colocalized with the chromosome during meiosis but not during mitosis. As histone acetylation is thought to play a role in propagating the gene expression pattern to the descendent generation during mitosis, and the gene expression pattern of differentiated oocytes is reprogrammed during meiosis to allow the initiation of a new program by totipotent zygotes of the next generation, our results suggest that the oocyte cytoplasm initializes a program of gene expression by deacetylating histones.     

Effects of histone acetylation on chromatin structure

The effect of histone acetylation was monitored on CHO chromatin structure, following the addition of 7 mM Na-butyrate to the cell culture medium. The properties of both control and hyperacetylated chromatins and nuclei were investigated by circular dichroism, ethidium bromide intercalation, differential scanning calorimetry, and affinity chromatography. Our results are compatible with modest but significant alterations in the various levels of chromatin organization, as a result of the charge neutralization of some lysine residues within the N-terminal region of the histonic octamer. Namely, large statistically significant differences do exist in the heat capacity thermograms of native nuclei, where unfolding into single nucleofilament of the highly packed native chromatin superfiber appears associated with acetylation; at the same time CD, EB, and affinity chromatography point to modest but consistent differences in the compactness of isolated nucleosomes and polynucleosomes. J. Cell. Biochem. 64:466–475. © 1997 Wiley-Liss, Inc.     

流式细胞术检测组蛋白乙酰化水平方法的建立与应用

组蛋白去乙酰化酶抑制剂是近年来出现的一类新的抗肿瘤药物,在艾滋病等其他疾病中同样也受到关注。但是在基础和临床研究中,目前还缺乏统一可靠的组蛋白乙酰化水平的检测手段。本文利用全血和外周血单个核细胞,通过一系列的对比实验,比较了不同样品处理温度(冰上和室温)、破膜方法(细胞内因子染色破膜和核内因子染色破膜)、抗体剂量(抗体滴定)和抗体孵育时间(时间梯度)等实验条件对流式细胞术检测的影响,最终建立了一套基于流式细胞术的组蛋白乙酰化水平检测手段。同时,将优化后的流式细胞检测技术应用于西达本胺(目前国内唯一上市的组蛋白去乙酰化酶抑制剂)的体外实验和临床试验,结果均证明本文建立的组蛋白乙酰化流式细胞检测方法可以作为基础和临床研究中一个可靠、快速、便捷的检测手段。     

Regulation of histone acetylation during meiotic maturation in mouse oocytes

Histone acetylation is an important epigenetic modification implicated in the regulation of chromatin structure and, subsequently, gene expression. Global histone deacetylation was reported in mouse oocytes during meiosis but not mitosis. The regulation of this meiosis-specific deacetylation has not been elucidated. Here, we demonstrate that p34(cdc2) kinase activity and protein synthesis are responsible for the activation of histone deacetylases and the inhibition of histone acetyltransferases (HATs), respectively, resulting in deacetylation of histone H4 at lysine-12 (H4K12) during mouse oocyte meiosis. Temporal changes in the acetylation state of H4K12 were examined immunocytochemically during meiotic maturation using an antibody specific for acetylated H4K12. H4K12 was deacetylated during the first meiosis, temporarily acetylated around the time of the first polar body (PB1) extrusion, and then deacetylated again during the second meiosis. Because these changes coincided with the known oscillation pattern of p34(cdc2) kinase activity, we investigated the involvement of the kinase in H4K12 deacetylation. Roscovitine, an inhibitor of cyclin-dependent kinase activity, prevented H4K12 deacetylation during both the first and second meiosis, suggesting that p34(cdc2) kinase activity is required for deacetylation during meiosis. In addition, cycloheximide, a protein synthesis inhibitor, also prevented deacetylation. After PB1 extrusion, at which time H4K12 had been deacetylated, H4K12 was re-acetylated in the condensed chromosomes by treatment with cycloheximide but not with roscovitine. These results demonstrate that HATs are present but inactivated by newly synthesized protein(s) that is (are) not involved in p34(cdc2) kinase activity. Our results suggest that p34(cdc2) kinase activity induces the deacetylation of H4K12 and that the deacetylated state is maintained by newly synthesized protein(s) that inhibits HAT activity during meiosis.     

Targeting histone acetylation dynamics and oncogenic transcription by catalytic P300/CBP inhibition

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Maternal and paternal alleles exhibit differential histone methylation and acetylation at maize imprinted genes

Imprinting is an epigenetically controlled form of gene regulation in which the expression of a gene is based on its parent of origin. This epigenetic regulation is likely to involve allele-specific DNA or histone modifications. The relative abundance of eight different histone modifications was tested at various regions in several imprinted maize (Zea mays) genes using a chromatin immunoprecipitation protocol coupled with quantitative allele-specific single nucleotide polymorphism assays. Histone H3 lysine-27 di- and tri-methylation are paternally enriched at the imprinted loci Mez1, ZmFie1 and Nrp1. In contrast, acetylation of histones H3 and H4 and H3K4 dimethylation are enriched at the maternal alleles of these genes. Di- and tri-methylation of H3 lysine-9, which is generally associated with constitutively silenced chromatin, was not enriched at either allele of imprinted loci. These patterns of enrichment were specific to tissues that exhibit imprinting. In addition, the enrichment of these modifications was dependent upon the parental origin of an allele and not sequence differences between the alleles, as demonstrated by reciprocal crosses. This study presents a detailed view of the chromatin modifications that are associated with the maternal and paternal alleles at imprinted loci and provides evidence for common histone modifications at multiple imprinted loci.