共查询到20条相似文献,搜索用时 46 毫秒
1.
Lauren P Blair Nathan L Avaritt Rong Huang Phillip A Cole Sean D Taverna Alan J Tackett 《Epigenetics》2011,6(4):490-499
In eukaryotes, DNA is wrapped around proteins called histones and is condensed into chromatin. Post-translational modification of histones can result in changes in gene expression. One of the most well-studied histone modifications is the methylation of lysine 4 on histone H3 (H3K4). This residue can be mono-, di- or tri-methylated and these varying methylation states have been associated with different levels of gene expression. Understanding exactly what the purpose of these methylation states is, in terms of gene expression, has been a topic of much research in recent years. Enzymes that can add (methyltransferases) and remove (demethylases) these modifications are of particular interest. The first demethylase discovered, LSD1, is the most well-classified and has been implicated in contributing to human cancers and to DNA damage response pathways. Currently, there are limited methods for accurately studying the activity of demethylases in vitro or in vivo. In this work, we present MassSQUIRM (mass spectrometric quantitation using isotopic reductive methylation), a quantitative method for studying the activity of demethylases capable of removing mono- and di-methyl marks from lysine residues. We focus specifically on LSD1 due to its potential as a prime therapeutic target for human disease. This quantitative approach will enable better characterization of the activity of LSD1 and other chromatin modifying enzymes in vitro, in vivo or in response to inhibitors.Key words: LSD1, lysine demethylase, mass spectrometry, reductive methylation, monoamine oxidase (MAO) inhibitors 相似文献
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《Epigenetics》2013,8(4):490-499
In eukaryotes, DNA is wrapped around proteins called histones and is condensed into chromatin. Post-translational modification of histones can result in changes in gene expression. One of the most well-studied histone modifications is the methylation of lysine 4 on histone H3 (H3K4). This residue can be mono-, di- or tri-methylated and these varying methylation states have been associated with different levels of gene expression. Understanding exactly what the purpose of these methylation states is, in terms of gene expression, has been a topic of much research in recent years. Enzymes that can add (methyltransferases) and remove (demethylases) these modifications are of particular interest. The first demethylase discovered, LSD1, is the most well-classified and has been implicated in contributing to human cancers and to DNA damage response pathways. Currently, there are limited methods for accurately studying the activity of demethylases in vitro or in vivo. In this work, we present MassSQUIRM (mass spectrometric quantitation using isotopic reductive methylation), a quantitative method for studying the activity of demethylases capable of removing mono- and di-methyl marks from lysine residues. We focus specifically on LSD1 due to its potential as a prime therapeutic target for human disease. This quantitative approach will enable better characterization of the activity of LSD1 and other chromatin modifying enzymes in vitro, in vivo or in response to inhibitors. 相似文献
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《Fly》2013,7(1):36-44
Epigenetic regulation of chromatin structure is a fundamental process for eukaryotes. Regulators include DNA methylation, microRNAs and chromatin modifications. Within the chromatin modifiers, one class of enzymes that can functionally bind and modify chromatin, through the removal of methyl marks, is the histone lysine demethylases. Here, we summarize the current findings of the 13 known histone lysine demethylases in Drosophila melanogaster, and discuss the critical role of these histone-modifying enzymes in the maintenance of genomic functions. Additionally, as histone demethylase dysregulation has been identified in cancer, we discuss the advantages for using Drosophila as a model system to study tumorigenesis. 相似文献
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Epigenetic regulation of chromatin structure is a fundamental process for eukaryotes. Regulators include DNA methylation, microRNAs and chromatin modifications. Within the chromatin modifiers, one class of enzymes that can functionally bind and modify chromatin, through the removal of methyl marks, is the histone lysine demethylases. Here, we summarize the current findings of the 13 known histone lysine demethylases in Drosophila melanogaster, and discuss the critical role of these histone-modifying enzymes in the maintenance of genomic functions. Additionally, as histone demethylase dysregulation has been identified in cancer, we discuss the advantages for using Drosophila as a model system to study tumorigenesis. 相似文献
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Wyatt W. Yue Wei Ge Christopher D.O. Cooper Kathryn L. Kavanagh Michael A. McDonough 《FEBS letters》2010,584(4):825-41625
Crystallographic analysis of the catalytic domain of PHD finger protein 8 (PHF8), an Nε-methyl lysine histone demethylase associated with mental retardation and cleft lip/palate, reveals a double-stranded β-helix fold with conserved Fe(II) and cosubstrate binding sites typical of the 2-oxoglutarate dependent oxygenases. The PHF8 active site is highly conserved with those of the FBXL10/11demethylases, which are also selective for the di-/mono-methylated lysine states, but differs from that of the JMJD2 demethylases which are selective for tri-/di-methylated states. The results rationalize the lack of activity for the clinically observed F279S PHF8 variant and they will help to identify inhibitors selective for specific Nε-methyl lysine demethylase subfamilies. 相似文献
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Histone methylation is involved in the regulation of many cellular processes. In the past 2 years, several histone demethylases including BHC110/LSD1 have been characterized. BHC110, the first known histone lysine demethylase, removes methyl groups from methylated histone H3 lysine 4 and has been found in many multi-protein complexes. Using one-step affinity purification, we have isolated enzymatically active BHC110-containing complexes. Here, we detail the methods used for the isolation and characterization of these histone demethylase complexes from a human stable cell line. 相似文献
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组蛋白甲基化修饰是一个可逆的动态的调节过程。甲基化和/或去甲基化状态与表观遗传、转录调控和维持基因组完整性等密切相关。组蛋白甲基化状态异常会直接或间接影响各种生理和病理过程。已知组蛋白去甲基化酶包括赖氨酸特异性去甲基化酶(LSD)家族和含JmjC结构域的JMJD家族。研究发现,两者与肿瘤的发生均有着密切的关系。本文总结了组蛋白去甲基化酶在组蛋白甲基化修饰及肿瘤研究方面的最新进展,为组蛋白修饰的功能及肿瘤诊断、治疗、预后监测等研究提供新思路。在胃癌、乳腺癌、结肠癌等常见肿瘤中,组蛋白去甲基化酶可改变组蛋白的甲基化水平或者直接作用于癌基因,也可调节microRNA或转录因子等,促进或抑制肿瘤的发生发展与影响肿瘤的预后。 相似文献
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The JmjC domain-containing histone demethylases can remove histone lysine methylation and thereby regulate gene expression. The JmjC domain uses iron Fe(II) and α-ketoglutarate (αKG) as cofactors in an oxidative demethylation reaction via hydroxymethyl lysine. We hypothesize that reactive oxygen species will oxidize Fe(II) to Fe(III), thereby attenuating the activity of JmjC domain-containing histone demethylases. To minimize secondary responses from cells, extremely short periods of oxidative stress (3 h) were used to investigate this question. Cells that were exposed to hydrogen peroxide (H2O2) for 3 h exhibited increases in several histone methylation marks including H3K4me3 and decreases of histone acetylation marks including H3K9ac and H4K8ac; preincubation with ascorbate attenuated these changes. The oxidative stress level was measured by generation of 2′,7′-dichlorofluorescein, GSH/GSSG ratio, and protein carbonyl content. A cell-free system indicated that H2O2 inhibited histone demethylase activity where increased Fe(II) rescued this inhibition. TET protein showed a decreased activity under oxidative stress. Cells exposed to a low-dose and long-term (3 weeks) oxidative stress also showed increased global levels of H3K4me3 and H3K27me3. However, these global methylation changes did not persist after washout. The cells exposed to short-term oxidative stress also appeared to have higher activity of class I/II histone deacetylase (HDAC) but not class III HDAC. In conclusion, we have found that oxidative stress transiently alters the epigenetic program process through modulating the activity of enzymes responsible for demethylation and deacetylation of histones. 相似文献
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The ε-amino group of lysine residues may be mono-, di- or tri-methylated by protein lysine methyltransferases. In the past few years it has been highly considered that methylation of both histone and non-histone proteins has fundamental role in development and progression of various human diseases. Thus, the establishment of tools to study lysine methylation that will distinguish between the different states of methylation is required to elucidate their cellular functions. The 3X malignant brain tumor domain (3XMBT) repeats of the Lethal(3)malignant brain tumor-like protein 1 (L3MBTL1) have been utilized in the past as an affinity reagent for the identification of mono- and di-methylated lysine residues on individual proteins and on a proteomic scale. Here, we have utilized the 3XMBT domain to develop an enzyme-linked immunosorbent assay (ELISA) that allows the high-throughput detection of 3XMBT binding to methylated lysines. We demonstrated that this system allows the detection of methylated peptides, methylated proteins and PKMT activity on both peptides and proteins. We also optimized the assay to detect 3XMBT binding in crude E. coli lysates which facilitated the high throughput screening of 3XMBT mutant libraries. We have utilized protein engineering tools and generated a double site saturation 3XMBT library of residues 361 and 411 that were shown before to be important for binding mono and di-methylated substrates and identified variants that can exclusively recognize only di-methylated peptides. Together, our results demonstrate a powerful new approach that will contribute to deeper understanding of lysine methylation biology and that can be utilized for the engineering of domains for specific binders of other post-translational modifications. 相似文献
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Jason R. Hickok Divya Vasudevan William E. Antholine Douglas D. Thomas 《The Journal of biological chemistry》2013,288(22):16004-16015
Methylation of lysine residues on histone tails is an important epigenetic modification that is dynamically regulated through the combined effects of methyltransferases and demethylases. The Jumonji C domain Fe(II) α-ketoglutarate family of proteins performs the majority of histone demethylation. We demonstrate that nitric oxide (•NO) directly inhibits the activity of the demethylase KDM3A by forming a nitrosyliron complex in the catalytic pocket. Exposing cells to either chemical or cellular sources of •NO resulted in a significant increase in dimethyl Lys-9 on histone 3 (H3K9me2), the preferred substrate for KDM3A. G9a, the primary methyltransferase acting on H3K9me2, was down-regulated in response to •NO, and changes in methylation state could not be accounted for by methylation in general. Furthermore, cellular iron sequestration via dinitrosyliron complex formation correlated with increased methylation. The mRNA of several histone demethylases and methyltransferases was also differentially regulated in response to •NO. Taken together, these data reveal three novel and distinct mechanisms whereby •NO can affect histone methylation as follows: direct inhibition of Jumonji C demethylase activity, reduction in iron cofactor availability, and regulation of expression of methyl-modifying enzymes. This model of •NO as an epigenetic modulator provides a novel explanation for nonclassical gene regulation by •NO. 相似文献
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Structural insights into histone lysine demethylation 总被引:1,自引:0,他引:1
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Both components of chromatin (DNA and histones) are subjected to dynamic postsynthetic covalent modifications. Dynamic histone lysine methylation involves the activities of modifying enzymes (writers), enzymes removing modifications (erasers), and readers of the epigenetic code. Known histone lysine demethylases include flavin-dependent monoamine oxidase lysine-specific demethylase 1 and α-ketoglutarate-Fe(II)-dependent dioxygenases containing Jumonji domains. Importantly, the Jumonji domain often associates with at least one additional recognizable domain (reader) within the same polypeptide that detects the methylation status of histones and/or DNA. Here, we summarize recent developments in characterizing structural and functional properties of various histone lysine demethylases, with emphasis on a mechanism of crosstalk between a Jumonji domain and its associated reader module(s). We further discuss the role of recently identified Tet1 enzyme in oxidizing 5-methylcytosine to 5-hydroxymethylcytosine in DNA. 相似文献
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Qingfeng Chen Xiangsong Chen Quan Wang Faben Zhang Zhiyong Lou Qifa Zhang Dao-Xiu Zhou 《PLoS genetics》2013,9(1)
Histone lysine methylation is an important epigenetic modification in regulating chromatin structure and gene expression. Histone H3 lysine 4 methylation (H3K4me), which can be in a mono-, di-, or trimethylated state, has been shown to play an important role in gene expression involved in plant developmental control and stress adaptation. However, the resetting mechanism of this epigenetic modification is not yet fully understood. In this work, we identified a JmjC domain-containing protein, JMJ703, as a histone lysine demethylase that specifically reverses all three forms of H3K4me in rice. Loss-of-function mutation of the gene affected stem elongation and plant growth, which may be related to increased expression of cytokinin oxidase genes in the mutant. Analysis of crystal structure of the catalytic core domain (c-JMJ703) of the protein revealed a general structural similarity with mammalian and yeast JMJD2 proteins that are H3K9 and H3K36 demethylases. However, several specific features were observed in the structure of c-JMJ703. Key residues that interact with cofactors Fe(II) and N-oxalylglycine and the methylated H3K4 substrate peptide were identified and were shown to be essential for the demethylase activity in vivo. Several key residues are specifically conserved in known H3K4 demethylases, suggesting that they may be involved in the specificity for H3K4 demethylation. 相似文献
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JMJD3 is a histone H3K27 demethylase 总被引:2,自引:0,他引:2
Histone methylation is an important epigenetic phenomenon that participates in a diverse array of cellular processes and has been found to be associated with cancer. Recent identification of several histone demethylases has proved that histone methylation is a reversible process. Through a candidate approach, we have biochemically identified JMJD3 as an H3K27 demethylase. Transfection of JMJD3 into HeLa cells caused a specific reduction oftrimethyl H3K27, but had no effect on di-and monomethyl H3K27, or histone lysine methylations on H3K4 and H3K9. The enzymatic activity requires the JmjC domain and the conserved histidine that has been suggested to be important for a cofactor binding. In vitro biochemical experiments demonstrated that JMJD3 directly catalyzes the demethylation. In addition, we found that JMJD3 is upregulated in prostate cancer, and its expression is higher in metastatic prostate cancer. Thus, we identified JMJD3 as a demethylase capable of removing the trimethyl group from histone H3 lysine 27 and upregulated in prostate cancer. 相似文献
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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 transmitted into daughter cells and through what mechanisms are
currently under active investigation. Previously, methylation was considered to be irreversible, but the recent discovery
of histone lysine demethylases 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, besides 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 progresses 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 相似文献