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Sample records for histone h3 lysine9

  1. The putative oncogene GASC1 demethylates tri- and dimethylated lysine 9 on histone H3

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    Cloos, Paul A C; Christensen, Jesper; Agger, Karl

    2006-01-01

    Methylation of lysine and arginine residues on histone tails affects chromatin structure and gene transcription. Tri- and dimethylation of lysine 9 on histone H3 (H3K9me3/me2) is required for the binding of the repressive protein HP1 and is associated with heterochromatin formation and transcript...

  2. Acetylation on histone H3 lysine 9 mediates a switch from transcription initiation to elongation.

    Science.gov (United States)

    Gates, Leah A; Shi, Jiejun; Rohira, Aarti D; Feng, Qin; Zhu, Bokai; Bedford, Mark T; Sagum, Cari A; Jung, Sung Yun; Qin, Jun; Tsai, Ming-Jer; Tsai, Sophia Y; Li, Wei; Foulds, Charles E; O'Malley, Bert W

    2017-09-01

    The transition from transcription initiation to elongation is a key regulatory step in gene expression, which requires RNA polymerase II (pol II) to escape promoter proximal pausing on chromatin. Although elongation factors promote pause release leading to transcription elongation, the role of epigenetic modifications during this critical transition step is poorly understood. Two histone marks on histone H3, lysine 4 trimethylation (H3K4me3) and lysine 9 acetylation (H3K9ac), co-localize on active gene promoters and are associated with active transcription. H3K4me3 can promote transcription initiation, yet the functional role of H3K9ac is much less understood. We hypothesized that H3K9ac may function downstream of transcription initiation by recruiting proteins important for the next step of transcription. Here, we describe a functional role for H3K9ac in promoting pol II pause release by directly recruiting the super elongation complex (SEC) to chromatin. H3K9ac serves as a substrate for direct binding of the SEC, as does acetylation of histone H4 lysine 5 to a lesser extent. Furthermore, lysine 9 on histone H3 is necessary for maximal pol II pause release through SEC action, and loss of H3K9ac increases the pol II pausing index on a subset of genes in HeLa cells. At select gene promoters, H3K9ac loss or SEC depletion reduces gene expression and increases paused pol II occupancy. We therefore propose that an ordered histone code can promote progression through the transcription cycle, providing new mechanistic insight indicating that SEC recruitment to certain acetylated histones on a subset of genes stimulates the subsequent release of paused pol II needed for transcription elongation. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Nickel ions increase histone H3 lysine 9 dimethylation and induce transgene silencing.

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    Chen, Haobin; Ke, Qingdong; Kluz, Thomas; Yan, Yan; Costa, Max

    2006-05-01

    We have previously reported that carcinogenic nickel compounds decreased global histone H4 acetylation and silenced the gpt transgene in G12 Chinese hamster cells. However, the nature of this silencing is still not clear. Here, we report that nickel ion exposure increases global H3K9 mono- and dimethylation, both of which are critical marks for DNA methylation and long-term gene silencing. In contrast to the up-regulation of global H3K9 dimethylation, nickel ions decreased the expression and activity of histone H3K9 specific methyltransferase G9a. Further investigation demonstrated that nickel ions interfered with the removal of histone methylation in vivo and directly decreased the activity of a Fe(II)-2-oxoglutarate-dependent histone H3K9 demethylase in nuclear extract in vitro. These results are the first to show a histone H3K9 demethylase activity dependent on both iron and 2-oxoglutarate. Exposure to nickel ions also increased H3K9 dimethylation at the gpt locus in G12 cells and repressed the expression of the gpt transgene. An extended nickel ion exposure led to increased frequency of the gpt transgene silencing, which was readily reversed by treatment with DNA-demethylating agent 5-aza-2'-deoxycytidine. Collectively, our data strongly indicate that nickel ions induce transgene silencing by increasing histone H3K9 dimethylation, and this effect is mediated by the inhibition of H3K9 demethylation.

  4. Nickel Ions Increase Histone H3 Lysine 9 Dimethylation and Induce Transgene Silencing

    OpenAIRE

    Chen, Haobin; Ke, Qingdong; Kluz, Thomas; Yan, Yan; Costa, Max

    2006-01-01

    We have previously reported that carcinogenic nickel compounds decreased global histone H4 acetylation and silenced the gpt transgene in G12 Chinese hamster cells. However, the nature of this silencing is still not clear. Here, we report that nickel ion exposure increases global H3K9 mono- and dimethylation, both of which are critical marks for DNA methylation and long-term gene silencing. In contrast to the up-regulation of global H3K9 dimethylation, nickel ions decreased the expression and ...

  5. Ornithine decarboxylase antizyme induces hypomethylation of genome DNA and histone H3 lysine 9 dimethylation (H3K9me2 in human oral cancer cell line.

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    Daisuke Yamamoto

    2010-09-01

    Full Text Available Methylation of CpG islands of genome DNA and lysine residues of histone H3 and H4 tails regulates gene transcription. Inhibition of polyamine synthesis by ornithine decarboxylase antizyme-1 (OAZ in human oral cancer cell line resulted in accumulation of decarboxylated S-adenosylmethionine (dcSAM, which acts as a competitive inhibitor of methylation reactions. We anticipated that accumulation of dcSAM impaired methylation reactions and resulted in hypomethylation of genome DNA and histone tails.Global methylation state of genome DNA and lysine residues of histone H3 and H4 tails were assayed by Methylation by Isoschizomers (MIAMI method and western blotting, respectively, in the presence or absence of OAZ expression. Ectopic expression of OAZ mediated hypomethylation of CpG islands of genome DNA and histone H3 lysine 9 dimethylation (H3K9me2. Protein level of DNA methyltransferase 3B (DNMT3B and histone H3K9me specific methyltransferase G9a were down-regulated in OAZ transfectant.OAZ induced hypomethylation of CpG islands of global genome DNA and H3K9me2 by down-regulating DNMT3B and G9a protein level. Hypomethylation of CpG islands of genome DNA and histone H3K9me2 is a potent mechanism of induction of the genes related to tumor suppression and DNA double strand break repair.

  6. Ornithine decarboxylase antizyme induces hypomethylation of genome DNA and histone H3 lysine 9 dimethylation (H3K9me2) in human oral cancer cell line.

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    Yamamoto, Daisuke; Shima, Kaori; Matsuo, Kou; Nishioka, Takashi; Chen, Chang Yan; Hu, Guo-Fu; Sasaki, Akira; Tsuji, Takanori

    2010-09-03

    Methylation of CpG islands of genome DNA and lysine residues of histone H3 and H4 tails regulates gene transcription. Inhibition of polyamine synthesis by ornithine decarboxylase antizyme-1 (OAZ) in human oral cancer cell line resulted in accumulation of decarboxylated S-adenosylmethionine (dcSAM), which acts as a competitive inhibitor of methylation reactions. We anticipated that accumulation of dcSAM impaired methylation reactions and resulted in hypomethylation of genome DNA and histone tails. Global methylation state of genome DNA and lysine residues of histone H3 and H4 tails were assayed by Methylation by Isoschizomers (MIAMI) method and western blotting, respectively, in the presence or absence of OAZ expression. Ectopic expression of OAZ mediated hypomethylation of CpG islands of genome DNA and histone H3 lysine 9 dimethylation (H3K9me2). Protein level of DNA methyltransferase 3B (DNMT3B) and histone H3K9me specific methyltransferase G9a were down-regulated in OAZ transfectant. OAZ induced hypomethylation of CpG islands of global genome DNA and H3K9me2 by down-regulating DNMT3B and G9a protein level. Hypomethylation of CpG islands of genome DNA and histone H3K9me2 is a potent mechanism of induction of the genes related to tumor suppression and DNA double strand break repair.

  7. [Histone H3 lysine 9 methylation is associated with the expression of hMLH1 and DNA methylation in gastric cancer cells].

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    Meng, Chun-Feng; Zhu, Xin-Jiang; Peng, Guo

    2008-09-16

    To identify the association of his tone H3 lysine 9 (H3-K9) methylation with DNA methylation and the expression of the mismatch repair gene hMLH1 in human gastric cancer cells. Gastric cancer cells of the lines BGC-823 and MGC-803 were cultured and treated with 5-Aza-2'-deoxycytidine (5-Aza-dC), a demethylation agent, for 72 hour. Chromatin immunoprecipitation (ChIP) assay was used to assess the status of histone H3 lysine 9 methylation in the promoter regions of hMLH1 gene. Methylation-specific PCR (MSP) was used to evaluate the effect of 5-Aza-dC on DNA methylation status. RT-PCR was used to examine the hMLH1 gene expression. In the MGC-803 cells, silenced hMLH1 gene was characterized by DNA methylation and histone H3-K9 hypermethylation; 5-Aza-dC demethylated the DNA and reduced the histone H3-K9 methylation at silenced loci and resulted in reactivation of hMLH1 gene therein. Contrary to the MGC-803 cells, BGC-823 cells expressed hMLH1 gene with DNA demethylation and histone H3-K9 hypomethylation; and 5-Aza-dC had no effects on the gene expression, DNA methylation, and histone H3-K9 methylation therein. Hypermethylation of DNA in the promoter region is related to transcriptional silencing of hMLH1 gene. Histone H3-K9 methylation in different regions of the promoter studied correlates with DNA methylation status of hMLH1 gene in gastric cancer cells. Alteration of DNA methylation affects histone H3-K9 methylation. 5-Aza-dC can control hMLH1 expression, DNA methylation, and histone H3-K9 methylation in the promoter.

  8. Alteration of histone H3 lysine 9 dimethylation in peripheral white blood cells of septic patients with trauma and cancer.

    Science.gov (United States)

    Jiang, Linghui; Wang, Yiqin; Zhu, Duming; Xue, Zhanggang; Mao, Hailei

    2016-12-01

    The present study aimed to investigate the clinical significance of histone methylation in sepsis. A total of 43 blood samples from trauma and esophageal cancer patients with or without sepsis were collected. Immunofluorescence staining of isolated peripheral white blood cells (WBCs) was conducted. Co‑stained 293T cells served as a reference, to allow the levels of histone methylation in different types of WBCs from patients to be determined. Immunostaining analyses revealed different levels of histone 3 lysine 9 dimethylation (H3K9me2) in neutrophils (Neu), lymphocytes (Lym), and monocytes (Mon) from trauma patients. Compared with trauma patients, the levels of H3K9me2 were elevated in the three types of WBCs from cancer patients. When combined with sepsis, trauma patients demonstrated increased H3K9me2 levels in Neu (P=0.0005) and Mon (P=0.0002), whereas cancer patients had a significant decrease of H3K9me2 levels in the three types of WBCs (Neu, P=0.0003; Lym, P=0.007; Mon, P=0.007). The H3K9me2 alterations in patients with trauma and cancer were different with the occurrence of sepsis. A larger cohort study is warranted to explore the diagnostic significance and prognostic implications of altered histone methylation in septic patients.

  9. Cancer-associated upregulation of histone H3 lysine 9 trimethylation promotes cell motility in vitro and drives tumor formation in vivo.

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    Yokoyama, Yuhki; Hieda, Miki; Nishioka, Yu; Matsumoto, Ayaka; Higashi, Satomi; Kimura, Hiroshi; Yamamoto, Hirofumi; Mori, Masaki; Matsuura, Shuji; Matsuura, Nariaki

    2013-07-01

    Global histone modification patterns correlate with tumor phenotypes and prognostic factors in multiple tumor types. Recent studies suggest that aberrant histone modifications play an important role in cancer. However, the effects of global epigenetic rearrangements on cell functions remain poorly understood. In this study, we show that the histone H3 lysine 9 (H3K9) methyltransferase SUV39H1 is clearly involved in regulating cell migration in vitro. Overexpression of wild-type SUV39H1, but not enzymatically inactive SUV39H1, activated migration in breast and colorectal cancer cells. Inversely, migration was reduced by knockdown of SUV39H1 or chemical inhibition by chaetocin. In addition, H3K9 trimethylation (H3K9me3) was specifically increased in invasive regions of colorectal cancer tissues. Moreover, the presence of H3K9me3 positively correlated with lymph node metastasis in colorectal cancer patients. Furthermore, overexpression of SUV39H1 drove tumorigenesis in mouse, resulting in a considerable decrease in survival rate. These data indicate that H3K9 trimethylation plays an important role in human colorectal cancer progression, possibly by promoting collective cell invasion. © 2013 Japanese Cancer Association.

  10. Nicotinamide N-Methyltransferase Suppression Participates in Nickel-Induced Histone H3 Lysine9 Dimethylation in BEAS-2B Cells

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    Qian Li

    2017-04-01

    Full Text Available Background: Nickel compounds are well-established human carcinogens with weak mutagenic activity. Histone methylation has been proposed to play an important role in nickel-induced carcinogenesis. Nicotinamide N-methyltransferase (NNMT decreases histone methylation in several cancer cells by altering the cellular ratio of S-adenosylmethionine (SAM to S-adenosylhomocysteine (SAH. However, the role of NNMT in nickel-induced histone methylation remains unclear. Methods: BEAS-2B cells were exposed to different concentrations of nickel chloride (NiCl2 for 72 h or 200 μM NiCl2 for different time periods. Histone H3 on lysine 9 (H3K9 mono-, di-, and trimethylation and NNMT protein levels were measured by western blot analysis. Expressions of NNMT mRNA and the H3k9me2-associated genes, mitogen-activated protein kinase 3 (MAP2K3 and dickkopf1 (DKK1, were determined by qPCR analysis. The cellular ratio of nicotinamide adenine dinucleotide (NAD+ to reduced NAD (NADH and SAM/SAH ratio were determined. Results: Exposure of BEAS-2B cells to nickel increased H3K9 dimethylation (H3K9me2, suppressed the expressions of H3K9me2-associated genes (MAP2K3 and DKK1, and induced NNMT repression at both the protein and mRNA levels. Furthermore, over-expression of NNMT inhibited nickel-induced H3K9me2 and altered the cellular SAM/SAH ratio. Additionally, the NADH oxidant phenazine methosulfate (PMS not only reversed the nickel-induced reduction in NAD+/NADH but also inhibited the increase in H3K9me2. Conclusions: These findings indicate that the repression of NNMT may underlie nickel-induced H3K9 dimethylation by altering the cellular SAM/SAH ratio.

  11. Nicotinamide N-Methyltransferase Suppression Participates in Nickel-Induced Histone H3 Lysine9 Dimethylation in BEAS-2B Cells.

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    Li, Qian; He, Min-Di; Mao, Lin; Wang, Xue; Jiang, Yu-Lin; Li, Min; Lu, Yong-Hui; Yu, Zheng-Ping; Zhou, Zhou

    2017-01-01

    Nickel compounds are well-established human carcinogens with weak mutagenic activity. Histone methylation has been proposed to play an important role in nickel-induced carcinogenesis. Nicotinamide N-methyltransferase (NNMT) decreases histone methylation in several cancer cells by altering the cellular ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH). However, the role of NNMT in nickel-induced histone methylation remains unclear. BEAS-2B cells were exposed to different concentrations of nickel chloride (NiCl2) for 72 h or 200 μM NiCl2 for different time periods. Histone H3 on lysine 9 (H3K9) mono-, di-, and trimethylation and NNMT protein levels were measured by western blot analysis. Expressions of NNMT mRNA and the H3k9me2-associated genes, mitogen-activated protein kinase 3 (MAP2K3) and dickkopf1 (DKK1), were determined by qPCR analysis. The cellular ratio of nicotinamide adenine dinucleotide (NAD+) to reduced NAD (NADH) and SAM/SAH ratio were determined. Exposure of BEAS-2B cells to nickel increased H3K9 dimethylation (H3K9me2), suppressed the expressions of H3K9me2-associated genes (MAP2K3 and DKK1), and induced NNMT repression at both the protein and mRNA levels. Furthermore, over-expression of NNMT inhibited nickel-induced H3K9me2 and altered the cellular SAM/SAH ratio. Additionally, the NADH oxidant phenazine methosulfate (PMS) not only reversed the nickel-induced reduction in NAD+/NADH but also inhibited the increase in H3K9me2. These findings indicate that the repression of NNMT may underlie nickel-induced H3K9 dimethylation by altering the cellular SAM/SAH ratio. © 2017 The Author(s)Published by S. Karger AG, Basel.

  12. Downregulation of histone H3 lysine 9 methyltransferase G9a induces centrosome disruption and chromosome instability in cancer cells.

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    Yutaka Kondo

    Full Text Available BACKGROUND: Modifications of the histone amino-terminal tails affect access of regulatory factors and complexes to chromatin and thereby influence biological processes. Cancer cells are characterized by prominent epigenetic dysregulation, including histone modifications. However, the functional roles of the histone methyltransferases (HMT in cancer remain unclear. METHODOLOGY/PRINCIPAL FINDINGS: We studied RNAi-based inhibition (knockdown, KD of 2 different H3K9 HMTs, SUV39H1 and G9a. Knockdown of the 2 HMTs in PC3 cancer cell line markedly inhibited cell growth and caused profound morphological changes with loss of telomerase activity and shortened telomeres. SUV39H1 KD cells showed substantial increase in G2/M fraction. G9a KD cells showed increased DNA content (1.7-fold in 2 independent clones compared with FACS analyses to control. Karyotype analyses showed that this was due to an increased number of chromosomes (from 61 to 102 in G9a KD cells compared to parental PC3. Intriguingly, we found abnormal centrosome morphology and number in about 25% of the G9a KD cells, while centrosomes were morphologically normal in control cells. Microarray analyses after KD of SUV39H1 or G9a showed very few genes up-regulated among the 39,000 genes. The silenced tumor-suppressor genes p16 and RASSF1A were not activated in KD cells. CONCLUSIONS/SIGNIFICANCE: These data suggest that the 2 HMTs, SUV39H1 and G9a are required to perpetuate the malignant phenotype. Furthermore, G9a plays a critical role in regulating centrosome duplication presumably through chromatin structure rather than through affecting gene expression in cancer cells. Targeting these histone methyltransferases may be of therapeutic benefit in cancers.

  13. Promoter histone H3 lysine 9 di-methylation is associated with DNA methylation and aberrant expression of p16 in gastric cancer cells.

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    Meng, Chun-Feng; Zhu, Xin-Jiang; Peng, Guo; Dai, Dong-Qiu

    2009-11-01

    In the course of gastric cancer development, gene silencing by DNA hypermethylation is an important mechanism. While DNA methylation often co-exists with histone modifications to regulate gene expression, the function of histone modifications in gene silencing in gastric cancer has not been evaluated in detail. p16, a well-known tumor suppressor gene, is frequently silenced in DNA hypermethylation manner in gastric cancer. Accordingly, we chose p16 to clarify whether there is a correlation among histone H3 lysine 9 (H3-K9) di-methylation, H3-K9 acetylation, DNA methylation and p16 expression in human gastric cancer. Three gastric cancer cells, MKN-45, SGC-7901 and BGC-823, were treated with 5-aza-2'-deoxycytidine (5-Aza-dC) and/or trichostatin A (TSA). We investigated p16 promoter DNA methylation status, p16 mRNA levels, regional and global levels of di-methyl-H3-K9 and acetyl-H3-K9 in four groups: i) 5-Aza-dC, ii) TSA, iii) the combination of 5-Aza-dC and TSA and iv) control group with no treatments. p16 silencing is characterized by DNA hypermethylation, H3-K9 hypoacetylation and H3-K9 hypermethylation at the promoter region. Treatment with TSA, increased H3-K9 acetylation at the hypermethylated promoter, but did not affect H3-K9 di-methylation or p16 expression. By contrast, treatment with 5-Aza-dC, reduced H3-K9 di-methylation, increased H3-K9 acetylation at the hypermethylated promoter and reactivated the expression of p16. Combined treatment restored the expression of p16 synergistically. In addition, 5-Aza-dC and the combined treatment did not result in global alteration of H3-K9 di-methylation. These results suggest that H3-K9 di-methylation, H3-K9 acetylation and DNA methylation work in combination to silence p16 in gastric cancer. The decreased H3-K9 di-methylation correlates with DNA demethylation and reactivation of p16. H3-K9 di-methylation as well as DNA methylation related to p16 silencing is limited to the promoter region. In addition to its effect

  14. Histone H3 lysine 9 acetylation pattern suggests that X and B chromosomes are silenced during entire male meiosis in a grasshopper.

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    Cabrero, J; Teruel, M; Carmona, F D; Jiménez, R; Camacho, J P M

    2007-01-01

    The facultative heterochromatic X chromosome in leptotene spermatocytes of the grasshopper Eyprepocnemis plorans showed marked hypoacetylation for lysine 9 in the H3 histone (H3-K9) with no sign of histone H2AX phosphorylation. Since H3-K9 hypoacetylation precedes the meiotic appearance of phosphorylated H2AX (gamma-H2AX), which marks the beginning of recombinational DNA double-strand breaks (DSBs), it seems that meiotic sex-chromosome inactivation (MSCI) in this grasshopper occurs prior to the beginning of recombination and hence synapsis (which in this species begins later than recombination). In addition, all constitutively heterochromatic chromosome regions harbouring a 180-bp tandem-repeat DNA and rDNA (B chromosomes and pericentromeric regions of A chromosomes) were H3-K9 hypoacetylated at early leptotene even though they will synapse at subsequent stages. This also suggests that meiotic silencing in this grasshopper might be independent of synapsis. The H3-K9 hypoacetylated state of facultative and constitutive heterochromatin persisted during subsequent meiotic stages and was even apparent in round spermatids. Finally, the fact that B chromosomes are differentially hypoacetylated in testis and embryo interphase cells suggests that they might be silenced early in development and remain this way for most (or all) life-cycle stages. Copyright (c) 2007 S. Karger AG, Basel.

  15. SIRT1-dependent modulation of methylation and acetylation of histone H3 on lysine 9 (H3K9 in the zygotic pronuclei improves porcine embryo development

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    Katerina Adamkova

    2017-11-01

    Full Text Available Abstract Background The histone code is an established epigenetic regulator of early embryonic development in mammals. The lysine residue K9 of histone H3 (H3K9 is a prime target of SIRT1, a member of NAD+-dependent histone deacetylase family of enzymes targeting both histone and non-histone substrates. At present, little is known about SIRT1-modulation of H3K9 in zygotic pronuclei and its association with the success of preimplantation embryo development. Therefore, we evaluated the effect of SIRT1 activity on H3K9 methylation and acetylation in porcine zygotes and the significance of H3K9 modifications for early embryonic development. Results Our results show that SIRT1 activators resveratrol and BML-278 increased H3K9 methylation and suppressed H3K9 acetylation in both the paternal and maternal pronucleus. Inversely, SIRT1 inhibitors nicotinamide and sirtinol suppressed methylation and increased acetylation of pronuclear H3K9. Evaluation of early embryonic development confirmed positive effect of selective SIRT1 activation on blastocyst formation rate (5.2 ± 2.9% versus 32.9 ± 8.1% in vehicle control and BML-278 group, respectively; P ≤ 0.05. Stimulation of SIRT1 activity coincided with fluorometric signal intensity of ooplasmic ubiquitin ligase MDM2, a known substrate of SIRT1 and known limiting factor of epigenome remodeling. Conclusions We conclude that SIRT1 modulates zygotic histone code, obviously through direct deacetylation and via non-histone targets resulting in increased H3K9me3. These changes in zygotes lead to more successful pre-implantation embryonic development and, indeed, the specific SIRT1 activation due to BML-278 is beneficial for in vitro embryo production and blastocyst achievement.

  16. Profiling of histone H3 lysine 9 trimethylation levels predicts transcription factor activity and survival in acute myeloid leukemia

    DEFF Research Database (Denmark)

    Müller-Tidow, Carsten; Klein, Hans-Ulrich; Hascher, Antje

    2010-01-01

    ) patterns in primary AML (n=109), ALL (n=28), CD34+ cells (n=21) and white blood cells (n=15) specimens. Hundreds of promoter regions in AML showed significant alterations in H3K9me3 levels. H3K9me3 deregulation in AML occurred preferentially as a decrease in H3K9me3 levels at core promoter regions...

  17. Regulation of DNA methyltransferase 1 transcription in BRCA1-mutated breast cancer: a novel crosstalk between E2F1 motif hypermethylation and loss of histone H3 lysine 9 acetylation.

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    Li, Da; Bi, Fang-Fang; Cao, Ji-Min; Cao, Chen; Liu, Bo; Yang, Qing

    2014-02-06

    DNA methyltransferase 1 (DNMT1) plays a critical role in breast cancer progression. However, the epigenetic mechanism regulating DNMT1 expression remains largely unknown. Epigenetic regulation of DNMT1 was assessed in 85 invasive ductal carcinomas from BRCA1 mutation carriers. Association between clinicopathological features and DNMT1 promoter methylation was determined using Fisher's exact test. Univariate analysis of survival was performed using the Kaplan-Meier method. Multivariate Cox regression analysis was performed to identify the independent prognostic factors for overall survival. Hypermethylated E2F transcription factor 1 (E2F1) motif is a key regulatory element for the DNMT1 gene in BRCA1-mutated breast cancer. Mechanistically, the abnormal E2F1 motif methylation-mediated loss of active histone H3 lysine 9 acetylation (H3K9ac) and transcription factor E2F1 enrichment synergistically inhibited the transcription of DNMT1. Clinicopathological data indicated that the hypermethylated E2F1 motif was associated with histological grade, lymph node, Ki67 and E-cadherin status; univariate survival and multivariate analyses demonstrated that lymph node metastasis was an independent and reliable prognostic factor for BRCA1-mutated breast cancer patients. Our findings imply that genetic (such as BRCA1 mutation) and epigenetic mechanisms (such as DNA methylation, histone modification, transcription factor binding) are jointly involved in the malignant progression of DNMT1-related breast cancer.

  18. Pattern of change in histone 3 lysine 9 acetylation and histone ...

    Indian Academy of Sciences (India)

    Pattern of change in histone 3 lysine 9 acetylation and histone deacetylases in development of zebrafish embryo. Yanning Li Junxia Wang Ying Xie Shufeng Liu Ye ... Wang1 Ying Xie1 Shufeng Liu1 Ye Tian1. Hebei Key Lab of Laboratory Animal, Hebei Medical University, Shijiazhuang 050017, People's Republic of China ...

  19. DNA topoisomerase 1α promotes transcriptional silencing of transposable elements through DNA methylation and histone lysine 9 dimethylation in Arabidopsis.

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    Thanh Theresa Dinh

    2014-07-01

    Full Text Available RNA-directed DNA methylation (RdDM and histone H3 lysine 9 dimethylation (H3K9me2 are related transcriptional silencing mechanisms that target transposable elements (TEs and repeats to maintain genome stability in plants. RdDM is mediated by small and long noncoding RNAs produced by the plant-specific RNA polymerases Pol IV and Pol V, respectively. Through a chemical genetics screen with a luciferase-based DNA methylation reporter, LUCL, we found that camptothecin, a compound with anti-cancer properties that targets DNA topoisomerase 1α (TOP1α was able to de-repress LUCL by reducing its DNA methylation and H3K9me2 levels. Further studies with Arabidopsis top1α mutants showed that TOP1α silences endogenous RdDM loci by facilitating the production of Pol V-dependent long non-coding RNAs, AGONAUTE4 recruitment and H3K9me2 deposition at TEs and repeats. This study assigned a new role in epigenetic silencing to an enzyme that affects DNA topology.

  20. Pattern of change in histone 3 lysine 9 acetylation and histone ...

    Indian Academy of Sciences (India)

    2014-08-11

    Aug 11, 2014 ... reported to be enriched in embryonic stem cells and may be a member of pluripotency (Hezroni et al. 2011). Embryonic stem cell differentiation and oocyte maturation are accom- panied by reduced H3K9ac (Krejcí et al. ..... implications for disease and therapy. Nat. Rev. Genet. 10, 32–42. Harrison M. R. ...

  1. Global histone modification of histone H3 in colorectal cancer and its precursor lesions.

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    Nakazawa, Tadao; Kondo, Tetsuo; Ma, Defu; Niu, Dongfeng; Mochizuki, Kunio; Kawasaki, Tomonori; Yamane, Tetsu; Iino, Hiroshi; Fujii, Hideki; Katoh, Ryohei

    2012-06-01

    Chromatin remodeling through histone modification is an important mechanism of epigenetic gene dysregulation in human cancers. However, little is known about global alteration of histone status during tumorigenesis and cancer progression. Histone H3 status was examined in benign and malignant colorectal tumors by immunohistochemistry and Western blotting. For immunohistochemical evaluation, 4 anti-histone H3 antibodies, specific to dimethylation at lysine 4 (H3K4me2), acetylation at lysine 9 (H3K9ac), dimethylation at lysine 9 (H3K9me2), and trimethylation at lysine 27 (H3K27me3), were used. On immunohistochemistry, H3K4me2, H3K9ac, and H3K27me3 showed no significant changes between normal and colorectal tumors. On the other hand, the global level of H3K9me2 was distinctly higher in neoplastic cells (adenoma and adenocarcinoma) than in normal glandular cells. In addition, it was significantly higher in adenocarcinoma than in adenoma. Correspondingly, Western blotting confirmed that H3K9me2 expression was significantly higher in adenocarcinomas than in normal colorectal mucosa. No alteration of H3K9me2 was observed with tumor differentiation and with the histological subtypes of colorectal cancers. These results suggest that aberration of the global H3K9me2 level is an important epigenetic event in colorectal tumorigenesis and carcinogenesis involved with gene regulation in neoplastic cells through chromatin remodeling. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. H3K9me-independent gene silencing in fission yeast heterochromatin by Clr5 and histone deacetylases

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    Hansen, Klavs R; Hazan, Idit; Shanker, Sreenath

    2011-01-01

    Nucleosomes in heterochromatic regions bear histone modifications that distinguish them from euchromatic nucleosomes. Among those, histone H3 lysine 9 methylation (H3K9me) and hypoacetylation have been evolutionarily conserved and are found in both multicellular eukaryotes and single-cell model o...

  3. Alcohol consumption during gestation causes histone3 lysine9 hyperacetylation and an alternation of expression of heart development-related genes in mice.

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    Pan, Bo; Zhu, Jing; Lv, Tiewei; Sun, Huichao; Huang, Xupei; Tian, Jie

    2014-09-01

    Alcohol abuse during gestation may cause congenital heart diseases (CHDs). The underlying mechanisms of alcohol-induced cardiac deformities are still not clear. Recent studies suggest that histone modification may play a crucial role in this pathological process. Moreover, our previous studies reported that ethanol could induce histone3 lysine9 (H3K9) hyperacetylation and overexpression of heart development-related genes in vitro. The aim of this study was to investigate the effect of alcohol consumption during gestation on the imbalance of H3K9 acetylation and the alternation of the expression of heart development-related genes during cardiogenesis. Pregnant mice were exposed to a single dose of alcohol (10 μl/g/d, 56% alcohol) by gavage every day in the morning from embryo day 7.5 (E7.5) to E15.5. Hematoxylin and eosin (H&E) staining was applied for observing the structure of the embryonic hearts. Western blotting and quantitative real-time polymerase chain reaction were used for detecting the level of H3K9 acetylation and gene expression. Histone acetyltransferase (HAT) and histone deacetylase (HDAC) activities were detected by colorimetric assay and fluorometric assay. H&E staining of cardiac tissue showed abnormalities of embryonic hearts at E17.5. The level of H3K9 acetylation reached peak at E17.5 and decreased sharply to a low level at birth and maintained at low level afterward. Alcohol exposure increased H3K9 acetylation at E11.5, E14.5, E17.5, and E18.5, respectively (p hearts at E14.5 and E17.5, Mef2c at E14.5, and Nkx2.5 at E14.5 and E17.5, (p 0.05). On embryonic day 17.5, HAT activities of embryonic hearts increased significantly, however alcohol exposure did not alter HDAC activities. These data indicate a time course of H3K9 acetylation change during heart development and demonstrate that alcohol exposure in utero may induce an increase of HAT activities, which results in H3K9 hyperacetylation and an increase of the expression of heart development

  4. 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.

  5. Effect of phenylhexyl isothiocyanate on aberrant histone H3 methylation in primary human acute leukemia

    Directory of Open Access Journals (Sweden)

    Zou Yong

    2012-07-01

    Full Text Available Abstract Background We have previously studied the histone acetylation in primary human leukemia cells. However, histone H3 methylation in these cells has not been characterized. Methods This study examined the methylation status at histone H3 lysine 4 (H3K4 and histone H3 lysine 9 (H3K9 in primary acute leukemia cells obtained from patients and compared with those in the non-leukemia and healthy cells. We further characterized the effect of phenylhexyl isothiocyanate (PHI, Trichostatin A (TSA, and 5-aza-2’-deoxycytidine (5-Aza on the cells. Results We found that methylation of histone H3K4 was virtually undetectable, while methylation at H3K9 was significantly higher in primary human leukemia cells. The histone H3K9 hypermethylation and histone H3K4 hypomethylation were observed in both myeloid and lymphoid leukemia cells. PHI was found to be able to normalize the methylation level in the primary leukemia cells. We further showed that PHI was able to enhance the methyltransferase activity of H3K4 and decrease the activity of H3K9 methyltransferase. 5-Aza had similar effect on H3K4, but minimal effect on H3K9, whereas TSA had no effect on H3K4 and H3K9 methyltransferases. Conclusions This study revealed opposite methylation level of H3K4 and H3K9 in primary human leukemia cells and demonstrated for the first time that PHI has different effects on the methyltransferases for H3K4 and H3K9.

  6. High Affinity Binding of Chp1 Chromodomain to K9 Methylated Histone H3 is Required to Establish Centromeric Hterochromatin

    Energy Technology Data Exchange (ETDEWEB)

    Schalch, T.; Job, G; Noffsinger, V; Shanker, S; Kuscu, C; Joshua-Tor, L; Partridge, J

    2009-01-01

    In fission yeast, assembly of centromeric heterochromatin requires the RITS complex, which consists of Ago1, Tas3, Chp1, and siRNAs derived from centromeric repeats. Recruitment of RITS to centromeres has been proposed to depend on siRNA-dependent targeting of Ago1 to centromeric sequences. Previously, we demonstrated that methylated lysine 9 of histone H3 (H3K9me) acts upstream of siRNAs during heterochromatin establishment. Our crystal structure of Chp1's chromodomain in complex with a trimethylated lysine 9 H3 peptide reveals extensive sites of contact that contribute to Chp1's high-affinity binding. We found that this high-affinity binding is critical for the efficient establishment of centromeric heterochromatin, but preassembled heterochromatin can be maintained when Chp1's affinity for H3K9me is greatly reduced.

  7. Histone H3 tail clipping regulates gene expression

    OpenAIRE

    Santos-Rosa, Helena; Kirmizis, Antonis; Nelson, Christopher; Bartke, Till; Saksouk, Nehme; Cote, Jacques; Kouzarides, Tony

    2008-01-01

    Induction of gene expression in yeast and human cells involves changes in histone modifications associated with promoters. Here we identify a histone H3 endopeptidase activity in S. cerevisiae that may regulate these events. The endopeptidase cleaves H3 after alanine 21, generating a histone lacking the first 21 residues and displays a preference for H3 tails carrying repressive modifications. In vivo, the H3 N-terminus is clipped, specifically within the promoter of genes following the induc...

  8. 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.

  9. Dynamic acetylation of all lysine 4-methylated histone H3 in the mouse nucleus: analysis at c-fos and c-jun.

    Directory of Open Access Journals (Sweden)

    Catherine A Hazzalin

    2005-12-01

    Full Text Available A major focus of current research into gene induction relates to chromatin and nucleosomal regulation, especially the significance of multiple histone modifications such as phosphorylation, acetylation, and methylation during this process. We have discovered a novel physiological characteristic of all lysine 4 (K4-methylated histone H3 in the mouse nucleus, distinguishing it from lysine 9-methylated H3. K4-methylated histone H3 is subject to continuous dynamic turnover of acetylation, whereas lysine 9-methylated H3 is not. We have previously reported dynamic histone H3 phosphorylation and acetylation as a key characteristic of the inducible proto-oncogenes c-fos and c-jun. We show here that dynamically acetylated histone H3 at these genes is also K4-methylated. Although all three modifications are proven to co-exist on the same nucleosome at these genes, phosphorylation and acetylation appear transiently during gene induction, whereas K4 methylation remains detectable throughout this process. Finally, we address the functional significance of the turnover of histone acetylation on the process of gene induction. We find that inhibition of turnover, despite causing enhanced histone acetylation at these genes, produces immediate inhibition of gene induction. These data show that all K4-methylated histone H3 is subject to the continuous action of HATs and HDACs, and indicates that at c-fos and c-jun, contrary to the predominant model, turnover and not stably enhanced acetylation is relevant for efficient gene induction.

  10. Rôle d'histones methyltransférases spécifiques de H3K9 dans l'équilibre prolifération et différenciation cellulaire

    OpenAIRE

    Battisti, Valentine

    2013-01-01

    In eukaryotes, gene expression partly relies on chromatin compaction degree. Chromatin status is controlled by epigenetic marks, such as histones (chromatin structural proteins) posttranslational modifications. As an example, histone H3 lysine 9 (H3K9) methylation on gene promoters is mainly associated with transcriptional repression. H3K9 is methylated by several enzymes called lysine methyltransferases (KMTs). The aim of my thesis project was to understand the role of the H3K9 KMTs, G9a, GL...

  11. DNA sequence-dependent epigenetic inheritance of gene silencing and histone H3K9 methylation.

    Science.gov (United States)

    Wang, Xiaoyi; Moazed, Danesh

    2017-04-07

    Epigenetic inheritance mechanisms play fundamental roles in maintaining cellular memory of gene expression states. In fission yeast, histone H3 lysine 9 (H3K9) is methylated (H3K9me) at heterochromatic domains. These domains can be epigenetically inherited when epe1+ , encoding an enzyme that promotes H3K9 demethylation, is deleted. How native epigenetic states are stably maintained in epe1+ cells remains unknown. Here, we developed a system to examine the role of DNA sequence and genomic context in propagation of a cis-heritable H3K9me-dependent silenced state. We show that in epe1+ cells, in addition to sequence-independent mechanisms that propagate H3K9me, epigenetic inheritance of silencing requires binding sites for sequence-dependent activating transcription factor (ATF)-adenosine 3',5'-monophosphate (cAMP) response element-binding protein (CREB) family transcription factors within their native chromosomal context. Thus, specific DNA sequences contribute to cis inheritance of H3K9me and silent epigenetic states. Copyright © 2017, American Association for the Advancement of Science.

  12. Distribution pattern of histone H3 phosphorylation at serine 10 ...

    Indian Academy of Sciences (India)

    2013-08-06

    Aug 6, 2013 ... in chromosome distribution of H3S10ph when mitosis and meiosis were compared. ... [Paula C. M. P., Techio V. H., Sobrinho F. S. and Freitas A. S. 2013 Distribution pattern of histone H3 phosphorylation at serine 10 during mitosis and meiosis in ... RDWebster], since current knowledge about specific roles ...

  13. 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

  14. The SUVR4 histone lysine methyltransferase binds ubiquitin and converts H3K9me1 to H3K9me3 on transposon chromatin in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Silje V Veiseth

    2011-03-01

    Full Text Available Chromatin structure and gene expression are regulated by posttranslational modifications (PTMs on the N-terminal tails of histones. Mono-, di-, or trimethylation of lysine residues by histone lysine methyltransferases (HKMTases can have activating or repressive functions depending on the position and context of the modified lysine. In Arabidopsis, trimethylation of lysine 9 on histone H3 (H3K9me3 is mainly associated with euchromatin and transcribed genes, although low levels of this mark are also detected at transposons and repeat sequences. Besides the evolutionarily conserved SET domain which is responsible for enzyme activity, most HKMTases also contain additional domains which enable them to respond to other PTMs or cellular signals. Here we show that the N-terminal WIYLD domain of the Arabidopsis SUVR4 HKMTase binds ubiquitin and that the SUVR4 product specificity shifts from di- to trimethylation in the presence of free ubiquitin, enabling conversion of H3K9me1 to H3K9me3 in vitro. Chromatin immunoprecipitation and immunocytological analysis showed that SUVR4 in vivo specifically converts H3K9me1 to H3K9me3 at transposons and pseudogenes and has a locus-specific repressive effect on the expression of such elements. Bisulfite sequencing indicates that this repression involves both DNA methylation-dependent and -independent mechanisms. Transcribed genes with high endogenous levels of H3K4me3, H3K9me3, and H2Bub1, but low H3K9me1, are generally unaffected by SUVR4 activity. Our results imply that SUVR4 is involved in the epigenetic defense mechanism by trimethylating H3K9 to suppress potentially harmful transposon activity.

  15. Histone Deacetylase Inhibitors Globally Enhance H3/H4 Tail Acetylation Without Affecting H3 Lysine 56 Acetylation

    OpenAIRE

    Drogaris, Paul; Villeneuve, Val?rie; Pomi?s, Christelle; Lee, Eun-Hye; Bourdeau, V?ronique; Bonneil, ?ric; Ferbeyre, Gerardo; Verreault, Alain; Thibault, Pierre

    2012-01-01

    Histone deacetylase inhibitors (HDACi) represent a promising avenue for cancer therapy. We applied mass spectrometry (MS) to determine the impact of clinically relevant HDACi on global levels of histone acetylation. Intact histone profiling revealed that the HDACi SAHA and MS-275 globally increased histone H3 and H4 acetylation in both normal diploid fibroblasts and transformed human cells. Histone H3 lysine 56 acetylation (H3K56ac) recently elicited much interest and controversy due to its p...

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

    Science.gov (United States)

    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.

  17. Histone H3 mutations--a special role for H3.3 in tumorigenesis?

    Science.gov (United States)

    Kallappagoudar, Satish; Yadav, Rajesh K; Lowe, Brandon R; Partridge, Janet F

    2015-06-01

    Brain tumors are the most common solid tumors in children. Pediatric high-grade glioma (HGG) accounts for ∼8-12 % of these brain tumors and is a devastating disease as 70-90 % of patients die within 2 years of diagnosis. The failure to advance therapy for these children over the last 30 years is largely due to limited knowledge of the molecular basis for these tumors and a lack of disease models. Recently, sequencing of tumor cells revealed that histone H3 is frequently mutated in pediatric HGG, with up to 78 % of diffuse intrinsic pontine gliomas (DIPGs) carrying K27M and 36 % of non-brainstem gliomas carrying either K27M or G34R/V mutations. Although mutations in many chromatin modifiers have been identified in cancer, this was the first demonstration that histone mutations may be drivers of disease. Subsequent studies have identified high-frequency mutation of histone H3 to K36M in chondroblastomas and to G34W/L in giant cell tumors of bone, which are diseases of adolescents and young adults. Interestingly, the G34 mutations, the K36M mutations, and the majority of K27M mutations occur in genes encoding the replacement histone H3.3. Here, we review the peculiar characteristics of histone H3.3 and use this information as a backdrop to highlight current thinking about how the identified mutations may contribute to disease development.

  18. Histone acetyl transferase 1 is essential for mammalian development, genome stability, and the processing of newly synthesized histones H3 and H4.

    Science.gov (United States)

    Nagarajan, Prabakaran; Ge, Zhongqi; Sirbu, Bianca; Doughty, Cheryl; Agudelo Garcia, Paula A; Schlederer, Michaela; Annunziato, Anthony T; Cortez, David; Kenner, Lukas; Parthun, Mark R

    2013-06-01

    Histone acetyltransferase 1 is an evolutionarily conserved type B histone acetyltransferase that is thought to be responsible for the diacetylation of newly synthesized histone H4 on lysines 5 and 12 during chromatin assembly. To understand the function of this enzyme in a complex organism, we have constructed a conditional mouse knockout model of Hat1. Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality. The lungs of embryos and pups genetically deficient in Hat1 were much less mature upon histological evaluation. The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress. Many of the Hat1(-/-) neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw. Hat1(-/-) mouse embryonic fibroblasts (MEFs) are defective in cell proliferation and are sensitive to DNA damaging agents. In addition, the Hat1(-/-) MEFs display a marked increase in genome instability. Analysis of histone dynamics at sites of replication-coupled chromatin assembly demonstrates that Hat1 is not only responsible for the acetylation of newly synthesized histone H4 but is also required to maintain the acetylation of histone H3 on lysines 9, 18, and 27 during replication-coupled chromatin assembly.

  19. Histone acetyl transferase 1 is essential for mammalian development, genome stability, and the processing of newly synthesized histones H3 and H4.

    Directory of Open Access Journals (Sweden)

    Prabakaran Nagarajan

    2013-06-01

    Full Text Available Histone acetyltransferase 1 is an evolutionarily conserved type B histone acetyltransferase that is thought to be responsible for the diacetylation of newly synthesized histone H4 on lysines 5 and 12 during chromatin assembly. To understand the function of this enzyme in a complex organism, we have constructed a conditional mouse knockout model of Hat1. Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality. The lungs of embryos and pups genetically deficient in Hat1 were much less mature upon histological evaluation. The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress. Many of the Hat1(-/- neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw. Hat1(-/- mouse embryonic fibroblasts (MEFs are defective in cell proliferation and are sensitive to DNA damaging agents. In addition, the Hat1(-/- MEFs display a marked increase in genome instability. Analysis of histone dynamics at sites of replication-coupled chromatin assembly demonstrates that Hat1 is not only responsible for the acetylation of newly synthesized histone H4 but is also required to maintain the acetylation of histone H3 on lysines 9, 18, and 27 during replication-coupled chromatin assembly.

  20. Histone Acetyl Transferase 1 Is Essential for Mammalian Development, Genome Stability, and the Processing of Newly Synthesized Histones H3 and H4

    Science.gov (United States)

    Nagarajan, Prabakaran; Ge, Zhongqi; Sirbu, Bianca; Doughty, Cheryl; Agudelo Garcia, Paula A.; Schlederer, Michaela; Annunziato, Anthony T.; Cortez, David; Kenner, Lukas; Parthun, Mark R.

    2013-01-01

    Histone acetyltransferase 1 is an evolutionarily conserved type B histone acetyltransferase that is thought to be responsible for the diacetylation of newly synthesized histone H4 on lysines 5 and 12 during chromatin assembly. To understand the function of this enzyme in a complex organism, we have constructed a conditional mouse knockout model of Hat1. Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality. The lungs of embryos and pups genetically deficient in Hat1 were much less mature upon histological evaluation. The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress. Many of the Hat1−/− neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw. Hat1−/− mouse embryonic fibroblasts (MEFs) are defective in cell proliferation and are sensitive to DNA damaging agents. In addition, the Hat1−/− MEFs display a marked increase in genome instability. Analysis of histone dynamics at sites of replication-coupled chromatin assembly demonstrates that Hat1 is not only responsible for the acetylation of newly synthesized histone H4 but is also required to maintain the acetylation of histone H3 on lysines 9, 18, and 27 during replication-coupled chromatin assembly. PMID:23754951

  1. Histone H3 lysine 4 methyltransferase KMT2D.

    Science.gov (United States)

    Froimchuk, Eugene; Jang, Younghoon; Ge, Kai

    2017-09-05

    Histone-lysine N-methyltransferase 2D (KMT2D), also known as MLL4 and MLL2 in humans and Mll4 in mice, belongs to a family of mammalian histone H3 lysine 4 (H3K4) methyltransferases. It is a large protein over 5500 amino acids in size and is partially functionally redundant with KMT2C. KMT2D is widely expressed in adult tissues and is essential for early embryonic development. The C-terminal SET domain is responsible for its H3K4 methyltransferase activity and is necessary for maintaining KMT2D protein stability in cells. KMT2D associates with WRAD (WDR5, RbBP5, ASH2L, and DPY30), NCOA6, PTIP, PA1, and H3K27 demethylase UTX in one protein complex. It acts as a scaffold protein within the complex and is responsible for maintaining the stability of UTX. KMT2D is a major mammalian H3K4 mono-methyltransferase and co-localizes with lineage determining transcription factors on transcriptional enhancers. It is required for the binding of histone H3K27 acetyltransferases CBP and p300 on enhancers, enhancer activation and cell-type specific gene expression during differentiation. KMT2D plays critical roles in regulating development, differentiation, metabolism, and tumor suppression. It is frequently mutated in developmental diseases, such as Kabuki syndrome and congenital heart disease, and various forms of cancer. Further understanding of the mechanism through which KMT2D regulates gene expression will reveal why KMT2D mutations are so harmful and may help generate novel therapeutic approaches. Published by Elsevier B.V.

  2. Epigenetically repressing human cytomegalovirus lytic infection and reactivation from latency in THP-1 model by targeting H3K9 and H3K27 histone demethylases.

    Directory of Open Access Journals (Sweden)

    Xin Gan

    Full Text Available Human Cytomegalovirus (hCMV infects a broad range of the population and establishes life-long latency in the infected individuals. Periodically the latently infected virus can reactivate and becomes a significant cause of morbidity and mortality in immunocompromised individuals. In latent infection, the viral genome is suppressed in a heterochromatic state and viral gene transcription is silenced. Upon reactivation, the repressive chromatin is remodeled to an active form, allowing viral lytic gene transcription, initiated by the expression of viral Immediate Early (IE genes. During this process, a number of histone modification enzymes, including histone demethylases (HDMs, play important roles in driving IE expression, but the mechanisms involved are not fully understood. To get a better understanding of these mechanisms, we focused on two HDMs, KDM4 and KDM6, which reverse the repressive histone H3-lysine 9 and lysine 27 methylation, respectively. Our studies show that in lytic infection, both demethylases are important in the activation of viral IE gene expression. Simultaneous disruption of both via genetic or chemical methods leads to severely impaired viral IE gene expression and viral replication. Additionally, in an experimental latency-reactivation model in THP-1 cells, the KDM6 family member JMJD3 is induced upon viral reactivation and its knockdown resulted in reduced IE gene transcription. These findings suggest pharmacological inhibition of these HDMs may potentially block hCMV lytic infection and reactivation, and control the viral infection associated diseases, which are of significant unmet medical needs.

  3. The cellular level of histone H3 lysine 4 dimethylation correlates with response to adjuvant gemcitabine in Japanese pancreatic cancer patients treated with surgery.

    Science.gov (United States)

    Watanabe, T; Morinaga, S; Akaike, M; Numata, M; Tamagawa, H; Yamamoto, N; Shiozawa, M; Ohkawa, S; Kameda, Y; Nakamura, Y; Miyagi, Y

    2012-11-01

    To search for biomarkers identifying pancreatic cancer patients likely to benefit from adjuvant gemcitabine chemotherapy, we investigated the status of several histone modifications in pancreatic tumors and their relationship to clinicopathological features and outcomes. Sixty one pancreatic cancer patients, primarily treated by surgical removal of tumors, were involved in the study. Thirty patients completed postoperative adjuvant gemcitabine, and in 31 it was discontinued. Tumor specimens were examined using immunohistochemistry for di- and tri-methylation of histone H3 lysine 4 (H3K4me2 and H3K4me3), dimethylation and acetylation of histone H3 lysine 9 (H3K9me2 and H3K9ac), and acetylation of histone H3 lysine 18 (H3K18ac). Positive tumor staining for each histone modification was used to classify patients into low- and high-staining groups, which were examined for relationships to clinicopathological features and clinical outcomes. High expression of H3K4me3 was related to the well and moderately differentiated tumor histological type (p = 0.012) and low expression of H3K4me2 was related to the presence of perineural invasion (p = 0.007). No cellular histone modifications were associated with overall or disease-free survival of patients as a whole. In the subgroup analyses, a low level of H3K4me2 was significantly associated with worse disease free survival in patients that completed adjuvant gemcitabine (p = 0.0239). Univariate and multivariate hazard models also indicated that a low level of H3K4me2 was a significant independent predictor of disease-free survival (p = 0.007). H3K4me2 was found to be a predictor of response to adjuvant gemcitabine in Asian patients with pancreatic cancer. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Histone H3.3 Mutations: A Variant Path to Cancer

    OpenAIRE

    Yuen, Benjamin T.K.; Knoepfler, Paul S.

    2013-01-01

    A host of cancer types exhibit aberrant histone modifications. Recently, distinct and recurrent mutations in a specific histone variant, histone H3.3, have been implicated in a high proportion of malignant pediatric brain cancers. The presence of mutant H3.3 histone disrupts epigenetic post-translational modifications near genes involved in cancer processes and in brain function. Here, we propose several possible mechanisms by which mutant H3.3 histones may act to promote tumorigenesis. Furth...

  5. The C Terminus of the Histone Chaperone Asf1 Cross-Links to Histone H3 in Yeast and Promotes Interaction with Histones H3 and H4

    OpenAIRE

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

    2013-01-01

    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, furthe...

  6. 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

  7. Poly(ADP-ribosylation) regulates chromatin organization through histone H3 modification and DNA methylation of the first cell cycle of mouse embryos

    Energy Technology Data Exchange (ETDEWEB)

    Osada, Tomoharu, E-mail: osada.tomoharu@mg.medience.co.jp [Advanced Medical Science Research Department, Mitsubishi Chemical Medience Corporation, 14-1 Sunayama, Kamisu-shi, Ibaragi 314-0255 (Japan); Department of Regenerative and Developmental Biology, Mitsubishi Kagaku Institute of Life Sciences (MITILS), 11 Minamiooya, Machida-shi, Tokyo 194-8511 (Japan); Rydén, Anna-Margareta [Division of Genome Stability Research, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Masutani, Mitsuko, E-mail: mmasutan@ncc.go.jp [Division of Genome Stability Research, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan)

    2013-04-26

    Highlights: •Histone modification of the mouse pronuclei is regulated by poly(ADP-ribosylation). •Hypermethylation of the mouse female pronuclei is maintained by poly(ADP-ribosylation). •Parp1 is physically interacted with Suz12, which may function in the pronuclei. •Poly(ADP-ribosylation) affects ultrastructure of chromatin of the mouse pronucleus. -- Abstract: We examined the roles of poly(ADP-ribosylation) in chromatin remodeling during the first cell cycle of mouse embryos. Drug-based inhibition of poly(ADP-ribosylation) by a PARP inhibitor, PJ-34, revealed up-regulation of dimethylation of histone H3 at lysine 4 in male pronuclei and down-regulation of dimethylation of histone H3 at lysine 9 (H3K9) and lysine 27 (H3K27). Association of poly(ADP-ribosylation) with histone modification was suggested to be supported by the interaction of Suz12, a histone methyltransferase in the polycomb complex, with Parp1. PARP activity was suggested to be required for a proper localization and maintenance of Suz12 on chromosomes. Notably, DNA methylation level of female pronuclei in one-cell embryos was robustly decreased by PJ-34. Electron microscopic analysis showed a frequent appearance of unusual electron-dense areas within the female pronuclei, implying the disorganized and hypercondensed chromatin ultrastructure. These results show that poly(ADP-ribosylation) is important for the integrity of non-equivalent epigenetic dynamics of pronuclei during the first cell cycle of mouse embryos.

  8. Histone H3.3 mutations: a variant path to cancer.

    Science.gov (United States)

    Yuen, Benjamin T K; Knoepfler, Paul S

    2013-11-11

    A host of cancer types exhibit aberrant histone modifications. Recently, distinct and recurrent mutations in a specific histone variant, histone H3.3, have been implicated in a high proportion of malignant pediatric brain cancers. The presence of mutant H3.3 histone disrupts epigenetic posttranslational modifications near genes involved in cancer processes and in brain function. Here, we review possible mechanisms by which mutant H3.3 histones may act to promote tumorigenesis. Furthermore, we discuss how perturbations in normal H3.3 chromatin-related and epigenetic functions may more broadly contribute to the formation of human cancers. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. 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. PMID:24130491

  10. Histone H3 lysine 56 acetylation and the response to DNA replication fork damage

    DEFF Research Database (Denmark)

    Wurtele, Hugo; Kaiser, Gitte Schalck; Bacal, Julien

    2012-01-01

    In Saccharomyces cerevisiae, histone H3 lysine 56 acetylation (H3K56ac) occurs in newly synthesized histones that are deposited throughout the genome during DNA replication. Defects in H3K56ac sensitize cells to genotoxic agents, suggesting that this modification plays an important role in the DNA...

  11. Histone H3 Acetyl K9 and Histone H3 Tri Methyl K4 as Prognostic Markers for Patients with Cervical Cancer.

    Science.gov (United States)

    Beyer, Susanne; Zhu, Junyan; Mayr, Doris; Kuhn, Christina; Schulze, Sandra; Hofmann, Simone; Dannecker, Christian; Jeschke, Udo; Kost, Bernd P

    2017-02-23

    Chromatin remodeling alters gene expression in carcinoma tissue. Although cervical cancer is the fourth most common cancer in women worldwide, a systematic study about the prognostic value of specific changes in the chromatin structure, such as histone acetylation or histone methylation, is missing. In this study, the expression of histone H3 acetyl K9, which is known to denote active regions at enhancers and promoters, and histone H3 tri methyl K4, which preferentially identifies active gene promoters, were examined as both show high metastatic potential. A panel of patients with cervical cancer was selected and the importance of the histone modifications concerning survival-time (overall survival and relapse-free survival) was analyzed in 250 cases. Histone H3 acetyl K9 staining was correlated with low grading, low FIGO (TNM classification and the International Federation of Gynecology and Obstetrics) status, negative N-status and low T-status in cervical cancer, showing a higher expression in adenocarcinoma than in squamous cell carcinoma. Cytoplasmic expression of histone H3 tri methyl K4 in a cervical cancer specimen was correlated with advanced T-status and poor prognosis. While cytoplasmic H3K4me3 expression seemed to be a marker of relapse-free survival, nuclear expression showed a correlation to poor prognosis in overall survival. Within this study, we analyzed the chemical modification of two histone proteins that are connected to active gene expression. Histone H3 acetyl K9 was found to be an independent marker of overall survival. Histone H3 tri methyl K4 was correlated with poor prognosis and it was found to be an independent marker of relapse-free survival. Therefore, we could show that chromatin remodeling plays an important role in cervical cancer biology.

  12. Histone H3 Acetyl K9 and Histone H3 Tri Methyl K4 as Prognostic Markers for Patients with Cervical Cancer

    Directory of Open Access Journals (Sweden)

    Susanne Beyer

    2017-02-01

    Full Text Available Chromatin remodeling alters gene expression in carcinoma tissue. Although cervical cancer is the fourth most common cancer in women worldwide, a systematic study about the prognostic value of specific changes in the chromatin structure, such as histone acetylation or histone methylation, is missing. In this study, the expression of histone H3 acetyl K9, which is known to denote active regions at enhancers and promoters, and histone H3 tri methyl K4, which preferentially identifies active gene promoters, were examined as both show high metastatic potential. A panel of patients with cervical cancer was selected and the importance of the histone modifications concerning survival-time (overall survival and relapse-free survival was analyzed in 250 cases. Histone H3 acetyl K9 staining was correlated with low grading, low FIGO (TNM classification and the International Federation of Gynecology and Obstetrics status, negative N-status and low T-status in cervical cancer, showing a higher expression in adenocarcinoma than in squamous cell carcinoma. Cytoplasmic expression of histone H3 tri methyl K4 in a cervical cancer specimen was correlated with advanced T-status and poor prognosis. While cytoplasmic H3K4me3 expression seemed to be a marker of relapse-free survival, nuclear expression showed a correlation to poor prognosis in overall survival. Within this study, we analyzed the chemical modification of two histone proteins that are connected to active gene expression. Histone H3 acetyl K9 was found to be an independent marker of overall survival. Histone H3 tri methyl K4 was correlated with poor prognosis and it was found to be an independent marker of relapse-free survival. Therefore, we could show that chromatin remodeling plays an important role in cervical cancer biology.

  13. Phosphorylation of Histone H3 at Serine 10 is Indispensable for Neoplastic Cell Transformation

    OpenAIRE

    Choi, Hong Seok; Choi, Bu Young; Cho, Yong-Yeon; Mizuno, Hideya; Kang, Bong Seok; Bode, Ann M.; Dong, Zigang

    2005-01-01

    Very little is known about the role of histone H3 phosphorylation in malignant transformation and cancer development. Here, we examine the function of H3 phosphorylation in cell transformation in vivo. Introduction of small interference RNA (siRNA)-H3 into JB6 cells resulted in decreased EGF-induced cell transformation. In contrast, wildtype histone H3-overexpressing cells markedly stimulated EGF-induced cell transformation, whereas the H3 mutant S10A cells suppressed transformation. When wil...

  14. Histone h3 glutathionylation in proliferating mammalian cells destabilizes nucleosomal structure.

    Science.gov (United States)

    García-Giménez, José Luis; Òlaso, Gloria; Hake, Sandra B; Bönisch, Clemens; Wiedemann, Sonja M; Markovic, Jelena; Dasí, Francisco; Gimeno, Amparo; Pérez-Quilis, Carme; Palacios, Oscar; Capdevila, Mercè; Viña, José; Pallardó, Federico V

    2013-10-20

    Here we report that chromatin, the complex and dynamic eukaryotic DNA packaging structure, is able to sense cellular redox changes. Histone H3, the only nucleosomal protein that possesses cysteine(s), can be modified by glutathione (GSH). Using Biotin labeled glutathione ethyl ester (BioGEE) treatment of nucleosomes in vitro, we show that GSH, the most abundant antioxidant in mammals, binds to histone H3. BioGEE treatment of NIH3T3 cells indicates that glutathionylation of H3 is maximal in fast proliferating cells, correlating well with enhanced levels of H3 glutathionylation in different tumor cell lines. Furthermore, glutathionylation of H3 in vivo decreases in livers from aged SAMP8 and C57BL/6J mice. We demonstrate biochemically and by mass spectrometry that histone variants H3.2/H3.3 are glutathionylated on their cysteine residue 110. Furthermore, circular dichroism, thermal denaturation of reconstituted nucleosomes, and molecular modeling indicate that glutathionylation of histone H3 produces structural changes affecting nucleosomal stability. We characterize the implications of histone H3 glutathionylation in cell physiology and the modulation of core histone proteins structure affected by this modification. Histone H3 senses cellular redox changes through glutathionylation of Cys, which increases during cell proliferation and decreases during aging. Glutathionylation of histone H3 affects nucleosome stability structure leading to a more open chromatin structure.

  15. Structural basis for recognition of H3K56-acetylated histone H3-H4 by the chaperone Rtt106

    Energy Technology Data Exchange (ETDEWEB)

    Su, Dan; Hu, Qi; Li, Qing; Thompson, James R; Cui, Gaofeng; Fazly, Ahmed; Davies, Brian A; Botuyan, Maria Victoria; Zhang, Zhiguo; Mer, Georges [Mayo

    2013-04-08

    Dynamic variations in the structure of chromatin influence virtually all DNA-related processes in eukaryotes and are controlled in part by post-translational modifications of histones. One such modification, the acetylation of lysine 56 (H3K56ac) in the amino-terminal α-helix (αN) of histone H3, has been implicated in the regulation of nucleosome assembly during DNA replication and repair, and nucleosome disassembly during gene transcription. In Saccharomyces cerevisiae, the histone chaperone Rtt106 contributes to the deposition of newly synthesized H3K56ac-carrying H3-H4 complex on replicating DNA, but it is unclear how Rtt106 binds H3-H4 and specifically recognizes H3K56ac as there is no apparent acetylated lysine reader domain in Rtt106. Here, we show that two domains of Rtt106 are involved in a combinatorial recognition of H3-H4. An N-terminal domain homodimerizes and interacts with H3-H4 independently of acetylation while a double pleckstrin-homology (PH) domain binds the K56-containing region of H3. Affinity is markedly enhanced upon acetylation of K56, an effect that is probably due to increased conformational entropy of the αN helix of H3. Our data support a mode of interaction where the N-terminal homodimeric domain of Rtt106 intercalates between the two H3-H4 components of the (H3-H4)2 tetramer while two double PH domains in the Rtt106 dimer interact with each of the two H3K56ac sites in (H3-H4)2. We show that the Rtt106-(H3-H4)2 interaction is important for gene silencing and the DNA damage response.

  16. Characterization of lysine 56 of histone H3 as an acetylation site in Saccharomyces cerevisiae.

    NARCIS (Netherlands)

    Ozdemir, A.; Spicuglia, S.; Lasonder, E.; Vermeulen, M.; Campsteijn, C.G.; Stunnenberg, H.G.; Logie, C.

    2005-01-01

    Post-translational histone modifications abound and regulate multiple nuclear processes. Most modifications are targeted to the amino-terminal domains of histones. Here we report the identification and characterization of acetylation of lysine 56 within the core domain of histone H3. In the crystal

  17. 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,

  18. 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…

  19. Apoptotic phosphorylation of histone H3 on Ser-10 by protein kinase Cδ.

    Science.gov (United States)

    Park, Choon-Ho; Kim, Kyong-Tai

    2012-01-01

    Phosphorylation of histone H3 on Ser-10 is regarded as an epigenetic mitotic marker and is tightly correlated with chromosome condensation during both mitosis and meiosis. However, it was also reported that histone H3 Ser-10 phosphorylation occurs when cells are exposed to various death stimuli, suggesting a potential role in the regulation of apoptosis. Here we report that histone H3 Ser-10 phosphorylation is mediated by the pro-apoptotic kinase protein kinase C (PKC) δ during apoptosis. We observed that PKCδ robustly phosphorylates histone H3 on Ser-10 both in vitro and in vivo. Ectopic expression of catalytically active PKCδ efficiently induces condensed chromatin structure in the nucleus. We also discovered that activation of PKCδ is required for histone H3 Ser-10 phosphorylation after treatment with DNA damaging agents during apoptosis. Collectively, these findings suggest that PKCδ is the kinase responsible for histone H3 Ser-10 phosphoryation during apoptosis and thus contributes to chromatin condensation together with other apoptosis-related histone modifications. As a result, histone H3 Ser-10 phosphorylation can be designated a new 'apoptotic histone code' mediated by PKCδ.

  20. Apoptotic phosphorylation of histone H3 on Ser-10 by protein kinase Cδ.

    Directory of Open Access Journals (Sweden)

    Choon-Ho Park

    Full Text Available Phosphorylation of histone H3 on Ser-10 is regarded as an epigenetic mitotic marker and is tightly correlated with chromosome condensation during both mitosis and meiosis. However, it was also reported that histone H3 Ser-10 phosphorylation occurs when cells are exposed to various death stimuli, suggesting a potential role in the regulation of apoptosis. Here we report that histone H3 Ser-10 phosphorylation is mediated by the pro-apoptotic kinase protein kinase C (PKC δ during apoptosis. We observed that PKCδ robustly phosphorylates histone H3 on Ser-10 both in vitro and in vivo. Ectopic expression of catalytically active PKCδ efficiently induces condensed chromatin structure in the nucleus. We also discovered that activation of PKCδ is required for histone H3 Ser-10 phosphorylation after treatment with DNA damaging agents during apoptosis. Collectively, these findings suggest that PKCδ is the kinase responsible for histone H3 Ser-10 phosphoryation during apoptosis and thus contributes to chromatin condensation together with other apoptosis-related histone modifications. As a result, histone H3 Ser-10 phosphorylation can be designated a new 'apoptotic histone code' mediated by PKCδ.

  1. Histone H3.3 and cancer: A potential reader connection

    OpenAIRE

    Lan, Fei; Shi, Yang

    2014-01-01

    The building block of chromatin is nucleosome, which consists of 146 base pairs of DNA wrapped around a histone octamer composed of two copies of histone H2A, H2B, H3, and H4. Significantly, the somatic missense mutations of the histone H3 variant, H3.3, are associated with childhood and young-adult tumors, such as pediatric high-grade astrocytomas, as well as chondroblastoma and giant-cell tumors of the bone. The mechanisms by which these histone mutations cause cancer are by and large uncle...

  2. Dynamic histone H3 epigenome marking during the intraerythrocytic cycle of Plasmodium falciparum

    DEFF Research Database (Denmark)

    Salcedo-Amaya, Adriana M; van Driel, Marc A; Alako, Blaise T

    2009-01-01

    Epigenome profiling has led to the paradigm that promoters of active genes are decorated with H3K4me3 and H3K9ac marks. To explore the epigenome of Plasmodium falciparum asexual stages, we performed MS analysis of histone modifications and found a general preponderance of H3/H4 acetylation and H3...

  3. Analysis of specific lysine histone H3 and H4 acetylation and methylation status in clones of cells with a gene silenced by nickel exposure.

    Science.gov (United States)

    Yan, Yan; Kluz, Thomas; Zhang, Ping; Chen, Hao-bin; Costa, Max

    2003-08-01

    We have previously reported that the gpt transgene in G12 Chinese hamster cells could be silenced by water-insoluble nickel compounds nickel sulfide (NiS) or nickel subsulfide (Ni(3)S(2)) and showed that the transgene was silenced by de novo DNA methylation and chromatin condensation. To further understand the nature of this silencing, we used the chromatin immunoprecipitation assay to elucidate the chromatin structure in nickel-induced silenced G12 clones. We also analyzed the effects of the DNA methyltransferase inhibitor 5-azacytidine (5-AzaC) and a histone deacetylase inhibitor trichostatin A (TSA) on histone H3 and H4 acetylation and gpt gene expression in selected nickel-silenced clones. We observed that both histone H3 and H4 were hypoacetylated and a methyl DNA-binding protein MeCP2 was targeted to the gpt gene locus, resulting in a localized inactive chromatin configuration in nickel-silenced cell clones. The histone H3K9 was also found methylated in three of four nickel- silenced cell clones, whereas the histone H3K9 was deacetylated in all four cell clones, indicating that the H3K9 methylation was involved in nickel-induced gene silencing. The acetylation of the gpt gene could be increased by a combination of 5-AzaC and TSA treatment, but not by either 5-AzaC or TSA alone. The gpt transcript was studied by either Northern blot or by semiquantitative RT-PCR following treatment of the silenced clones with TSA or 5-AzaC. An increase in gpt mRNA could be detected by RT-PCR in the clones that regained acetylation of H3 and H4. These data show that gene silencing induced by nickel in the gpt transgenic cell line involved a loss of histone acetylation and an activation of histone methylation. Both H4 and H3 histone acetylation were lost in the silenced clones and these clones exhibited an increase in the methylation of the lysine 9 in histone H3.

  4. A trans-tail histone code defined by monomethylated H4 Lys-20 and H3 Lys-9 demarcates distinct regions of silent chromatin.

    Science.gov (United States)

    Sims, Jennifer K; Houston, Sabrina I; Magazinnik, Tanya; Rice, Judd C

    2006-05-05

    The specific post-translational modifications of the histone proteins are associated with specific DNA-templated processes, such as transcriptional activation or repression. To investigate the biological role(s) of histone H4 lysine 20 (H4 Lys-20) methylation, we created a novel panel of antibodies that specifically detected mono-, di-, or trimethylated H4 Lys-20. We report that the different methylated forms of H4 Lys-20 are compartmentalized within visually distinct, transcriptionally silent regions in the mammalian nucleus. Interestingly, direct comparison of methylated H4 Lys-20 with the different methylated states of histone H3 lysine 9 (H3 Lys-9) revealed significant overlap and exclusion between the specific groups of methyl modifications. Trimethylated H4 Lys-20 and H3 Lys-9 were both selectively enriched within pericentric heterochromatin. Similarly, monomethylated H4 Lys-20 and H3 Lys-9 partitioned together and the dimethylated forms partitioned together within the chromosome arms; however, the mono- and dimethylated modifications were virtually exclusive. These findings strongly suggest that the combinatorial presence or absence of the different methylated states of H4 Lys-20 and H3 Lys-9 define particular types of silent chromatin. Consistent with this, detailed analysis of monomethylated H4 Lys-20 and H3 Lys-9 revealed that both were preferentially and selectively enriched within the same nucleosome particle in vivo. Collectively, these findings define a novel trans-tail histone code involving monomethylated H4 Lys-20 and H3 Lys-9 that act cooperatively to mark distinct regions of silent chromatin within the mammalian epigenome.

  5. Quantitative Histone Mass Spectrometry Identifies Elevated Histone H3 Lysine 27 (Lys27) Trimethylation in Melanoma.

    Science.gov (United States)

    Sengupta, Deepanwita; Byrum, Stephanie D; Avaritt, Nathan L; Davis, Lauren; Shields, Bradley; Mahmoud, Fade; Reynolds, Matthew; Orr, Lisa M; Mackintosh, Samuel G; Shalin, Sara C; Tackett, Alan J

    2016-03-01

    Normal cell growth is characterized by a regulated epigenetic program that drives cellular activities such as gene transcription, DNA replication, and DNA damage repair. Perturbation of this epigenetic program can lead to events such as mis-regulation of gene transcription and diseases such as cancer. To begin to understand the epigenetic program correlated to the development of melanoma, we performed a novel quantitative mass spectrometric analysis of histone post-translational modifications mis-regulated in melanoma cell culture as well as patient tumors. Aggressive melanoma cell lines as well as metastatic melanoma were found to have elevated histone H3 Lys(27) trimethylation (H3K27me3) accompanied by overexpressed methyltransferase EZH2 that adds the specific modification. The altered epigenetic program that led to elevated H3K27me3 in melanoma cell culture was found to directly silence transcription of the tumor suppressor genes RUNX3 and E-cadherin. The EZH2-mediated silencing of RUNX3 and E-cadherin transcription was also validated in advanced stage human melanoma tissues. This is the first study focusing on the detailed epigenetic mechanisms leading to EZH2-mediated silencing of RUNX3 and E-cadherin tumor suppressors in melanoma. This study underscores the utility of using high resolution mass spectrometry to identify mis-regulated epigenetic programs in diseases such as cancer, which could ultimately lead to the identification of biological markers for diagnostic and prognostic applications. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. 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

  7. Sgf29 binds histone H3K4me2/3 and is required for SAGA complex recruitment and histone H3 acetylation

    Energy Technology Data Exchange (ETDEWEB)

    Bian, Chuanbing; Xu, Chao; Ruan, Jianbin; Lee, Kenneth K.; Burke, Tara L.; Tempel, Wolfram; Barsyte, Dalia; Li, Jing; Wu, Minhao; Zhou, Bo O.; Fleharty, Brian E.; Paulson, Ariel; Allali-Hassani, Abdellah; Zhou, Jin-Qiu; Mer, Georges; Grant, Patrick A.; Workman, Jerry L.; Zang, Jianye; Min, Jinrong (Toronto); (Stowers); (UST - China); (UV); (Chinese Aca. Sci.); (MCCM)

    2011-09-28

    The SAGA (Spt-Ada-Gcn5 acetyltransferase) complex is an important chromatin modifying complex that can both acetylate and deubiquitinate histones. Sgf29 is a novel component of the SAGA complex. Here, we report the crystal structures of the tandem Tudor domains of Saccharomyces cerevisiae and human Sgf29 and their complexes with H3K4me2 and H3K4me3 peptides, respectively, and show that Sgf29 selectively binds H3K4me2/3 marks. Our crystal structures reveal that Sgf29 harbours unique tandem Tudor domains in its C-terminus. The tandem Tudor domains in Sgf29 tightly pack against each other face-to-face with each Tudor domain harbouring a negatively charged pocket accommodating the first residue alanine and methylated K4 residue of histone H3, respectively. The H3A1 and K4me3 binding pockets and the limited binding cleft length between these two binding pockets are the structural determinants in conferring the ability of Sgf29 to selectively recognize H3K4me2/3. Our in vitro and in vivo functional assays show that Sgf29 recognizes methylated H3K4 to recruit the SAGA complex to its targets sites and mediates histone H3 acetylation, underscoring the importance of Sgf29 in gene regulation.

  8. A histone H3K9M mutation traps histone methyltransferase Clr4 to prevent heterochromatin spreading

    Energy Technology Data Exchange (ETDEWEB)

    Shan, Chun-Min; Wang, Jiyong; Xu, Ke; Chen, Huijie; Yue, Jia-Xing; Andrews, Stuart; Moresco, James J.; Yates, John R.; Nagy, Peter L.; Tong, Liang; Jia, Songtao

    2016-09-20

    Histone lysine-to-methionine (K-to-M) mutations are associated with multiple cancers, and they function in a dominant fashion to block the methylation of corresponding lysines on wild type histones. However, their mechanisms of function are controversial. Here we show that in fission yeast, introducing the K9M mutation into one of the three histone H3 genes dominantly blocks H3K9 methylation on wild type H3 across the genome. In addition, H3K9M enhances the interaction of histone H3 tail with the H3K9 methyltransferase Clr4 in a SAM (S-adenosyl-methionine)-dependent manner, and Clr4 is trapped at nucleation sites to prevent its spreading and the formation of large heterochromatin domains. We further determined the crystal structure of an H3K9M peptide in complex with human H3K9 methyltransferase G9a and SAM, which reveales that the methionine side chain had enhanced van der Waals interactions with G9a. Therefore, our results provide a detailed mechanism by which H3K9M regulates H3K9 methylation.

  9. Hypoacetylation of acetyl-histone H3 (H3K9ac) as marker of poor prognosis in oral cancer.

    Science.gov (United States)

    Webber, Liana P; Wagner, Vivian P; Curra, Marina; Vargas, Pablo A; Meurer, Luise; Carrard, Vinícius C; Squarize, Cristiane H; Castilho, Rogério M; Martins, Manoela D

    2017-08-01

    Epigenetics refers to changes in cell characteristics that occur independently of modifications to the DNA sequence. Oral carcinogenesis is influenced by modifications in epigenetic mechanisms, including changes in histones, which are proteins that support chromatin remodelling for the dynamic regulation of gene expression and silencing. The dysregulation of histone acetylation can lead to the uncontrolled activity of different genes, thereby triggering events associated with malignant transformation. The aim of this study was to analyse the expression of acetyl-histone H3 at lys9 (H3K9ac) in oral leucoplakia (OL) and oral squamous cell carcinoma (OSCC) in addition to its association with cell proliferation, epithelial-mesenchymal transition (EMT) and clinical-pathological findings. Samples of normal oral mucosa (NOM), OL and OSCC were submitted to immunohistochemical analysis using anti-H3K9ac, Ki67 and vimentin. Slides were submitted to quantitative analysis regarding the percentage of positive cells. OSCC presented less expression of H3K9ac in comparison to OL (P oral carcinogenesis along with an increase in cell proliferation and EMT. The results demonstrate that H3K9ac may be a useful novel prognostic marker for OSCC. © 2017 John Wiley & Sons Ltd.

  10. Histone H3K4 and H3K36 Methylation Independently Recruit the NuA3 Histone Acetyltransferase in Saccharomyces cerevisiae.

    Science.gov (United States)

    Martin, Benjamin J E; McBurney, Kristina L; Maltby, Vicki E; Jensen, Kristoffer N; Brind'Amour, Julie; Howe, LeAnn J

    2017-03-01

    Histone post-translational modifications (PTMs) alter chromatin structure by promoting the interaction of chromatin-modifying complexes with nucleosomes. The majority of chromatin-modifying complexes contain multiple domains that preferentially interact with modified histones, leading to speculation that these domains function in concert to target nucleosomes with distinct combinations of histone PTMs. In Saccharomyces cerevisiae, the NuA3 histone acetyltransferase complex contains three domains, the PHD finger in Yng1, the PWWP domain in Pdp3, and the YEATS domain in Taf14; which in vitro bind to H3K4 methylation, H3K36 methylation, and acetylated and crotonylated H3K9, respectively. While the in vitro binding has been well characterized, the relative in vivo contributions of these histone PTMs in targeting NuA3 is unknown. Here, through genome-wide colocalization and by mutational interrogation, we demonstrate that the PHD finger of Yng1, and the PWWP domain of Pdp3 independently target NuA3 to H3K4 and H3K36 methylated chromatin, respectively. In contrast, we find no evidence to support the YEATS domain of Taf14 functioning in NuA3 recruitment. Collectively our results suggest that the presence of multiple histone PTM binding domains within NuA3, rather than restricting it to nucleosomes containing distinct combinations of histone PTMs, can serve to increase the range of nucleosomes bound by the complex. Interestingly, however, the simple presence of NuA3 is insufficient to ensure acetylation of the associated nucleosomes, suggesting a secondary level of acetylation regulation that does not involve control of HAT-nucleosome interactions. Copyright © 2017 by the Genetics Society of America.

  11. Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Roche, Joëlle, E-mail: joelle.roche@univ-poitiers.fr [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States); CNRS FRE 3511, University of Poitiers, 1 rue Georges Bonnet, F-86022 Poitiers Cédex (France); Nasarre, Patrick; Gemmill, Robert [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States); Baldys, Aleksander [Department of Medicine, Nephrology Division, MUSC, Ralph H. Johnson Veterans Affairs Medical Center, Charleston, SC 29425 (United States); Pontis, Julien [Epigénétique & Destin Cellulaire, CNRS UMR 7216, University of Paris Diderot, Sorbonne Paris Cité, F-75013 Paris (France); Korch, Christopher [CU DNA Sequencing and Analysis Core, University of Colorado, School of Medicine, Anschutz Medical Campus, 12801 E. 17th Ave., Aurora, CO 80045 (United States); Guilhot, Joëlle [INSERM, CIC 0802, CHU de Poitiers, F-86021 (France); Ait-Si-Ali, Slimane [Epigénétique & Destin Cellulaire, CNRS UMR 7216, University of Paris Diderot, Sorbonne Paris Cité, F-75013 Paris (France); Drabkin, Harry [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States)

    2013-04-03

    The epithelial to mesenchymal transition (EMT) enables epithelial cells with a migratory mesenchymal phenotype. It is activated in cancer cells and is involved in invasion, metastasis and stem-like properties. ZEB1, an E-box binding transcription factor, is a major suppressor of epithelial genes in lung cancer. In the present study, we show that in H358 non-small cell lung cancer cells, ZEB1 downregulates EpCAM (coding for an epithelial cell adhesion molecule), ESRP1 (epithelial splicing regulatory protein), ST14 (a membrane associated serine protease involved in HGF processing) and RAB25 (a small G-protein) by direct binding to these genes. Following ZEB1 induction, acetylation of histone H4 and histone H3 on lysine 9 (H3K9) and 27 (H3K27) was decreased on ZEB1 binding sites on these genes as demonstrated by chromatin immunoprecipitation. Of note, decreased H3K27 acetylation could be also detected by western blot and immunocytochemistry in ZEB1 induced cells. In lung cancers, H3K27 acetylation level was higher in the tumor compartment than in the corresponding stroma where ZEB1 was more often expressed. Since HDAC and DNA methylation inhibitors increased expression of ZEB1 target genes, targeting these epigenetic modifications would be expected to reduce metastasis.

  12. Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Slimane Ait-Si-Ali

    2013-04-01

    Full Text Available The epithelial to mesenchymal transition (EMT enables epithelial cells with a migratory mesenchymal phenotype. It is activated in cancer cells and is involved in invasion, metastasis and stem-like properties. ZEB1, an E-box binding transcription factor, is a major suppressor of epithelial genes in lung cancer. In the present study, we show that in H358 non-small cell lung cancer cells, ZEB1 downregulates EpCAM (coding for an epithelial cell adhesion molecule, ESRP1 (epithelial splicing regulatory protein, ST14 (a membrane associated serine protease involved in HGF processing and RAB25 (a small G-protein by direct binding to these genes. Following ZEB1 induction, acetylation of histone H4 and histone H3 on lysine 9 (H3K9 and 27 (H3K27 was decreased on ZEB1 binding sites on these genes as demonstrated by chromatin immunoprecipitation. Of note, decreased H3K27 acetylation could be also detected by western blot and immunocytochemistry in ZEB1 induced cells. In lung cancers, H3K27 acetylation level was higher in the tumor compartment than in the corresponding stroma where ZEB1 was more often expressed. Since HDAC and DNA methylation inhibitors increased expression of ZEB1 target genes, targeting these epigenetic modifications would be expected to reduce metastasis.

  13. Association Between Phosphorylated Histone H3 and Oncotype DX Recurrence Scores in Breast Cancer.

    Science.gov (United States)

    Lee, Lik Hang; Swanson, Paul E; Tang, Patricia A; Bigras, Gilbert; Yang, Hua

    2017-01-01

    We investigate the association between phosphorylated histone H3 (PhH3) and Oncotype DX recurrence score (RS). All invasive breast carcinoma with RS results from our city between 2007 and 2010 (n=47) were reviewed. Whole-tumor sections were stained for PhH3. Mitotic and PhH3 counts were performed and clinical charts reviewed. PhH3 correlated well with RS (r=0.69, Pcancer.

  14. The Histone Variant H3.3 in Transcriptional Regulation and Human Disease.

    Science.gov (United States)

    Shi, Leilei; Wen, Hong; Shi, Xiaobing

    2017-06-30

    Histone proteins wrap around DNA to form nucleosomes, which further compact into the higher-order structure of chromatin. In addition to the canonical histones, there are also variant histones that often have pivotal roles in regulating chromatin dynamics and in the accessibility of the underlying DNA. H3.3 is the most common non-centromeric variant of histone H3 that differs from the canonical H3 by just 4-5 aa. Here, we discuss the current knowledge of H3.3 in transcriptional regulation and the recent discoveries and molecular mechanisms of H3.3 mutations in human cancer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Dynamic histone H3 epigenome marking during the intraerythrocytic cycle of Plasmodium falciparum.

    NARCIS (Netherlands)

    Salcedo-Amaya, A.M.; Driel, M.A. van; Alako, B.T.; Trelle, M. B.; Elzen, A.M.G. van den; Cohen, A.M.; Janssen-Megens, E.M.; Vegte-Bolmer, M.G. van de; Selzer, R.R.; Iniguez, A.L.; Green, R.D.; Sauerwein, R.W.; Jensen, O.N.; Stunnenberg, H.G.

    2009-01-01

    Epigenome profiling has led to the paradigm that promoters of active genes are decorated with H3K4me3 and H3K9ac marks. To explore the epigenome of Plasmodium falciparum asexual stages, we performed MS analysis of histone modifications and found a general preponderance of H3/H4 acetylation and

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

    Directory of Open Access Journals (Sweden)

    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.

  17. Expression of acetylated and dimethylated histone H3 in colorectal cancer.

    Science.gov (United States)

    Hashimoto, Toshio; Yamakawa, Mitsunori; Kimura, Shingo; Usuba, Osamu; Toyono, Mitsuru

    2013-01-01

    The expression of acetylated and dimethylated histone H3 in colorectal cancer was examined by immunohistochemistry and chromatin immunoprecipitation (ChIP)/Western blot (WB) assay. The correlation between the expression of histone H3 and clinicopathological findings was analyzed. Formalin-fixed and paraffin-embedded sections obtained from 80 operated cases of colorectal cancer were immunostained with anti-acetylated histone H3 (H3Ac) antibody and anti-dimethylated histone H3 lysine 4 (H3K4) antibody. Positive immunoreactivity was evaluated using the Allred scoring system. Furthermore, the expression was confirmed by ChIP/WB assay using formalin-fixed and paraffin-embedded sections. There was good correlation between immunostaining and expression on ChIP/WB assay (p = 0.0005). There was a significant difference between the Allred score of H3K4 and the depth of tumor invasion (p = 0.0003) and the pathological stages (p = 0.0065). In overall survival classified by Allred scores of H3Ac (p = 0.0072) and H3K4 (p = 0.0187), the highest scores represented significantly worse prognoses than the other scores. Specifically, in stages II and III, the highest scores represented significantly worse prognoses than the other scores (p < 0.0001 and p = 0.0173, respectively). The expression of H3Ac and H3K4 may estimate patient prognosis. © 2013 S. Karger AG, Basel.

  18. The histone H3.3K36M mutation reprograms the epigenome of chondroblastomas

    OpenAIRE

    Fang, Dong; Gan, Haiyun; Lee, Jeong-Heon; Han, Jing; Wang, Zhiquan; Riester, Scott M; Jin, Long; Chen, Jianji; Zhou, Hui; Wang, Jinglong; Zhang, Honglian; Yang, Na; Bradley, Elizabeth W.; Ho, Thai H.; Rubin, Brian P.

    2016-01-01

    Over 90% of chondroblastomas contain a heterozygous mutation replacing lysine 36 with methionine (K36M) in the histone H3 variant H3.3. Here, we show that H3K36 methylation is reduced globally in chondroblastomas and in chondrocytes harboring the same genetic mutation due to inhibition of at least two H3K36 methyltransferases, MMSET and SETD2, by the H3.3K36M mutant proteins. Genes with altered expression as well as H3K36 di- and trimethylation in H3.3K36M cells are enriched in cancer pathway...

  19. Somatic mutations of the histone H3K27 demethylase, UTX, in human cancer

    OpenAIRE

    van Haaften, Gijs; Dalgliesh, Gillian L; Davies, Helen; Chen, Lina; Bignell, Graham; Greenman, Chris; Edkins, Sarah; Hardy, Claire; O?Meara, Sarah; Teague, Jon; Butler, Adam; Hinton, Jonathan; Latimer, Calli; Andrews, Jenny; Barthorpe, Syd

    2009-01-01

    Somatically acquired epigenetic changes are present in many cancers. Epigenetic regulation is maintained via post-translational modifications of core histones. Here, we describe inactivating somatic mutations in the histone lysine demethylase, UTX, pointing to histone H3 lysine methylation deregulation in multiple tumour types. UTX reintroduction into cancer cells with inactivating UTX mutations resulted in slowing of proliferation and marked transcriptional changes. These data identify UTX a...

  20. Histone H3 lysine-to-methionine mutants as a paradigm to study chromatin signaling.

    Science.gov (United States)

    Herz, Hans-Martin; Morgan, Marc; Gao, Xin; Jackson, Jessica; Rickels, Ryan; Swanson, Selene K; Florens, Laurence; Washburn, Michael P; Eissenberg, Joel C; Shilatifard, Ali

    2014-08-29

    Histone H3 lysine(27)-to-methionine (H3K27M) gain-of-function mutations occur in highly aggressive pediatric gliomas. We established a Drosophila animal model for the pathogenic histone H3K27M mutation and show that its overexpression resembles polycomb repressive complex 2 (PRC2) loss-of-function phenotypes, causing derepression of PRC2 target genes and developmental perturbations. Similarly, an H3K9M mutant depletes H3K9 methylation levels and suppresses position-effect variegation in various Drosophila tissues. The histone H3K9 demethylase KDM3B/JHDM2 associates with H3K9M-containing nucleosomes, and its misregulation in Drosophila results in changes of H3K9 methylation levels and heterochromatic silencing defects. We have established histone lysine-to-methionine mutants as robust in vivo tools for inhibiting methylation pathways that also function as biochemical reagents for capturing site-specific histone-modifying enzymes, thus providing molecular insight into chromatin signaling pathways. Copyright © 2014, American Association for the Advancement of Science.

  1. Formaldehyde-induced histone H3 phosphorylation via JNK and the expression of proto-oncogenes

    Energy Technology Data Exchange (ETDEWEB)

    Yoshida, Ikuma; Ibuki, Yuko, E-mail: ibuki@u-shizuoka-ken.ac.jp

    2014-12-15

    Graphical abstract: - Highlights: • Formaldehyde modified histones. • The phosphorylation of H3S10 was increased at the promoter regions of proto-oncogenes. • The phosphorylation of H2AXS139 was attributed to FA-induced DNA damage. • The FA-induced initiation and promotion of cancer could be judged by these modifications. - Abstract: Formaldehyde (FA) is a very reactive compound that forms DNA adducts and DNA-protein crosslinks, which are known to contribute to FA-induced mutations and carcinogenesis. Post-translational modifications to histones have recently attracted attention due to their link with cancer. In the present study, we examined histone modifications following a treatment with FA. FA significantly phosphorylated histone H3 at serine 10 (H3S10), and at serine 28 (H3S28), the time-course of which was similar to the phosphorylation of H2AX at serine 139 (γ-H2AX), a marker of DNA double strand breaks. The temporal deacetylation of H3 was observed due to the reaction of FA with the lysine residues of histones. The phosphorylation mechanism was then analyzed by focusing on H3S10. The nuclear distribution of the phosphorylation of H3S10 and γ-H2AX did not overlap, and the phosphorylation of H3S10 could not be suppressed with an inhibitor of ATM/ATR, suggesting that the phosphorylation of H3S10 was independent of the DNA damage response. ERK and JNK in the MAPK pathways were phosphorylated by the treatment with FA, in which the JNK pathway was the main target for phosphorylation. The phosphorylation of H3S10 increased at the promoter regions of c-fos and c-jun, indicating a relationship between FA-induced tumor promotion activity and phosphorylation of H3S10. These results suggested that FA both initiates and promotes cancer, as judged by an analysis of histone modifications.

  2. Loss of H3K9me3 Correlates with ATM Activation and Histone H2AX Phosphorylation Deficiencies in Hutchinson-Gilford Progeria Syndrome.

    Directory of Open Access Journals (Sweden)

    Haoyue Zhang

    Full Text Available Compelling evidence suggests that defective DNA damage response (DDR plays a key role in the premature aging phenotypes in Hutchinson-Gilford progeria syndrome (HGPS. Studies document widespread alterations in histone modifications in HGPS cells, especially, the global loss of histone H3 trimethylated on lysine 9 (H3K9me3. In this study, we explore the potential connection(s between H3K9me3 loss and the impaired DDR in HGPS. When cells are exposed to a DNA-damaging agent Doxorubicin (Dox, double strand breaks (DSBs are generated that result in the phosphorylation of histone H2A variant H2AX (gammaH2AX within an hour. We find that the intensities of gammaH2AX foci appear significantly weaker in the G0/G1 phase HGPS cells compared to control cells. This reduction is associated with a delay in the recruitment of essential DDR factors. We further demonstrate that ataxia-telangiectasia mutated (ATM is responsible for the amplification of gammaH2AX signals at DSBs during G0/G1 phase, and its activation is inhibited in the HGPS cells that display significant loss of H3K9me3. Moreover, methylene (MB blue treatment, which is known to save heterochromatin loss in HGPS, restores H3K9me3, stimulates ATM activity, increases gammaH2AX signals and rescues deficient DDR. In summary, this study demonstrates an early DDR defect of attenuated gammaH2AX signals in G0/G1 phase HGPS cells and provides a plausible connection between H3K9me3 loss and DDR deficiency.

  3. Recognition Elements in the Histone H3 and H4 Tails for Seven Different Importins.

    Science.gov (United States)

    Soniat, Michael; Cağatay, Tolga; Chook, Yuh Min

    2016-09-30

    N-terminal tails of histones H3 and H4 are known to bind several different Importins to import the histones into the cell nucleus. However, it is not known what binding elements in the histone tails are recognized by the individual Importins. Biochemical studies of H3 and H4 tails binding to seven Importins, Impβ, Kapβ2, Imp4, Imp5, Imp7, Imp9, and Impα, show the H3 tail binding more tightly than the H4 tail. The H3 tail binds Kapβ2 and Imp5 with KD values of 77 and 57 nm, respectively, and binds the other five Importins more weakly. Mutagenic analysis shows H3 tail residues 11-27 to be the sole binding segment for Impβ, Kapβ2, and Imp4. However, Imp5, Imp7, Imp9, and Impα bind two separate elements in the H3 tail: the segment at residues 11-27 and an isoleucine-lysine nuclear localization signal (IK-NLS) motif at residues 35-40. The H4 tail also uses either one or two basic segments to bind the same set of Importins with a similar trend of relative affinities as the H3 tail, albeit at least 10-fold weaker. Of the many lysine residues in the H3 and H4 tails, only acetylation of the H3 Lys14 substantially decreased binding to several Importins. Lastly, we show that, in addition to the N-terminal tails, the histone fold domains of H3 and H4 and/or the histone chaperone Asf1b are important for Importin-histone recognition. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Recognition Elements in the Histone H3 and H4 Tails for Seven Different Importins*

    Science.gov (United States)

    Soniat, Michael; Cağatay, Tolga; Chook, Yuh Min

    2016-01-01

    N-terminal tails of histones H3 and H4 are known to bind several different Importins to import the histones into the cell nucleus. However, it is not known what binding elements in the histone tails are recognized by the individual Importins. Biochemical studies of H3 and H4 tails binding to seven Importins, Impβ, Kapβ2, Imp4, Imp5, Imp7, Imp9, and Impα, show the H3 tail binding more tightly than the H4 tail. The H3 tail binds Kapβ2 and Imp5 with KD values of 77 and 57 nm, respectively, and binds the other five Importins more weakly. Mutagenic analysis shows H3 tail residues 11–27 to be the sole binding segment for Impβ, Kapβ2, and Imp4. However, Imp5, Imp7, Imp9, and Impα bind two separate elements in the H3 tail: the segment at residues 11–27 and an isoleucine-lysine nuclear localization signal (IK-NLS) motif at residues 35–40. The H4 tail also uses either one or two basic segments to bind the same set of Importins with a similar trend of relative affinities as the H3 tail, albeit at least 10-fold weaker. Of the many lysine residues in the H3 and H4 tails, only acetylation of the H3 Lys14 substantially decreased binding to several Importins. Lastly, we show that, in addition to the N-terminal tails, the histone fold domains of H3 and H4 and/or the histone chaperone Asf1b are important for Importin-histone recognition. PMID:27528606

  5. Characterization of histone H3K27 modifications in the {beta}-globin locus

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yea Woon [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Pusan 609-735 (Korea, Republic of); Kim, AeRi, E-mail: kimaeri@pusan.ac.kr [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Pusan 609-735 (Korea, Republic of)

    2011-02-11

    Research highlights: {yields} The {beta}-globin locus control region is hyperacetylated and monomethylated at histone H3K27. {yields} Highly transcribed globin genes are marked by H3K27ac, but H3K27me2 is remarkable at silent globin genes in erythroid K562 cells. {yields} Association of PRC2 subunits is comparable with H3K27me3 pattern. {yields} Modifications of histone H3K27 are established in an enhancer-dependent manner. -- Abstract: Histone H3K27 is acetylated or methylated in the environment of nuclear chromatin. Here, to characterize the modification pattern of H3K27 in locus control region (LCR) and to understand the correlation of various H3K27 modifications with transcriptional activity of genes, we analyzed the human {beta}-globin locus using the ChIP assay. The LCR of the human {beta}-globin locus was enriched by H3K27ac and H3K27me1 in erythroid K562 cells. The highly transcribed globin genes were hyperacetylated at H3K27, but the repressed globin genes were highly dimethylated at this lysine in these cells. However, in non-erythroid 293FT cells, the {beta}-globin locus was marked by a high level of H3K27me3. EZH2 and SUZ12, subunits of polycomb repressive complex 2, were comparably detected with the H3K27me3 pattern in K562 and 293FT cells. In addition, H3K27ac, H3K27me1 and H3K27me3 were established in an enhancer-dependent manner in a model minichromosomal locus containing an enhancer and its target gene. Taken together, these results show that H3K27 modifications have distinctive correlations with the chromatin state or transcription level of genes and are influenced by an enhancer.

  6. Histone H3 gene in the Pacific oyster, Crassostrea gigas Thunberg, 1793: molecular and cytogenetic characterisations

    Directory of Open Access Journals (Sweden)

    Karine Bouilly

    2010-12-01

    Full Text Available The Pacific oyster, Crassostrea gigas Thunberg, 1793 (2n = 20 is an economically important mollusc species cultured throughout the world. The most frequently used technique for molecular cytogenetic studies is fluorescence in situ hybridisation which offers new opportunities for the identification of oyster chromosomes. In oysters, it has been used to locate telomeric sequences, satellite DNA, simple sequence repeats, ribosomal RNA genes, and bacteriophage P1 clones. However, regarding chromosome identification, no study has been done with histone H3 gene. Histone H3 is among the most conserved eukaryotic proteins. Most histone H3 genes are repeatedly organised into clusters, which make them an ideal chromosomal marker. In bivalves, some data exist concerning sequence information but little knowledge is available concerning the physical mapping of histone genes. The histone H3 gene was sequenced in C. gigas and phylogenetic analysis revealed that C. gigas was more closely related to Ostrea edulis Linnaeus, 1758 and species of the genus Mytilus Linnaeus, 1758. In C. gigas, the histone H3 gene was mapped on two different pairs of chromosomes, one at an interstitial site on the long arm of chromosome pair 4, and the other on the telomeres of the smaller chromosome pair (pair 10. Polymorphism was detected on the telomeres of pair 10, once it was possible to observe single or double signals. Comparative chromosomal mapping should improve our understanding of bivalve genome organisation.

  7. Post-translational modification and mitochondrial relocalization of histone H3 during apoptosis induced by staurosporine.

    Science.gov (United States)

    Han, Min-Joon; Koc, Emine Cavdar; Koc, Hasan

    2014-07-18

    Post-translational modifications (PTMs) of histones such as phosphorylation, acetylation, and ubiquitination, collectively referred to as the "histone-code", have been known to regulate gene expression and chromatin condensation for over a decade. They are also implicated in processes such as DNA repair and apoptosis. However, the study of the phosphorylation of histones has been mainly focused on chromosome condensation and mitosis. Therefore, the phosphorylation of histones in apoptosis is not fully understood. It was recently demonstrated by Tang et al. that histones are released from nucleosome during apoptosis, an observation that is in agreement with our findings. In addition to the release of histones, the dephosphorylation of histone H3 at Thr-3 and Ser-10 was observed during apoptosis in some cancer cells. Our data suggest that the modification and release of histones could serve markers of apoptosis in human cancer cells. We also suggest that the released histones, especially H3, could be translocated to mitochondria during apoptosis. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Histone H3.3 and cancer: A potential reader connection.

    Science.gov (United States)

    Lan, Fei; Shi, Yang

    2015-06-02

    The building block of chromatin is nucleosome, which consists of 146 base pairs of DNA wrapped around a histone octamer composed of two copies of histone H2A, H2B, H3, and H4. Significantly, the somatic missense mutations of the histone H3 variant, H3.3, are associated with childhood and young-adult tumors, such as pediatric high-grade astrocytomas, as well as chondroblastoma and giant-cell tumors of the bone. The mechanisms by which these histone mutations cause cancer are by and large unclear. Interestingly, two recent studies identified BS69/ZMYND11, which was proposed to be a candidate tumor suppressor, as a specific reader for a modified form of H3.3 (H3.3K36me3). Importantly, some H3.3 cancer mutations are predicted to abrogate the H3.3K36me3/BS69 interaction, suggesting that this interaction may play an important role in tumor suppression. These new findings also raise the question of whether H3.3 cancer mutations may lead to the disruption and/or gain of interactions of additional cellular factors that contribute to tumorigenesis.

  9. Double chromodomains cooperate to recognize the methylated histone H3 tail

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, John F.; Mi, Li-Zhi; Chruszcz, Maksymilian; Cymborowski, Marcin; Clines, Katrina L.; Kim, Youngchang; Minor, Wladek; Rastinejad, Fraydoon; Khorasanizadeh, Sepideh (ANL/SBC); (UV)

    2010-07-19

    Chromodomains are modules implicated in the recognition of lysine-methylated histone tails and nucleic acids. CHD (for chromo-ATPase/helicase-DNA-binding) proteins regulate ATP-dependent nucleosome assembly and mobilization through their conserved double chromodomains and SWI2/SNF2 helicase/ATPase domain. The Drosophila CHD1 localizes to the interbands and puffs of the polytene chromosomes, which are classic sites of transcriptional activity. Other CHD isoforms (CHD3/4 or Mi-2) are important for nucleosome remodelling in histone deacetylase complexes. Deletion of chromodomains impairs nucleosome binding and remodelling by CHD proteins. Here we describe the structure of the tandem arrangement of the human CHD1 chromodomains, and its interactions with histone tails. Unlike HP1 and Polycomb proteins that use single chromodomains to bind to their respective methylated histone H3 tails, the two chromodomains of CHD1 cooperate to interact with one methylated H3 tail. We show that the human CHD1 double chromodomains target the lysine 4-methylated histone H3 tail (H3K4me), a hallmark of active chromatin. Methylammonium recognition involves two aromatic residues, not the three-residue aromatic cage used by chromodomains of HP1 and Polycomb proteins. Furthermore, unique inserts within chromodomain 1 of CHD1 block the expected site of H3 tail binding seen in HP1 and Polycomb, instead directing H3 binding to a groove at the inter-chromodomain junction.

  10. Histone H3 Serine 28 Is Essential for Efficient Polycomb-Mediated Gene Repression in Drosophila

    Directory of Open Access Journals (Sweden)

    Philip Yuk Kwong Yung

    2015-06-01

    Full Text Available Trimethylation at histone H3K27 is central to the polycomb repression system. Juxtaposed to H3K27 is a widely conserved phosphorylatable serine residue (H3S28 whose function is unclear. To assess the importance of H3S28, we generated a Drosophila H3 histone mutant with a serine-to-alanine mutation at position 28. H3S28A mutant cells lack H3S28ph on mitotic chromosomes but support normal mitosis. Strikingly, all methylation states of H3K27 drop in H3S28A cells, leading to Hox gene derepression and to homeotic transformations in adult tissues. These defects are not caused by active H3K27 demethylation nor by the loss of H3S28ph. Biochemical assays show that H3S28A nucleosomes are a suboptimal substrate for PRC2, suggesting that the unphosphorylated state of serine 28 is important for assisting in the function of polycomb complexes. Collectively, our data indicate that the conserved H3S28 residue in metazoans has a role in supporting PRC2 catalysis.

  11. Formaldehyde-induced histone H3 phosphorylation via JNK and the expression of proto-oncogenes.

    Science.gov (United States)

    Yoshida, Ikuma; Ibuki, Yuko

    2014-12-01

    Formaldehyde (FA) is a very reactive compound that forms DNA adducts and DNA-protein crosslinks, which are known to contribute to FA-induced mutations and carcinogenesis. Post-translational modifications to histones have recently attracted attention due to their link with cancer. In the present study, we examined histone modifications following a treatment with FA. FA significantly phosphorylated histone H3 at serine 10 (H3S10), and at serine 28 (H3S28), the time-course of which was similar to the phosphorylation of H2AX at serine 139 (γ-H2AX), a marker of DNA double strand breaks. The temporal deacetylation of H3 was observed due to the reaction of FA with the lysine residues of histones. The phosphorylation mechanism was then analyzed by focusing on H3S10. The nuclear distribution of the phosphorylation of H3S10 and γ-H2AX did not overlap, and the phosphorylation of H3S10 could not be suppressed with an inhibitor of ATM/ATR, suggesting that the phosphorylation of H3S10 was independent of the DNA damage response. ERK and JNK in the MAPK pathways were phosphorylated by the treatment with FA, in which the JNK pathway was the main target for phosphorylation. The phosphorylation of H3S10 increased at the promoter regions of c-fos and c-jun, indicating a relationship between FA-induced tumor promotion activity and phosphorylation of H3S10. These results suggested that FA both initiates and promotes cancer, as judged by an analysis of histone modifications. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Excess free histone H3 localizes to centrosomes for proteasome-mediated degradation during mitosis in metazoans.

    Science.gov (United States)

    Wike, Candice L; Graves, Hillary K; Wason, Arpit; Hawkins, Reva; Gopalakrishnan, Jay; Schumacher, Jill; Tyler, Jessica K

    2016-08-17

    The cell tightly controls histone protein levels in order to achieve proper packaging of the genome into chromatin, while avoiding the deleterious consequences of excess free histones. Our accompanying study has shown that a histone modification that loosens the intrinsic structure of the nucleosome, phosphorylation of histone H3 on threonine 118 (H3 T118ph), exists on centromeres and chromosome arms during mitosis. Here, we show that H3 T118ph localizes to centrosomes in humans, flies, and worms during all stages of mitosis. H3 abundance at the centrosome increased upon proteasome inhibition, suggesting that excess free histone H3 localizes to centrosomes for degradation during mitosis. In agreement, we find ubiquitinated H3 specifically during mitosis and within purified centrosomes. These results suggest that targeting of histone H3 to the centrosome for proteasome-mediated degradation is a novel pathway for controlling histone supply, specifically during mitosis.

  13. 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.

  14. Histone H3.3 maintains genome integrity during mammalian development.

    Science.gov (United States)

    Jang, Chuan-Wei; Shibata, Yoichiro; Starmer, Joshua; Yee, Della; Magnuson, Terry

    2015-07-01

    Histone H3.3 is a highly conserved histone H3 replacement variant in metazoans and has been implicated in many important biological processes, including cell differentiation and reprogramming. Germline and somatic mutations in H3.3 genomic incorporation pathway components or in H3.3 encoding genes have been associated with human congenital diseases and cancers, respectively. However, the role of H3.3 in mammalian development remains unclear. To address this question, we generated H3.3-null mouse models through classical genetic approaches. We found that H3.3 plays an essential role in mouse development. Complete depletion of H3.3 leads to developmental retardation and early embryonic lethality. At the cellular level, H3.3 loss triggers cell cycle suppression and cell death. Surprisingly, H3.3 depletion does not dramatically disrupt gene regulation in the developing embryo. Instead, H3.3 depletion causes dysfunction of heterochromatin structures at telomeres, centromeres, and pericentromeric regions of chromosomes, leading to mitotic defects. The resulting karyotypical abnormalities and DNA damage lead to p53 pathway activation. In summary, our results reveal that an important function of H3.3 is to support chromosomal heterochromatic structures, thus maintaining genome integrity during mammalian development. © 2015 Jang et al.; Published by Cold Spring Harbor Laboratory Press.

  15. WHSC1 links transcription elongation to HIRA-mediated histone H3.3 deposition

    Science.gov (United States)

    Sarai, Naoyuki; Nimura, Keisuke; Tamura, Tomohiko; Kanno, Tomohiko; Patel, Mira C; Heightman, Tom D; Ura, Kiyoe; Ozato, Keiko

    2013-01-01

    Actively transcribed genes are enriched with the histone variant H3.3. Although H3.3 deposition has been linked to transcription, mechanisms controlling this process remain elusive. We investigated the role of the histone methyltransferase Wolf–Hirschhorn syndrome candidate 1 (WHSC1) (NSD2/MMSET) in H3.3 deposition into interferon (IFN) response genes. IFN treatment triggered robust H3.3 incorporation into activated genes, which continued even after cessation of transcription. Likewise, UV radiation caused H3.3 deposition in UV-activated genes. However, in Whsc1−/− cells IFN- or UV-triggered H3.3 deposition was absent, along with a marked reduction in IFN- or UV-induced transcription. We found that WHSC1 interacted with the bromodomain protein 4 (BRD4) and the positive transcription elongation factor b (P-TEFb) and facilitated transcriptional elongation. WHSC1 also associated with HIRA, the H3.3-specific histone chaperone, independent of BRD4 and P-TEFb. WHSC1 and HIRA co-occupied IFN-stimulated genes and supported prolonged H3.3 incorporation, leaving a lasting transcriptional mark. Our results reveal a previously unrecognized role of WHSC1, which links transcriptional elongation and H3.3 deposition into activated genes through two molecularly distinct pathways. PMID:23921552

  16. Transcriptional regulation of 15-lipoxygenase expression by histone h3 lysine 4 methylation/demethylation.

    Science.gov (United States)

    Liu, Cheng; Xu, Dawei; Han, Hongya; Fan, Yidong; Schain, Frida; Xu, Zhonghua; Claesson, Hans-Erik; Björkholm, Magnus; Sjöberg, Jan

    2012-01-01

    15-Lipoxygenase-1 (15-LOX-1) oxidizes polyunsaturated fatty acids to a rich spectrum of biologically active metabolites and is implicated in physiological membrane remodelling, inflammation and apoptosis. Its deregulation is involved in the pathogenesis of diverse cancer and immune diseases. Recent experimental evidence reveals that dynamic histone methylation/demethylation mediated by histone methyltransferases and demethylases plays a critical role in regulation of chromatin remodelling and gene expression. In the present study, we compared the histone 3 lysine 4 (H3-K4) methylation status of the 15-LOX-1 promoter region of the two Hodgkin lymphoma (HL) cell lines L1236 and L428 with abundant and undetectable 15-LOX-1 expression, respectively. We identified a potential role of H3-K4 methylation in positive regulation of 15-LOX-1 transcription. Furthermore, we found that histone methyltransferase SMYD3 inhibition reduced 15-LOX-1 expression by decreasing promoter activity in L1236 cells. SMYD3 knock down in these cells abolished di-/trimethylation of H3-K4, attenuated the occupancy by the transactivator STAT6, and led to diminished histone H3 acetylation at the 15-LOX-1 promoter. In contrast, inhibition of SMCX, a JmjC-domain-containing H3-K4 tri-demethylase, upregulated 15-LOX-1 expression through induction of H3-K4 trimethylation, histone acetylation and STAT6 recruitment at the 15-LOX-1 promoter in L428 cells. In addition, we observed strong SMYD3 expression in the prostate cancer cell line LNCaP and its inhibition led to decreased 15-LOX-1 expression. Taken together, our data suggest that regulation of histone methylation/demethylation at the 15-LOX-1 promoter is important in 15-LOX-1 expression.

  17. Transcriptional regulation of 15-lipoxygenase expression by histone h3 lysine 4 methylation/demethylation.

    Directory of Open Access Journals (Sweden)

    Cheng Liu

    Full Text Available 15-Lipoxygenase-1 (15-LOX-1 oxidizes polyunsaturated fatty acids to a rich spectrum of biologically active metabolites and is implicated in physiological membrane remodelling, inflammation and apoptosis. Its deregulation is involved in the pathogenesis of diverse cancer and immune diseases. Recent experimental evidence reveals that dynamic histone methylation/demethylation mediated by histone methyltransferases and demethylases plays a critical role in regulation of chromatin remodelling and gene expression. In the present study, we compared the histone 3 lysine 4 (H3-K4 methylation status of the 15-LOX-1 promoter region of the two Hodgkin lymphoma (HL cell lines L1236 and L428 with abundant and undetectable 15-LOX-1 expression, respectively. We identified a potential role of H3-K4 methylation in positive regulation of 15-LOX-1 transcription. Furthermore, we found that histone methyltransferase SMYD3 inhibition reduced 15-LOX-1 expression by decreasing promoter activity in L1236 cells. SMYD3 knock down in these cells abolished di-/trimethylation of H3-K4, attenuated the occupancy by the transactivator STAT6, and led to diminished histone H3 acetylation at the 15-LOX-1 promoter. In contrast, inhibition of SMCX, a JmjC-domain-containing H3-K4 tri-demethylase, upregulated 15-LOX-1 expression through induction of H3-K4 trimethylation, histone acetylation and STAT6 recruitment at the 15-LOX-1 promoter in L428 cells. In addition, we observed strong SMYD3 expression in the prostate cancer cell line LNCaP and its inhibition led to decreased 15-LOX-1 expression. Taken together, our data suggest that regulation of histone methylation/demethylation at the 15-LOX-1 promoter is important in 15-LOX-1 expression.

  18. 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.

  19. A role for CARM1-mediated histone H3 arginine methylation in protecting histone acetylation by releasing corepressors from chromatin.

    Directory of Open Access Journals (Sweden)

    Jing Wu

    Full Text Available Arginine methylation broadly occurs in histones and has been linked to transcriptional regulation, cell cycle regulation and DNA repair. While numerous proteins (histone code effectors that specifically recognize or read the methylated lysine residues in core histones have been identified, little is known for effectors specific for methylated arginines in histones. In this study, we attempted to identify effector(s recognizing asymmetrically methylated R17 and R26 in H3, which are catalyzed by CARM1/PRMT4, through an unbiased biochemical approach. Although we have yet to identify such effector using this approach, we find that these modifications function cooperatively with histone acetylation to inhibit the binding of the nucleosome remodeling and deacetylase complex (NuRD and TIF1 family corepressors to H3 tail in vitro. In support of this finding, we show that overexpression of CARM1 in 293 T cells leads to reduced association of NuRD with chromatin, whereas knockdown of CARM1 in HeLa cells leads to increased association of NuRD with chromatin and decreased level of histone acetylation. Furthermore, in the Carm1-/- MEF cells there is an increased association of NuRD and TIF1β with chromatin and a global decrease in histone acetylation. By chromatin immunoprecipitation assay, we show that overexpression of CARM1 results in reduced association of NuRD complex and TIF1β with an episomal reporter and that CARM1 is required in MEF cells for LPS-induced dissociation of NuRD from a NF-κb target gene. Taking together, our study provides evidence for a role of CARM1-mediated arginine methylation in regulation of histone acetylation and transcription: facilitating transcription by discharging corepressors from chromatin.

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

    Directory of Open Access Journals (Sweden)

    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.

  1. Histone H3 lysine 4 acetylation and methylation dynamics define breast cancer subtypes.

    Science.gov (United States)

    Messier, Terri L; Gordon, Jonathan A R; Boyd, Joseph R; Tye, Coralee E; Browne, Gillian; Stein, Janet L; Lian, Jane B; Stein, Gary S

    2016-02-02

    The onset and progression of breast cancer are linked to genetic and epigenetic changes that alter the normal programming of cells. Epigenetic modifications of DNA and histones contribute to chromatin structure that result in the activation or repression of gene expression. Several epigenetic pathways have been shown to be highly deregulated in cancer cells. Targeting specific histone modifications represents a viable strategy to prevent oncogenic transformation, tumor growth or metastasis. Methylation of histone H3 lysine 4 has been extensively studied and shown to mark genes for expression; however this residue can also be acetylated and the specific function of this alteration is less well known. To define the relative roles of histone H3 methylation (H3K4me3) and acetylation (H3K4ac) in breast cancer, we determined genomic regions enriched for both marks in normal-like (MCF10A), transformed (MCF7) and metastatic (MDA-MB-231) cells using a genome-wide ChIP-Seq approach. Our data revealed a genome-wide gain of H3K4ac associated with both early and late breast cancer cell phenotypes, while gain of H3K4me3 was predominantly associated with late stage cancer cells. Enrichment of H3K4ac was over-represented at promoters of genes associated with cancer-related phenotypic traits, such as estrogen response and epithelial-to-mesenchymal transition pathways. Our findings highlight an important role for H3K4ac in predicting epigenetic changes associated with early stages of transformation. In addition, our data provide a valuable resource for understanding epigenetic signatures that correlate with known breast cancer-associated oncogenic pathways.

  2. In silico probing and biological evaluation of SETDB1/ESET-targeted novel compounds that reduce tri-methylated histone H3K9 (H3K9me3) level

    Science.gov (United States)

    Park, Insun; Hwang, Yu Jin; Kim, TaeHun; Viswanath, Ambily Nath Indu; Londhe, Ashwini M.; Jung, Seo Yun; Sim, Kyoung Mi; Min, Sun-Joon; Lee, Ji Eun; Seong, Jihye; Kim, Yun Kyung; No, Kyoung Tai; Ryu, Hoon; Pae, Ae Nim

    2017-09-01

    ERG-associated protein with the SET domain (ESET/SET domain bifurcated 1/SETDB1/KMT1E) is a histone lysine methyltransferase (HKMT) and it preferentially tri-methylates lysine 9 of histone H3 (H3K9me3). SETDB1/ESET leads to heterochromatin condensation and epigenetic gene silencing. These functional changes are reported to correlate with Huntington's disease (HD) progression and mood-related disorders which make SETDB1/ESET a viable drug target. In this context, the present investigation was performed to identify novel peptide-competitive small molecule inhibitors of the SETDB1/ESET by a combined in silico-in vitro approach. A ligand-based pharmacophore model was built and employed for the virtual screening of ChemDiv and Asinex database. Also, a human SETDB1/ESET homology model was constructed to supplement the data further. Biological evaluation of the selected 21 candidates singled out 5 compounds exhibiting a notable reduction of the H3K9me3 level via inhibitory potential of SETDB1/ESET activity in SETDB1/ESET-inducible cell line and HD striatal cells. Later on, we identified two compounds as final hits that appear to have neuronal effects without cytotoxicity based on the result from MTT assay. These compounds hold the calibre to become the future lead compounds and can provide structural insights into more SETDB1/ESET-focused drug discovery research. Moreover, these SETDB1/ESET inhibitors may be applicable for the preclinical study to ameliorate neurodegenerative disorders via epigenetic regulation.

  3. In silico probing and biological evaluation of SETDB1/ESET-targeted novel compounds that reduce tri-methylated histone H3K9 (H3K9me3) level

    Science.gov (United States)

    Park, Insun; Hwang, Yu Jin; Kim, TaeHun; Viswanath, Ambily Nath Indu; Londhe, Ashwini M.; Jung, Seo Yun; Sim, Kyoung Mi; Min, Sun-Joon; Lee, Ji Eun; Seong, Jihye; Kim, Yun Kyung; No, Kyoung Tai; Ryu, Hoon; Pae, Ae Nim

    2017-10-01

    ERG-associated protein with the SET domain (ESET/SET domain bifurcated 1/SETDB1/KMT1E) is a histone lysine methyltransferase (HKMT) and it preferentially tri-methylates lysine 9 of histone H3 (H3K9me3). SETDB1/ESET leads to heterochromatin condensation and epigenetic gene silencing. These functional changes are reported to correlate with Huntington's disease (HD) progression and mood-related disorders which make SETDB1/ESET a viable drug target. In this context, the present investigation was performed to identify novel peptide-competitive small molecule inhibitors of the SETDB1/ESET by a combined in silico-in vitro approach. A ligand-based pharmacophore model was built and employed for the virtual screening of ChemDiv and Asinex database. Also, a human SETDB1/ESET homology model was constructed to supplement the data further. Biological evaluation of the selected 21 candidates singled out 5 compounds exhibiting a notable reduction of the H3K9me3 level via inhibitory potential of SETDB1/ESET activity in SETDB1/ESET-inducible cell line and HD striatal cells. Later on, we identified two compounds as final hits that appear to have neuronal effects without cytotoxicity based on the result from MTT assay. These compounds hold the calibre to become the future lead compounds and can provide structural insights into more SETDB1/ESET-focused drug discovery research. Moreover, these SETDB1/ESET inhibitors may be applicable for the preclinical study to ameliorate neurodegenerative disorders via epigenetic regulation.

  4. 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

    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...... histones genome wide during DNA replication....

  5. BRPF3-HBO1 regulates replication origin activation and histone H3K14 acetylation

    DEFF Research Database (Denmark)

    Feng, Yunpeng; Vlassis, Arsenios; Roques, Céline

    2016-01-01

    recruitment, but not MCM2-7 loading, is impaired in BRPF3-depleted cells, identifying a BRPF3-dependent function of HBO1 in origin activation that is complementary to its role in licencing. We thus propose that BRPF3-HBO1 acetylation of histone H3K14 around TSS facilitates efficient activation of nearby...

  6. Lysyl oxidase-like 2 (LOXL2) oxidizes trimethylated lysine 4 in histone H3.

    Science.gov (United States)

    Herranz, Nicolás; Dave, Natàlia; Millanes-Romero, Alba; Pascual-Reguant, Laura; Morey, Lluis; Díaz, Víctor M; Lórenz-Fonfría, Víctor; Gutierrez-Gallego, Ricardo; Jerónimo, Celia; Iturbide, Ane; Di Croce, Luciano; García de Herreros, Antonio; Peiró, Sandra

    2016-12-01

    Methylation of histone H3 lysine 4 is linked to active transcription and can be removed by LSD1 or the JmjC domain-containing proteins by amino-oxidation or hydroxylation, respectively. Here we describe that its deamination can be catalyzed by lysyl oxidase-like 2 protein (LOXL2), presenting an unconventional chemical mechanism for H3K4 modification. Infrared spectroscopy and mass spectrometry analyses demonstrated that recombinant LOXL2 specifically deaminates trimethylated H3K4. Moreover, by regulating H3K4me3 deamination, LOXL2 activity is linked with the transcriptional control of the CDH1 gene. These results reveal the existence of further H3 modification as well as a novel mechanism for H3K4me3 demethylation. The GEO accession number for the data referred to this paper is GSE35600. © 2016 Federation of European Biochemical Societies.

  7. The histone H3 methyltransferase G9A epigenetically activates the serine-glycine synthesis pathway to sustain cancer cell survival and proliferation.

    Science.gov (United States)

    Ding, Jane; Li, Tai; Wang, Xiangwei; Zhao, Erhu; Choi, Jeong-Hyeon; Yang, Liqun; Zha, Yunhong; Dong, Zheng; Huang, Shuang; Asara, John M; Cui, Hongjuan; Ding, Han-Fei

    2013-12-03

    Increased activation of the serine-glycine biosynthetic pathway is an integral part of cancer metabolism that drives macromolecule synthesis needed for cell proliferation. Whether this pathway is under epigenetic control is unknown. Here we show that the histone H3 lysine 9 (H3K9) methyltransferase G9A is required for maintaining the pathway enzyme genes in an active state marked by H3K9 monomethylation and for the transcriptional activation of this pathway in response to serine deprivation. G9A inactivation depletes serine and its downstream metabolites, triggering cell death with autophagy in cancer cell lines of different tissue origins. Higher G9A expression, which is observed in various cancers and is associated with greater mortality in cancer patients, increases serine production and enhances the proliferation and tumorigenicity of cancer cells. These findings identify a G9A-dependent epigenetic program in the control of cancer metabolism, providing a rationale for G9A inhibition as a therapeutic strategy for cancer. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Relationship between lunasin's sequence and its inhibitory activity of histones H3 and H4 acetylation.

    Science.gov (United States)

    Hernández-Ledesma, Blanca; Hsieh, Chia-Chien; de Lumen, Ben O

    2011-07-01

    Dysfunction of histone acetyltransferases (HATs) or histone deacetylases (HDACs) involved in histones acetylation has been associated with cancer. Inhibitors of these enzymes are becoming potential targets for new therapies. This study reports by Western-Blot analysis, that peptide lunasin is mainly an in vitro inhibitor of histone H4 acetylation by P300/cAMP-response element-binding protein (CBP)-associated factor (PCAF), with IC₅₀ values dependent on the lysine position sensitive to be acetylated (0.83 μM (H4-Lys 8), 1.27 μM (H4-Lys 12) and 0.40 μM (H4-Lys 5, 8, 12, 16)). Lunasin is also capable of inhibiting H3 acetylation (IC₅₀ of 5.91 μM (H3-Lys 9) and 7.81 μM (H3-Lys 9, 14)). Studies on structure-activity relationship establish that lunasin's sequence are essential for inhibiting H4 acetylation whereas poly-D sequence is the main active sequence responsible for H3 acetylation inhibition. Lunasin also inhibits H3 and H4 acetylation and cell proliferation (IC₅₀ of 181 μM) in breast cancer MDA-MB-231 cells. Moreover, this peptide decreases expression of cyclins and cyclin dependent kinases-4 and -6, implicated in cell cycle pathways. Results from this study demonstrates lunasin's role as modulator of histone acetylation and protein expression that might contribute on its chemopreventive properties against breast cancer. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Histone H3 Methylated at Arginine 17 Is Essential for Reprogramming the Paternal Genome in Zygotes

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    Yuki Hatanaka

    2017-09-01

    Full Text Available At fertilization, the paternal genome undergoes extensive reprogramming through protamine-histone exchange and active DNA demethylation, but only a few maternal factors have been defined in these processes. We identified maternal Mettl23 as a protein arginine methyltransferase (PRMT, which most likely catalyzes the asymmetric dimethylation of histone H3R17 (H3R17me2a, as indicated by in vitro assays and treatment with TBBD, an H3R17 PRMT inhibitor. Maternal histone H3.3, which is essential for paternal nucleosomal assembly, is unable to be incorporated into the male pronucleus when it lacks R17me2a. Mettl23 interacts with Tet3, a 5mC-oxidizing enzyme responsible for active DNA demethylation, by binding to another maternal factor, GSE (gonad-specific expression. Depletion of Mettl23 from oocytes resulted in impaired accumulation of GSE, Tet3, and 5hmC in the male pronucleus, suggesting that Mettl23 may recruit GSE-Tet3 to chromatin. Our findings establish H3R17me2a and its catalyzing enzyme Mettl23 as key regulators of paternal genome reprogramming.

  10. Changes in histone H3 lysine 36 methylation in porcine oocytes and preimplantation embryos.

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    Yun Fei Diao

    Full Text Available Histone H3 lysine 36 (H3K36 methylation is known to be associated with transcriptionally active genes, and is considered a genomic marker of active loci. To investigate the changes in H3K36 methylation in pig, we determined the mono-, di-, and tri-methylations of H3K36 (H3K36me1, H3K36me2 and H3K36me3, respectively in porcine fetal fibroblasts, oocytes and preimplantation embryos by immunocytochemistry using specific antibodies and confocal microscopy. These analyses revealed that only H3K36me3 in porcine fetal fibroblasts consistently colocalized with transcription sites identified as actively synthesizing RNA based on fluorouridine (FU incorporation. Treatment of cells with flavopiridol, which blocks transcription elongation, completely abrogated both H3K36me3 signals and RNA synthesis. All three types of H3K36 methylation were present and did not significantly differ during oocyte maturation. In parthenogenetic embryos, H3K36me1 and -me2 were detected in 1-cell through blastocyst-stage embryos. In contrast, H3K36me3 was not detected in most 1-cell stage embryos. H3K36me3 signals became detectable in 2-cell stage embryos, peaked at the 4-cell stage, decreased at the 8-cell stage, and then became undetectable at blastocyst stages in both parthenogenetic and in vitro-fertilized (IVF embryos. Unlike the case in IVF embryos, H3K36me3 could not be demethylated completely during the 1-cell stage in somatic cell nuclear transfer (SCNT embryos. These results collectively indicate that H3K36me3, but not H3K36me1 or -me2, is associated with transcription elongation in porcine fetal fibroblasts. H3K36me3 is developmentally regulated and may be a histone mark of embryonic gene activation in pig. Aberrant H3K36 tri-methylation occurred during the nuclear reprogramming of SCNT embryos.

  11. Nucleosome Binding Alters the Substrate Bonding Environment of Histone H3 Lysine 36 Methyltransferase NSD2.

    Science.gov (United States)

    Poulin, Myles B; Schneck, Jessica L; Matico, Rosalie E; Hou, Wangfang; McDevitt, Patrick J; Holbert, Marc; Schramm, Vern L

    2016-06-01

    Nuclear receptor-binding SET domain protein 2 (NSD2) is a histone H3 lysine 36 (H3K36)-specific methyltransferase enzyme that is overexpressed in a number of cancers, including multiple myeloma. NSD2 binds to S-adenosyl-l-methionine (SAM) and nucleosome substrates to catalyze the transfer of a methyl group from SAM to the ε-amino group of histone H3K36. Equilibrium binding isotope effects and density functional theory calculations indicate that the SAM methyl group is sterically constrained in complex with NSD2, and that this steric constraint is released upon nucleosome binding. Together, these results show that nucleosome binding to NSD2 induces a significant change in the chemical environment of enzyme-bound SAM.

  12. RPA binds histone H3-H4 and functions in DNA replication-coupled nucleosome assembly.

    Science.gov (United States)

    Liu, Shaofeng; Xu, Zhiyun; Leng, He; Zheng, Pu; Yang, Jiayi; Chen, Kaifu; Feng, Jianxun; Li, Qing

    2017-01-27

    DNA replication-coupled nucleosome assembly is essential to maintain genome integrity and retain epigenetic information. Multiple involved histone chaperones have been identified, but how nucleosome assembly is coupled to DNA replication remains elusive. Here we show that replication protein A (RPA), an essential replisome component that binds single-stranded DNA, has a role in replication-coupled nucleosome assembly. RPA directly binds free H3-H4. Assays using a synthetic sequence that mimics freshly unwound single-stranded DNA at replication fork showed that RPA promotes DNA-(H3-H4) complex formation immediately adjacent to double-stranded DNA. Further, an RPA mutant defective in H3-H4 binding exhibited attenuated nucleosome assembly on nascent chromatin. Thus, we propose that RPA functions as a platform for targeting histone deposition to replication fork, through which RPA couples nucleosome assembly with ongoing DNA replication. Copyright © 2017, American Association for the Advancement of Science.

  13. Histone H3 variants and their chaperones during development and disease: contributing to epigenetic control.

    Science.gov (United States)

    Filipescu, Dan; Müller, Sebastian; Almouzni, Geneviève

    2014-01-01

    Within the nucleus, the interplay between lineage-specific transcription factors and chromatin dynamics defines cellular identity. Control of this interplay is necessary to properly balance stability and plasticity during the development and entire life span of multicellular organisms. Here, we present our current knowledge of the contribution of histone H3 variants to chromatin dynamics during development. We review the network of histone chaperones that governs their deposition timing and sites of incorporation and highlight how their distinct distribution impacts genome organization and function. We integrate the importance of H3 variants in the context of nuclear reprogramming and cell differentiation, and, using the centromere as a paradigm, we describe a case in which the identity of a given genomic locus is propagated across different cell types. Finally, we compare development to changes in stress and disease. Both physiological and pathological settings underline the importance of H3 dynamics for genome and chromatin integrity.

  14. Molecular pathways: deregulation of histone h3 lysine 27 methylation in cancer-different paths, same destination

    National Research Council Canada - National Science Library

    Ezponda, Teresa; Licht, Jonathan D

    2014-01-01

    Methylation of lysine 27 on histone H3 (H3K27me), a modification associated with gene repression, plays a critical role in regulating the expression of genes that determine the balance between cell differentiation and proliferation...

  15. Distinct factors control histone variant H3.3 localization at specific genomic regions

    Science.gov (United States)

    Goldberg, Aaron D.; Banaszynski, Laura A.; Noh, Kyung-Min; Lewis, Peter W.; Elsaesser, Simon J.; Stadler, Sonja; Dewell, Scott; Law, Martin; Guo, Xingyi; Li, Xuan; Wen, Duancheng; Chapgier, Ariane; DeKelver, Russell C.; Miller, Jeffrey C.; Lee, Ya-Li; Boydston, Elizabeth A.; Holmes, Michael C.; Gregory, Philip D.; Greally, John M.; Rafii, Shahin; Yang, Chingwen; Scambler, Peter J.; Garrick, David; Gibbons, Richard J.; Higgs, Douglas R.; Cristea, Ileana M.; Urnov, Fyodor D.; Zheng, Deyou; Allis, C. David

    2010-01-01

    Summary The incorporation of histone H3 variants has been implicated in the epigenetic memory of cellular state. Using genome editing with zinc finger nucleases to tag endogenous H3.3, we report genome-wide profiles of H3 variants in mammalian embryonic stem (ES) cells and neuronal precursor cells. Genome-wide patterns of H3.3 are dependent on amino acid sequence, and change with cellular differentiation at developmentally regulated loci. The H3.3 chaperone Hira is required for H3.3 enrichment at active and repressed genes. Strikingly, Hira is not essential for localization of H3.3 at telomeres and many transcription factor binding sites. Immunoaffinity purification and mass spectrometry reveal that the proteins Atrx and Daxx associate with H3.3 in a Hira-independent manner. Atrx is required for Hira-independent localization of H3.3 at telomeres, and for the repression of telomeric RNA. Our data demonstrate that multiple and distinct factors are responsible for H3.3 localization at specific genomic locations in mammalian cells. PMID:20211137

  16. The histone H3.3K36M mutation reprograms the epigenome of chondroblastomas.

    Science.gov (United States)

    Fang, Dong; Gan, Haiyun; Lee, Jeong-Heon; Han, Jing; Wang, Zhiquan; Riester, Scott M; Jin, Long; Chen, Jianji; Zhou, Hui; Wang, Jinglong; Zhang, Honglian; Yang, Na; Bradley, Elizabeth W; Ho, Thai H; Rubin, Brian P; Bridge, Julia A; Thibodeau, Stephen N; Ordog, Tamas; Chen, Yue; van Wijnen, Andre J; Oliveira, Andre M; Xu, Rui-Ming; Westendorf, Jennifer J; Zhang, Zhiguo

    2016-06-10

    More than 90% of chondroblastomas contain a heterozygous mutation replacing lysine-36 with methionine-36 (K36M) in the histone H3 variant H3.3. Here we show that H3K36 methylation is reduced globally in human chondroblastomas and in chondrocytes harboring the same genetic mutation, due to inhibition of at least two H3K36 methyltransferases, MMSET and SETD2, by the H3.3K36M mutant proteins. Genes with altered expression as well as H3K36 di- and trimethylation in H3.3K36M cells are enriched in cancer pathways. In addition, H3.3K36M chondrocytes exhibit several hallmarks of cancer cells, including increased ability to form colonies, resistance to apoptosis, and defects in differentiation. Thus, H3.3K36M proteins reprogram the H3K36 methylation landscape and contribute to tumorigenesis, in part through altering the expression of cancer-associated genes. Copyright © 2016, American Association for the Advancement of Science.

  17. Holocarboxylase Synthetase 1 Physically Interacts with Histone H3 in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Xi Chen

    2013-01-01

    Full Text Available Biotin is a water-soluble vitamin required by all organisms, but only synthesized by plants and some bacterial and fungal species. As a cofactor, biotin is responsible for carbon dioxide transfer in all biotin-dependent carboxylases, including acetyl-CoA carboxylase, methylcrotonyl-CoA carboxylase, and pyruvate carboxylase. Adding biotin to carboxylases is catalyzed by the enzyme holocarboxylase synthetase (HCS. Biotin is also involved in gene regulation, and there is some indication that histones can be biotinylated in humans. Histone proteins and most histone modifications are highly conserved among eukaryotes. HCS1 is the only functional biotin ligase in Arabidopsis and has a high homology with human HCS. Therefore, we hypothesized that HCS1 also biotinylates histone proteins in Arabidopsis. A comparison of the catalytic domain of HCS proteins was performed among eukaryotes, prokaryotes, and archaea, and this domain is highly conserved across the selected organisms. Biotinylated histones could not be identified in vivo by using avidin precipitation or two-dimensional gel analysis. However, HCS1 physically interacts with Arabidopsis histone H3 in vitro, indicating the possibility of the role of this enzyme in the regulation of gene expression.

  18. ASH1L Links Histone H3 Lysine 36 Dimethylation to MLL Leukemia.

    Science.gov (United States)

    Zhu, Li; Li, Qin; Wong, Stephen H K; Huang, Min; Klein, Brianna J; Shen, Jinfeng; Ikenouye, Larissa; Onishi, Masayuki; Schneidawind, Dominik; Buechele, Corina; Hansen, Loren; Duque-Afonso, Jesús; Zhu, Fangfang; Martin, Gloria Mas; Gozani, Or; Majeti, Ravindra; Kutateladze, Tatiana G; Cleary, Michael L

    2016-07-01

    Numerous studies in multiple systems support that histone H3 lysine 36 dimethylation (H3K36me2) is associated with transcriptional activation; however, the underlying mechanisms are not well defined. Here, we show that the H3K36me2 chromatin mark written by the ASH1L histone methyltransferase is preferentially bound in vivo by LEDGF, a mixed-lineage leukemia (MLL)-associated protein that colocalizes with MLL, ASH1L, and H3K36me2 on chromatin genome wide. Furthermore, ASH1L facilitates recruitment of LEDGF and wild-type MLL proteins to chromatin at key leukemia target genes and is a crucial regulator of MLL-dependent transcription and leukemic transformation. Conversely, KDM2A, an H3K36me2 demethylase and Polycomb group silencing protein, antagonizes MLL-associated leukemogenesis. Our studies are the first to provide a basic mechanistic insight into epigenetic interactions wherein placement, interpretation, and removal of H3K36me2 contribute to the regulation of gene expression and MLL leukemia, and suggest ASH1L as a novel target for therapeutic intervention. Epigenetic regulators play vital roles in cancer pathogenesis and represent a new frontier in therapeutic targeting. Our studies provide basic mechanistic insight into the role of H3K36me2 in transcription activation and MLL leukemia pathogenesis and implicate ASH1L histone methyltransferase as a promising target for novel molecular therapy. Cancer Discov; 6(7); 770-83. ©2016 AACR.See related commentary by Balbach and Orkin, p. 700This article is highlighted in the In This Issue feature, p. 681. ©2016 American Association for Cancer Research.

  19. Linker histone H1 and H3K56 acetylation are antagonistic regulators of nucleosome dynamics.

    Science.gov (United States)

    Bernier, Morgan; Luo, Yi; Nwokelo, Kingsley C; Goodwin, Michelle; Dreher, Sarah J; Zhang, Pei; Parthun, Mark R; Fondufe-Mittendorf, Yvonne; Ottesen, Jennifer J; Poirier, Michael G

    2015-12-09

    H1 linker histones are highly abundant proteins that compact nucleosomes and chromatin to regulate DNA accessibility and transcription. However, the mechanisms that target H1 regulation to specific regions of eukaryotic genomes are unknown. Here we report fluorescence measurements of human H1 regulation of nucleosome dynamics and transcription factor (TF) binding within nucleosomes. H1 does not block TF binding, instead it suppresses nucleosome unwrapping to reduce DNA accessibility within H1-bound nucleosomes. We then investigated H1 regulation by H3K56 and H3K122 acetylation, two transcriptional activating histone post translational modifications (PTMs). Only H3K56 acetylation, which increases nucleosome unwrapping, abolishes H1.0 reduction of TF binding. These findings show that nucleosomes remain dynamic, while H1 is bound and H1 dissociation is not required for TF binding within the nucleosome. Furthermore, our H3K56 acetylation measurements suggest that a single-histone PTM can define regions of the genome that are not regulated by H1.

  20. Contributions of histone H3 nucleosome core surface mutations to chromatin structures, silencing and DNA repair.

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    Michel Fink

    Full Text Available Histone H3 mutations in residues that cluster in a discrete region on the nucleosome surface around lysine 79 of H3 affect H3-K79 methylation, impair transcriptional silencing in subtelomeric chromatin, and reveal distinct contributions of histone H3 to various DNA-damage response and repair pathways. These residues might act by recruitment of silencing and DNA-damage response factors. Alternatively, their location on the nucleosome surface suggests a possible involvement in nucleosome positioning, stability and nucleosome interactions. Here, we show that the yeast H3 mutants hht2-T80A, hht2-K79E, hht2-L70S, and hht2-E73D show normal nucleosome positioning and stability in minichromosomes. However, loss of silencing in a subtelomeric URA3 gene correlates with a shift of the promoter nucleosome, while nucleosome positions and stability in the coding region are maintained. Moreover, the H3 mutants show normal repair of UV lesions by photolyase and nucleotide excision repair in minichromosomes and slightly enhanced repair in the subtelomeric region. Thus, these results support a role of those residues in the recruitment of silencing proteins and argue against a general role in nucleosome organization.

  1. DNA promoter and histone H3 methylation downregulate NGX6 in gastric cancer cells.

    Science.gov (United States)

    Liu, Jian; Zhu, Xinjiang; Xu, Xiaoyang; Dai, Dongqiu

    2014-01-01

    Nasopharyngeal carcinoma-associated gene 6 (NGX6) is a novel candidate tumor metastasis suppressor gene. Our study was to determine whether DNA hypermethylation and histone modification at the NGX6 gene promoter play important roles in silencing NGX6 expression in gastric cancer. NGX6 expression was downregulated in all gastric cancer cells and 76.19 % tissues. In three GC cell lines, hypermethylated NGX6 loci were characterized by histone H3-K9 hypoacetylation and hypermethylation. Trichostatin A treatment could moderately increase H3-K9 acetylation at the silenced loci; however, it had no effect on DNA and H3-K9 methylation and minimal effects on NGX6 expression. In contrast, 5'aza-2'-deoxycytidine treatment could rapidly decrease DNA and H3-K9 methylation at the silenced loci, leading to the reexpression of NGX6. Combined treatment with 5'aza-2'-deoxycytidine and trichostatin A had synergistic effects on the reexpression of NGX6 at the hypermethylation loci. Our current study shows that NGX6 expression is downregulated in GC cancer cells and tissues due to NGX6 promoter methylation and H3-K9 methylation, but not H3-K9 acetylation. Our findings indicate that the downregulation of NGX6 expression contributes to the development and progression of gastric cancer. More studies are needed to determine the precise mechanism of NGX6 in the progression of gastric cancer.

  2. Basal aurora kinase B activity is sufficient for histone H3 phosphorylation in prophase

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    Ly-Thuy-Tram Le

    2013-02-01

    Histone H3 phosphorylation is the hallmark of mitosis deposited by aurora kinase B. Benzo[e]pyridoindoles are a family of potent, broad, ATP-competitive aurora kinase inhibitors. However, benzo[e]pyridoindole C4 only inhibits histone H3 phosphorylation in prophase but not in metaphase. Under the C4 treatment, the cells enter into mitosis with dephosphorylated histone H3, assemble chromosomes normally and progress to metaphase, and then to anaphase. C4 also induces lagging chromosome in anaphase but we demonstrated that these chromosome compaction defects are not related to the absence of H3 phosphorylation in prophase. As a result of C4 action, mitosis lasts longer and the cell cycle is slowed down. We reproduced the mitotic defects with reduced concentrations of potent pan aurora kinase as well as with a specific aurora B ATP-competitive inhibitor; we therefore propose that histone H3 phosphorylation and anaphase chromosome compaction involve the basal activity of aurora kinase B. Our data suggest that aurora kinase B is progressively activated at mitosis entry and at anaphase onset. The full activation of aurora kinase B by its partners, in prometaphase, induces a shift in the catalytic domain of aurora B that modifies its affinity for ATP. These waves of activation/deactivation of aurora B correspond to different conformations of the chromosomal complex revealed by FRAP. The presence of lagging chromosomes may have deleterious consequences on the daughter cells and, unfortunately, the situation may be encountered in patients receiving treatment with aurora kinase inhibitors.

  3. The histone H3.3K27M mutation in pediatric glioma reprograms H3K27 methylation and gene expression

    OpenAIRE

    Chan, Kui-Ming; Fang, Dong; Gan, Haiyun; Hashizume, Rintaro; Yu, Chuanhe; Schroeder, Mark; Gupta, Nalin; Mueller, Sabine; James, C. David; Jenkins, Robert; Sarkaria, Jann; Zhang, Zhiguo

    2013-01-01

    Sixty percent of high-grade pediatric glioma cases harbor a K27M mutation at one allele of H3F3A, one of the two genes encoding histone H3 variant H3.3. Chan et al. now find that the levels of H3K27 di- and trimethylation (H3K27me2 and H3K27me3) are reduced globally in H3.3K27M patient samples. H3K27me3 and Ezh2 (the catalytic subunit of H3K27 methyltransferase) are increased locally at hundreds of gene loci in H3.3K27M patient cells, altering the expression of genes that are associated with ...

  4. DNA Mismatch Repair Interacts with CAF-1- and ASF1A-H3-H4-dependent Histone (H3-H4)2 Tetramer Deposition.

    Science.gov (United States)

    Rodriges Blanko, Elena; Kadyrova, Lyudmila Y; Kadyrov, Farid A

    2016-04-22

    DNA mismatch repair (MMR) is required for the maintenance of genome stability and protection of humans from several types of cancer. Human MMR occurs in the chromatin environment, but little is known about the interactions between MMR and the chromatin environment. Previous research has suggested that MMR coincides with replication-coupled assembly of the newly synthesized DNA into nucleosomes. The first step in replication-coupled nucleosome assembly is CAF-1-dependent histone (H3-H4)2 tetramer deposition, a process that involves ASF1A-H3-H4 complex. In this work we used reconstituted human systems to investigate interactions between MMR and CAF-1- and ASF1A-H3-H4-dependent histone (H3-H4)2 tetramer deposition. We have found that MutSα inhibits CAF-1- and ASF1A-H3-H4-dependent packaging of a DNA mismatch into a tetrasome. This finding supports the idea that MMR occurs before the DNA mismatch is packaged into the tetrasome. Our experiments have also revealed that CAF-1- and ASF1A-H3-H4-dependent deposition of the histone (H3-H4)2 tetramers does not interfere with MMR reactions. In addition, we have established that unnecessary degradation of the discontinuous strand that takes place in both DNA polymerase δ (Pol δ)- and DNA polymerase ϵ (Pol ϵ)-dependent MMR reactions is suppressed by CAF-1- and ASF1A-H3-H4-dependent deposition of the histone (H3-H4)2 tetramers. These data suggest that CAF-1- and ASF1A-H3-H4-dependent deposition of the histone (H3-H4)2 tetramers is compatible with MMR and protects the discontinuous daughter strand from unnecessary degradation by MMR machinery. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Genome-wide mapping of histone H3K9me2 in acute myeloid leukemia reveals large chromosomal domains associated with massive gene silencing and sites of genome instability.

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    Anna C Salzberg

    Full Text Available A facultative heterochromatin mark, histone H3 lysine 9 dimethylation (H3K9me2, which is mediated by histone methyltransferases G9a/GLP (EHMT2/1, undergoes dramatic rearrangements during myeloid cell differentiation as observed by chromatin imaging. To determine whether these structural transitions also involve genomic repositioning of H3K9me2, we used ChIP-sequencing to map genome-wide topography of H3K9me2 in normal human granulocytes, normal CD34+ hematopoietic progenitors, primary myeloblasts from acute myeloid leukemia (AML patients, and a model leukemia cell line K562. We observe that H3K9me2 naturally repositions from the previously designated "repressed" chromatin state in hematopoietic progenitors to predominant association with heterochromatin regions in granulocytes. In contrast, AML cells accumulate H3K9me2 on previously undefined large (> 100 Kb genomic blocks that are enriched with AML-specific single nucleotide variants, sites of chromosomal translocations, and genes downregulated in AML. Specifically, the AML-specific H3K9me2 blocks are enriched with genes regulated by the proto-oncogene ERG that promotes stem cell characteristics. The AML-enriched H3K9me2 blocks (in contrast to the heterochromatin-associated H3K9me2 blocks enriched in granulocytes are reduced by pharmacological inhibition of the histone methyltransferase G9a/GLP in K562 cells concomitantly with transcriptional activation of ERG and ETS1 oncogenes. Our data suggest that G9a/GLP mediate formation of transient H3K9me2 blocks that are preserved in AML myeloblasts and may lead to an increased rate of AML-specific mutagenesis and chromosomal translocations.

  6. Altered global histone-trimethylation code and H3F3A-ATRX mutation in pediatric GBM.

    Science.gov (United States)

    Pathak, Pankaj; Jha, Prerana; Purkait, Suvendu; Sharma, Vikas; Suri, Vaishali; Sharma, Mehar C; Faruq, Mohammed; Suri, Ashish; Sarkar, Chitra

    2015-02-01

    Mutations in H3.3-ATRX-DAXX chromatin remodeling pathway have been reported in pediatric GBMs. H3.3 (H3F3A) mutations may affect transcriptional regulation by altered global histone-methylation. Therefore, we analyzed yet partly understood global histone code (H3K-4/9/27/36) trimethylation pattern in H3F3A-ATRX mutants and wild-type. H3F3A, HIST1H3B, IDH1, ATRX, DAXX and Tp53 mutations were identified by sequencing/immunohistochemistry in 27 pediatric GBMs. Global histone-methylation H3K-4/9/27/36me3 and Polycomb-protein EZH2 expression were evaluated by immunohistochemistry. H3F3A-ATRX mutation was observed in 66.7 % (18/27) of pediatric GBMs. K27M and G34R-H3F3A mutations were found in 37 % (10/27) and 14.8 % (4/27) patients respectively. G34V-H3F3A, HIST1H3B and IDH1 mutations were absent. Notably, commonest global histone-methylation mark lost was H3K27me3 (17/25, 68 %) followed by H3K4me3 (45.5 %, 10/22) and H3K9me3 (18.2 %, 4/22). Global H3K36me3 showed no loss. Most significant observation was loss of one or more histone-trimethylation mark in 80 % (20/25) pediatric GBMs. Notably, simultaneous loss of H3K27me3 and H3K4me3 were present in 7/22 (31.8 %) of pediatric GBMs. Low expression of EZH2 was found in 12/24 (50 %) of cases. However no significant correlation of loss of histone-marks or EZH2 expression with H3F3A-ATRX mutants (loss of at least one histone-marks in 87.5 % (14/16) cases) versus wild-types (loss of at least one histone-marks in 75 % (6/8) cases) was seen. The present study highlights for the first time combinatorial loss of one or more histone-trimethylation marks associated with majority of pediatric GBMs and the finding suggests significant role of histone-code in the molecular biology that underlies pediatric GBMs. Hence therapies for patients with particular combinations of histone modifications present opportunity to design innovative patient-tailored treatment protocols.

  7. Synthesis of ubiquitylated histone H3 using a thiirane linker for chemical ligation.

    Science.gov (United States)

    Kawakami, Toru; Mishima, Yuichi; Hojo, Hironobu; Suetake, Isao

    2017-07-01

    Post-translational modifications of histone proteins, which form nucleosome cores, play an important role in gene regulation. Ubiquitin modification is one such modification. We previously reported on the use of a thiirane linker to introduce a 1,2-aminothiol moiety at a cysteine residue for native chemical ligation with peptide thioesters, which permitted isopeptide mimetics to be produced. In this report, we describe the preparation of the ubiquitylated full length histone H3 at the 18 position and the construction of tetranucleosomes with recombinant histones H2A, H2B, H4, and DNA, which are slightly more stable than those that are prepared without ubiquitin modification. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.

  8. Context dependency of Set1/COMPASS-mediated histone H3 Lys4 trimethylation.

    Science.gov (United States)

    Thornton, Janet L; Westfield, Gerwin H; Takahashi, Yoh-Hei; Cook, Malcolm; Gao, Xin; Woodfin, Ashley R; Lee, Jung-Shin; Morgan, Marc A; Jackson, Jessica; Smith, Edwin R; Couture, Jean-Francois; Skiniotis, Georgios; Shilatifard, Ali

    2014-01-15

    The stimulation of trimethylation of histone H3 Lys4 (H3K4) by H2B monoubiquitination (H2Bub) has been widely studied, with multiple mechanisms having been proposed for this form of histone cross-talk. Cps35/Swd2 within COMPASS (complex of proteins associated with Set1) is considered to bridge these different processes. However, a truncated form of Set1 (762-Set1) is reported to function in H3K4 trimethylation (H3K4me3) without interacting with Cps35/Swd2, and such cross-talk is attributed to the n-SET domain of Set1 and its interaction with the Cps40/Spp1 subunit of COMPASS. Here, we used biochemical, structural, in vivo, and chromatin immunoprecipitation (ChIP) sequencing (ChIP-seq) approaches to demonstrate that Cps40/Spp1 and the n-SET domain of Set1 are required for the stability of Set1 and not the cross-talk. Furthermore, the apparent wild-type levels of H3K4me3 in the 762-Set1 strain are due to the rogue methylase activity of this mutant, resulting in the mislocalization of H3K4me3 from the promoter-proximal regions to the gene bodies and intergenic regions. We also performed detailed screens and identified yeast strains lacking H2Bub but containing intact H2Bub enzymes that have normal levels of H3K4me3, suggesting that monoubiquitination may not directly stimulate COMPASS but rather works in the context of the PAF and Rad6/Bre1 complexes. Our study demonstrates that the monoubiquitination machinery and Cps35/Swd2 function to focus COMPASS's H3K4me3 activity at promoter-proximal regions in a context-dependent manner.

  9. The PR-Set7 binding domain of Riz1 is required for the H4K20me1-H3K9me1 trans-tail 'histone code' and Riz1 tumor suppressor function.

    Science.gov (United States)

    Congdon, Lauren M; Sims, Jennifer K; Tuzon, Creighton T; Rice, Judd C

    2014-04-01

    PR-Set7/Set8/KMT5a is the sole histone H4 lysine 20 monomethyltransferase (H4K20me1) in metazoans and is essential for proper cell division and genomic stability. We unexpectedly discovered that normal cellular levels of monomethylated histone H3 lysine 9 (H3K9me1) were also dependent on PR-Set7, but independent of its catalytic activity. This observation suggested that PR-Set7 interacts with an H3K9 monomethyltransferase to establish the previously reported H4K20me1-H3K9me1 trans-tail 'histone code'. Here we show that PR-Set7 specifically and directly binds the C-terminus of the Riz1/PRDM2/KMT8 tumor suppressor and demonstrate that the N-terminal PR/SET domain of Riz1 preferentially monomethylates H3K9. The PR-Set7 binding domain was required for Riz1 nuclear localization and maintenance of the H4K20me1-H3K9me1 trans-tail 'histone code'. Although Riz1 can function as a repressor, Riz1/H3K9me1 was dispensable for the repression of genes regulated by PR-Set7/H4K20me1. Frameshift mutations resulting in a truncated Riz1 incapable of binding PR-Set7 occur frequently in various aggressive cancers. In these cancer cells, expression of wild-type Riz1 restored tumor suppression by decreasing proliferation and increasing apoptosis. These phenotypes were not observed in cells expressing either the Riz1 PR/SET domain or PR-Set7 binding domain indicating that Riz1 methyltransferase activity and PR-Set7 binding domain are both essential for Riz1 tumor suppressor function.

  10. Systematic screen reveals new functional dynamics of histones H3 and H4 during gametogenesis.

    Science.gov (United States)

    Govin, Jérôme; Dorsey, Jean; Gaucher, Jonathan; Rousseaux, Sophie; Khochbin, Saadi; Berger, Shelley L

    2010-08-15

    Profound epigenetic differences exist between genomes derived from male and female gametes; however, the nature of these changes remains largely unknown. We undertook a systematic investigation of chromatin reorganization during gametogenesis, using the model eukaryote Saccharomyces cerevisiae to examine sporulation, which has strong similarities with higher eukaryotic spermatogenesis. We established a mutational screen of histones H3 and H4 to uncover substitutions that reduce sporulation efficiency. We discovered two patches of residues-one on H3 and a second on H4-that are crucial for sporulation but not critical for mitotic growth, and likely comprise interactive nucleosomal surfaces. Furthermore, we identified novel histone post-translational modifications that mark the chromatin reorganization process during sporulation. First, phosphorylation of H3T11 appears to be a key modification during meiosis, and requires the meiotic-specific kinase Mek1. Second, H4 undergoes amino tail acetylation at Lys 5, Lys 8, and Lys 12, and these are synergistically important for post-meiotic chromatin compaction, occurring subsequent to the post-meiotic transient peak in phosphorylation at H4S1, and crucial for recruitment of Bdf1, a bromodomain protein, to chromatin in mature spores. Strikingly, the presence and temporal succession of the new H3 and H4 modifications are detected during mouse spermatogenesis, indicating that they are conserved through evolution. Thus, our results show that investigation of gametogenesis in yeast provides novel insights into chromatin dynamics, which are potentially relevant to epigenetic modulation of the mammalian process.

  11. Drosophila Kismet regulates histone H3 lysine 27 methylation and early elongation by RNA polymerase II.

    Directory of Open Access Journals (Sweden)

    Shrividhya Srinivasan

    2008-10-01

    Full Text Available Polycomb and trithorax group proteins regulate cellular pluripotency and differentiation by maintaining hereditable states of transcription. Many Polycomb and trithorax group proteins have been implicated in the covalent modification or remodeling of chromatin, but how they interact with each other and the general transcription machinery to regulate transcription is not well understood. The trithorax group protein Kismet-L (KIS-L is a member of the CHD subfamily of chromatin-remodeling factors that plays a global role in transcription by RNA polymerase II (Pol II. Mutations in CHD7, the human counterpart of kis, are associated with CHARGE syndrome, a developmental disorder affecting multiple tissues and organs. To clarify how KIS-L activates gene expression and counteracts Polycomb group silencing, we characterized defects resulting from the loss of KIS-L function in Drosophila. These studies revealed that KIS-L acts downstream of P-TEFb recruitment to stimulate elongation by Pol II. The presence of two chromodomains in KIS-L suggested that its recruitment or function might be regulated by the methylation of histone H3 lysine 4 by the trithorax group proteins ASH1 and TRX. Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin. By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression. A similar increase in H3 lysine 27 methylation was observed in ash1 and trx mutant larvae. Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

  12. Systems Level Analysis of Histone H3 Post-translational Modifications (PTMs) Reveals Features of PTM Crosstalk in Chromatin Regulation*

    Science.gov (United States)

    Schwämmle, Veit; Sidoli, Simone; Ruminowicz, Chrystian; Wu, Xudong; Lee, Chung-Fan; Helin, Kristian; Jensen, Ole N.

    2016-01-01

    Histones are abundant chromatin constituents carrying numerous post-translational modifications (PTMs). Such PTMs mediate a variety of biological functions, including recruitment of enzymatic readers, writers and erasers that modulate DNA replication, transcription and repair. Individual histone molecules contain multiple coexisting PTMs, some of which exhibit crosstalk, i.e. coordinated or mutually exclusive activities. Here, we present an integrated experimental and computational systems level molecular characterization of histone PTMs and PTM crosstalk. Using wild type and engineered mouse embryonic stem cells (mESCs) knocked out in components of the Polycomb Repressive Complex 2 (PRC2, Suz12−/−), PRC1 (Ring1A/B−/−) and (Dnmt1/3a/3b−/−) we performed comprehensive PTM analysis of histone H3 tails (50 aa) by utilizing quantitative middle-down proteome analysis by tandem mass spectrometry. We characterized combinatorial PTM features across the four mESC lines and then applied statistical data analysis to predict crosstalk between histone H3 PTMs. We detected an overrepresentation of positive crosstalk (codependent marks) between adjacent mono-methylated and acetylated marks, and negative crosstalk (mutually exclusive marks) among most of the seven characterized di- and tri-methylated lysine residues in the H3 tails. We report novel features of PTM interplay involving hitherto poorly characterized arginine methylation and lysine methylation sites, including H3R2me, H3R8me and H3K37me. Integration of the H3 data with RNAseq data by coabundance clustering analysis of histone PTMs and histone modifying enzymes revealed correlations between PTM and enzyme levels. We conclude that middle-down proteomics is a powerful tool to determine conserved or dynamic interdependencies between histone marks, which paves the way for detailed investigations of the histone code. Histone H3 PTM data is publicly available in the CrossTalkDB repository at http

  13. Systems Level Analysis of Histone H3 Post-translational Modifications (PTMs) Reveals Features of PTM Crosstalk in Chromatin Regulation.

    Science.gov (United States)

    Schwämmle, Veit; Sidoli, Simone; Ruminowicz, Chrystian; Wu, Xudong; Lee, Chung-Fan; Helin, Kristian; Jensen, Ole N

    2016-08-01

    Histones are abundant chromatin constituents carrying numerous post-translational modifications (PTMs). Such PTMs mediate a variety of biological functions, including recruitment of enzymatic readers, writers and erasers that modulate DNA replication, transcription and repair. Individual histone molecules contain multiple coexisting PTMs, some of which exhibit crosstalk, i.e. coordinated or mutually exclusive activities. Here, we present an integrated experimental and computational systems level molecular characterization of histone PTMs and PTM crosstalk. Using wild type and engineered mouse embryonic stem cells (mESCs) knocked out in components of the Polycomb Repressive Complex 2 (PRC2, Suz12(-/-)), PRC1 (Ring1A/B(-/-)) and (Dnmt1/3a/3b(-/-)) we performed comprehensive PTM analysis of histone H3 tails (50 aa) by utilizing quantitative middle-down proteome analysis by tandem mass spectrometry. We characterized combinatorial PTM features across the four mESC lines and then applied statistical data analysis to predict crosstalk between histone H3 PTMs. We detected an overrepresentation of positive crosstalk (codependent marks) between adjacent mono-methylated and acetylated marks, and negative crosstalk (mutually exclusive marks) among most of the seven characterized di- and tri-methylated lysine residues in the H3 tails. We report novel features of PTM interplay involving hitherto poorly characterized arginine methylation and lysine methylation sites, including H3R2me, H3R8me and H3K37me. Integration of the H3 data with RNAseq data by coabundance clustering analysis of histone PTMs and histone modifying enzymes revealed correlations between PTM and enzyme levels. We conclude that middle-down proteomics is a powerful tool to determine conserved or dynamic interdependencies between histone marks, which paves the way for detailed investigations of the histone code. Histone H3 PTM data is publicly available in the CrossTalkDB repository at http

  14. KMT2D maintains neoplastic cell proliferation and global histone H3 lysine 4 monomethylation.

    Science.gov (United States)

    Guo, Changcun; Chen, Lee H; Huang, Yafen; Chang, Chun-Chi; Wang, Ping; Pirozzi, Christopher J; Qin, Xiaoxia; Bao, Xuhui; Greer, Paula K; McLendon, Roger E; Yan, Hai; Keir, Stephen T; Bigner, Darell D; He, Yiping

    2013-11-01

    KMT2D (lysine (K)-specific methyltransferase 2D), formerly named MLL2 (myeloid/lymphoid or mixed-lineage leukemia 2, also known as ALR/MLL4), is a histone methyltransferase that plays an important role in regulating gene transcription. In particular, it targets histone H3 lysine 4 (H3K4), whose methylations serve as a gene activation mark. Recently, KMT2D has emerged as one of the most frequently mutated genes in a variety of cancers and in other human diseases, including lymphoma, medulloblastoma, gastric cancer, and Kabuki syndrome. Mutations in KMT2D identified thus far point to its loss-of-function in pathogenesis and suggest its role as a tumor suppressor in various tissues. To determine the effect of a KMT2D deficiency on neoplastic cells, we used homologous recombination- and nuclease-mediated gene editing approaches to generate a panel of isogenic colorectal and medulloblastoma cancer cell lines that differ with respect to their endogenous KMT2D status. We found that a KMT2D deficiency resulted in attenuated cancer cell proliferation and defective cell migration. Analysis of histone H3 modifications revealed that KMT2D was essential for maintaining the level of global H3K4 monomethylation and that its enzymatic SET domain was directly responsible for this function. Furthermore, we found that a majority of KMT2D binding sites are located in regions of potential enhancer elements. Together, these findings revealed the role of KMT2D in regulating enhancer elements in human cells and shed light on the tumorigenic role of its deficiency. Our study supports that KMT2D has distinct roles in neoplastic cells, as opposed to normal cells, and that inhibiting KMT2D may be a viable strategy for cancer therapeutics.

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

    Science.gov (United States)

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

    2016-01-01

    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 1H-15N 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. PMID:26673727

  16. Combinatorial readout of histone H3 modifications specifies localization of ATRX to heterochromatin.

    Science.gov (United States)

    Eustermann, Sebastian; Yang, Ji-Chun; Law, Martin J; Amos, Rachel; Chapman, Lynda M; Jelinska, Clare; Garrick, David; Clynes, David; Gibbons, Richard J; Rhodes, Daniela; Higgs, Douglas R; Neuhaus, David

    2011-06-12

    Accurate read-out of chromatin modifications is essential for eukaryotic life. Mutations in the gene encoding X-linked ATRX protein cause a mental-retardation syndrome, whereas wild-type ATRX protein targets pericentric and telomeric heterochromatin for deposition of the histone variant H3.3 by means of a largely unknown mechanism. Here we show that the ADD domain of ATRX, in which most syndrome-causing mutations occur, engages the N-terminal tail of histone H3 through two rigidly oriented binding pockets, one for unmodified Lys4 and the other for di- or trimethylated Lys9. In vivo experiments show this combinatorial readout is required for ATRX localization, with recruitment enhanced by a third interaction through heterochromatin protein-1 (HP1) that also recognizes trimethylated Lys9. The cooperation of ATRX ADD domain and HP1 in chromatin recruitment results in a tripartite interaction that may span neighboring nucleosomes and illustrates how the 'histone-code' is interpreted by a combination of multivalent effector-chromatin interactions.

  17. Identification of lysine K18 acetylation on histone H3 peptide using gold nanoparticles' aggregation behaviour.

    Science.gov (United States)

    Li, Ning; Sutarlie, Laura; Lew, Qiao Jing; Chao, Sheng-Hao; Su, Xiaodi

    2016-04-01

    Acetylation of histones, the major protein component of eukaryotic chromosomes, contributes to the epigenetic regulation of gene expression and is also involved in cancer development. A recent study revealed the correlation between tumour formation and acetylation level of lysine K18 on histone H3. In this study, we developed two colorimetric in vitro assays using gold nanoparticles (AuNPs) for identification of lysine K18 acetylation on histone H3 peptide. In assay I, citrate ion-capped AuNP without further modification was employed. Simply mixing the K18 peptide with AuNP solution leads to distinct particle aggregation, relative to that by non-acetylated or lysine K14 acetylated control peptides. In assay II, an AuNP-peptide-antibody composite was synthesized and used as both the sensing probe and the transducing element. By mixing the sample peptides with the composite solution followed by PBS screening, different aggregation behaviours were observed between the K18 acetylated target peptide and the control sequences. Both assays are capable of identifying the acetylated peptides, and also differentiating the distinctive acetylation positions that differ merely by a distance of three amino acids.

  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. Distinct Roles of Histone H3 and H2A Tails in Nucleosome Stability

    Science.gov (United States)

    Li, Zhenhai; Kono, Hidetoshi

    2016-01-01

    Nucleosome breathing potentially increases the DNA exposure, which in turn recruits DNA-binding protein and regulates gene transcription. Numerous studies have shown the critical roles of N-terminal tails of histones H3 and H4 in gene expression; however, few studies have focused on the H2A C-terminal tail. Here we present thorough computational studies on a single nucleosome particle showing the linker DNA closing and opening, which is thought to be nucleosome breathing. With our simulation, the H2A C-terminal and H3 N-terminal tails were found to modulate the nucleosome conformation differently. The H2A C-terminal tail regulates nucleosome conformation by binding to linker DNA at different locations, whereas the H3 N-terminal tail regulates linker DNA by binding to it in different patterns. Further MD simulation on tail truncated structures corroborates this analysis. These findings replenish our understanding of the histone tail regulation mechanism on atomic level. PMID:27527579

  20. The CHD3 Remodeler PICKLE Promotes Trimethylation of Histone H3 Lysine 27*S⃞

    Science.gov (United States)

    Zhang, Heng; Rider, Stanley Dean; Henderson, James T.; Fountain, Matthew; Chuang, King; Kandachar, Vasundhara; Simons, Alexis; Edenberg, Howard J.; Romero-Severson, Jeanne; Muir, William M.; Ogas, Joe

    2008-01-01

    CHD3 proteins are ATP-dependent chromatin remodelers that contribute to repression of developmentally regulated genes in both animal and plant systems. In animals, this repression has been linked to a multiple subunit complex, Mi-2/NuRD, whose constituents include a CHD3 protein, a histone deacetylase, and a methyl-CpG-binding domain protein. In Arabidopsis, PICKLE (PKL) codes for a CHD3 protein that acts during germination to repress expression of seed-associated genes. Repression of seed-associated traits is promoted in pkl seedlings by the plant growth regulator gibberellin (GA). We undertook a microarray analysis to determine how PKL and GA act to promote the transition from seed to seedling. We found that PKL and GA act in separate pathways to repress expression of seed-specific genes. Comparison of genomic datasets revealed that PKL-dependent genes are enriched for trimethylation of histone H3 lysine 27 (H3K27me3), a repressive epigenetic mark. Chromatin immunoprecipitation studies demonstrate that PKL promotes H3K27me3 in both germinating seedlings and in adult plants but do not identify a connection between PKL-dependent expression and acetylation levels. Taken together, our analyses illuminate a new pathway by which CHD3 remodelers contribute to repression in eukaryotes. PMID:18539592

  1. Analysis of Histones H3 and H4 Reveals Novel and Conserved Post-Translational Modifications in Sugarcane.

    Science.gov (United States)

    Moraes, Izabel; Yuan, Zuo-Fei; Liu, Shichong; Souza, Glaucia Mendes; Garcia, Benjamin A; Casas-Mollano, J Armando

    2015-01-01

    Histones are the main structural components of the nucleosome, hence targets of many regulatory proteins that mediate processes involving changes in chromatin. The functional outcome of many pathways is "written" in the histones in the form of post-translational modifications that determine the final gene expression readout. As a result, modifications, alone or in combination, are important determinants of chromatin states. Histone modifications are accomplished by the addition of different chemical groups such as methyl, acetyl and phosphate. Thus, identifying and characterizing these modifications and the proteins related to them is the initial step to understanding the mechanisms of gene regulation and in the future may even provide tools for breeding programs. Several studies over the past years have contributed to increase our knowledge of epigenetic gene regulation in model organisms like Arabidopsis, yet this field remains relatively unexplored in crops. In this study we identified and initially characterized histones H3 and H4 in the monocot crop sugarcane. We discovered a number of histone genes by searching the sugarcane ESTs database. The proteins encoded correspond to canonical histones, and their variants. We also purified bulk histones and used them to map post-translational modifications in the histones H3 and H4 using mass spectrometry. Several modifications conserved in other plants, and also novel modified residues, were identified. In particular, we report O-acetylation of serine, threonine and tyrosine, a recently identified modification conserved in several eukaryotes. Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species. To our knowledge, this is the first report of histones H3 and H4 as well as their post-translational modifications in sugarcane, and will provide a starting point for the study of chromatin regulation in

  2. PITX2 associates with PTIP-containing histone H3 lysine 4 methyltransferase complex.

    Science.gov (United States)

    Liu, Yan; Huang, Yue; Fan, Jun; Zhu, Guo-Zhang

    2014-02-21

    Pituitary homeobox 2 (PITX2), a Paired-like homeodomain transcription factor and a downstream effector of Wnt/β-catenin signaling, plays substantial roles in embryonic development and human disorders. The mechanism of its functions, however, is not fully understood. In this study, we demonstrated that PITX2 associated with histone H3 lysine 4 (H3K4) methyltransferase (HKMT) mixed-lineage leukemia 4 (MLL4/KMT2D), Pax transactivation domain-interacting protein (PTIP), and other H3K4·HKMT core subunits. This association of PITX2 with H3K4·HKMT complex was dependent on PITX2's homeodomain. Consistently, the PITX2 protein complex was shown to possess H3K4·HKMT activity. Furthermore, the chromatin immunoprecipitation result revealed co-occupancy of PITX2 and PTIP on the promoter of the PITX2's transcriptional target. Taken together, our data provide new mechanistic perspectives on PITX2's functions and its related biological processes. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Three Distinct Patterns of Histone H3Y41 Phosphorylation Mark Active Genes

    Directory of Open Access Journals (Sweden)

    Mark A. Dawson

    2012-09-01

    Full Text Available The JAK2 tyrosine kinase is a critical mediator of cytokine-induced signaling. It plays a role in the nucleus, where it regulates transcription by phosphorylating histone H3 at tyrosine 41 (H3Y41ph. We used chromatin immunoprecipitation coupled to massively parallel DNA sequencing (ChIP-seq to define the genome-wide pattern of H3Y41ph in human erythroid leukemia cells. Our results indicate that H3Y41ph is located at three distinct sites: (1 at a subset of active promoters, where it overlaps with H3K4me3, (2 at distal cis-regulatory elements, where it coincides with the binding of STAT5, and (3 throughout the transcribed regions of active, tissue-specific hematopoietic genes. Together, these data extend our understanding of this conserved and essential signaling pathway and provide insight into the mechanisms by which extracellular stimuli may lead to the coordinated regulation of transcription.

  4. Transition state for the NSD2-catalyzed methylation of histone H3 lysine 36.

    Science.gov (United States)

    Poulin, Myles B; Schneck, Jessica L; Matico, Rosalie E; McDevitt, Patrick J; Huddleston, Michael J; Hou, Wangfang; Johnson, Neil W; Thrall, Sara H; Meek, Thomas D; Schramm, Vern L

    2016-02-02

    Nuclear receptor SET domain containing protein 2 (NSD2) catalyzes the methylation of histone H3 lysine 36 (H3K36). It is a determinant in Wolf-Hirschhorn syndrome and is overexpressed in human multiple myeloma. Despite the relevance of NSD2 to cancer, there are no potent, selective inhibitors of this enzyme reported. Here, a combination of kinetic isotope effect measurements and quantum chemical modeling was used to provide subangstrom details of the transition state structure for NSD2 enzymatic activity. Kinetic isotope effects were measured for the methylation of isolated HeLa cell nucleosomes by NSD2. NSD2 preferentially catalyzes the dimethylation of H3K36 along with a reduced preference for H3K36 monomethylation. Primary Me-(14)C and (36)S and secondary Me-(3)H3, Me-(2)H3, 5'-(14)C, and 5'-(3)H2 kinetic isotope effects were measured for the methylation of H3K36 using specifically labeled S-adenosyl-l-methionine. The intrinsic kinetic isotope effects were used as boundary constraints for quantum mechanical calculations for the NSD2 transition state. The experimental and calculated kinetic isotope effects are consistent with an SN2 chemical mechanism with methyl transfer as the first irreversible chemical step in the reaction mechanism. The transition state is a late, asymmetric nucleophilic displacement with bond separation from the leaving group at (2.53 Å) and bond making to the attacking nucleophile (2.10 Å) advanced at the transition state. The transition state structure can be represented in a molecular electrostatic potential map to guide the design of inhibitors that mimic the transition state geometry and charge.

  5. Reduced Histone H3 Acetylation in CD4+ T Lymphocytes: Potential Mechanism of Latent Autoimmune Diabetes in Adults

    Directory of Open Access Journals (Sweden)

    Xi-yu Liu

    2015-01-01

    Full Text Available Aims. Latent autoimmune diabetes in adults (LADA is the result of gene-environment interactions. Histone acetylation regulates gene expression and maybe interpret how environmental factors modify LADA. Hence, we studied the histone acetylation patterns in CD4+ T lymphocytes from LADA patients. Methods. Blood CD4+ T lymphocytes from 28 patients with LADA and 28 healthy controls were obtained to detect histone H3 acetylation and H4 acetylation. The gene expression of histone acetyltransferases (P300 and CREBBP and histone deacetylases (HDAC1, HDAC2, and HDAC7 was measured by real-time polymerase chain reaction (RT-PCR. Results. Compared to healthy controls, reduced global H3 acetylation was observed in LADA patients’ CD4+ T lymphocytes (P<0.05. Global level of H4 acetylation was not statistically different. Among LADA, CD4+ T lymphocytes H3 acetylation was associated with glycosylated hemoglobin (HbA1c and GADA titer. Compared to healthy controls, the expression of histone acetyltransferases CREBBP in LADA patients was downregulated, and the expression of histone deacetylases HDAC1 and HDAC7 was upregulated. Conclusion. A concerted downregulation of histone H3 acetylation was found in CD4+ T lymphocytes of LADA patients, and this might provide evidence of a novel epigenetic explanation for the pathogenesis of LADA and its complications.

  6. Immunostaining analysis of tissue cultured cells and tissue sections using phospho-Histone H3 (Serine 10) antibody.

    Science.gov (United States)

    Padmanabhan, Jaya

    2015-01-01

    Post-translational modifications of histones play an important role in regulation of gene expression through condensation and decondensation of chromatin structure. These modifications include acetylation, methylation, phosphorylation and ubiquitination. Phosphorylation on histones is associated with cellular responses such as DNA damage, transcription, chromatin compaction and mitosis or meiosis. One of the most extensively studied modifications of histones is the Serine 10 phosphorylation on histone H3 N-terminal tail. This specific phosphorylation on Histone H3 has been associated with condensation and transcriptional inactivation of mitotic chromosomes, but recent studies have suggested a role for this specific phosphorylation in chromatin relaxation and activation of transcription in interphase cells. Co-immunostaining analysis of cells using antibodies specific to serine 10P-Histone H3 together with those to cell cycle specific markers will allow us to determine the nature of phosphorylation in a cell cycle-specific manner. In a complex system, such as tissue specimens, analysis using P-Histone H3 and a cell type specific antibody will allow identification of specific cells that are affected by this histone modification. This is of particular interest in the field of cancer biology or neurobiology where identification or quantification of the transcriptionally active or mitotic cells will enable one to evaluate the progression of the disease development. The protocol described here provides details on how co-immunostaining and analysis can be performed in tissue cultured cells or tissue sections.

  7. The COMPASS Family of Histone H3K4 Methylases: Mechanisms of Regulation in Development and Disease Pathogenesis

    OpenAIRE

    Shilatifard, Ali

    2012-01-01

    The Saccharomyces cerevisiae Set1/COMPASS was the first histone H3 lysine 4 (H3K4) methylase identified over ten years ago. Since then, it has been demonstrated that Set1/COMPASS and its enzymatic product, H3K4 methylation, is highly conserved across the evolutionary tree. Although there is only one COMPASS in yeast, human cells bear at least six COMPASS family members each capable of methylating H3K4 with non-redundant functions. In yeast, the monoubiquitination of histone H2B by Rad6/Bre1 i...

  8. Overlapping regulation of CenH3 localization and histone H3 turnover by CAF-1 and HIR proteins in Saccharomyces cerevisiae.

    Science.gov (United States)

    Lopes da Rosa, Jessica; Holik, John; Green, Erin M; Rando, Oliver J; Kaufman, Paul D

    2011-01-01

    Accurate chromosome segregation is dependent on the centromere-specific histone H3 isoform known generally as CenH3, or as Cse4 in budding yeast. Cytological experiments have shown that Cse4 appears at extracentromeric loci in yeast cells deficient for both the CAF-1 and HIR histone H3/H4 deposition complexes, consistent with increased nondisjunction in these double mutant cells. Here, we examined molecular aspects of this Cse4 mislocalization. Genome-scale chromatin immunoprecipitation analyses demonstrated broader distribution of Cse4 outside of centromeres in cac1Δ hir1Δ double mutant cells that lack both CAF-1 and HIR complexes than in either single mutant. However, cytological localization showed that the essential inner kinetochore component Mif2 (CENP-C) was not recruited to extracentromeric Cse4 in cac1Δ hir1Δ double mutant cells. We also observed that rpb1-1 mutants displayed a modestly increased Cse4 half-life at nonpermissive temperatures, suggesting that turnover of Cse4 is partially dependent on Pol II transcription. We used genome-scale assays to demonstrate that the CAF-1 and HIR complexes independently stimulate replication-independent histone H3 turnover rates. We discuss ways in which altered histone exchange kinetics may affect eviction of Cse4 from noncentromeric loci.

  9. The Histone H3K27 Demethylase UTX Regulates Synaptic Plasticity and Cognitive Behaviors in Mice

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    Gang-Bin Tang

    2017-08-01

    Full Text Available Histone demethylase UTX mediates removal of repressive trimethylation of histone H3 lysine 27 (H3K27me3 to establish a mechanistic switch to activate large sets of genes. Mutation of Utx has recently been shown to be associated with Kabuki syndrome, a rare congenital anomaly syndrome with dementia. However, its biological function in the brain is largely unknown. Here, we observe that deletion of Utx results in increased anxiety-like behaviors and impaired spatial learning and memory in mice. Loss of Utx in the hippocampus leads to reduced long-term potentiation and amplitude of miniature excitatory postsynaptic current, aberrant dendrite development and defective synapse formation. Transcriptional profiling reveals that Utx regulates a subset of genes that are involved in the regulation of dendritic morphology, synaptic transmission, and cognition. Specifically, Utx deletion disrupts expression of neurotransmitter 5-hydroxytryptamine receptor 5B (Htr5b. Restoration of Htr5b expression in newborn hippocampal neurons rescues the defects of neuronal morphology by Utx ablation. Therefore, we provide evidence that Utx plays a critical role in modulating synaptic transmission and cognitive behaviors. Utx cKO mouse models like ours provide a valuable means to study the underlying mechanisms of the etiology of Kabuki syndrome.

  10. H3K23me1 is an evolutionarily conserved histone modification associated with CG DNA methylation in Arabidopsis.

    Science.gov (United States)

    Trejo-Arellano, Minerva S; Mahrez, Walid; Nakamura, Miyuki; Moreno-Romero, Jordi; Nanni, Paolo; Köhler, Claudia; Hennig, Lars

    2017-04-01

    Amino-terminal tails of histones are targets for diverse post-translational modifications whose combinatorial action may constitute a code that will be read and interpreted by cellular proteins to define particular transcriptional states. Here, we describe monomethylation of histone H3 lysine 23 (H3K23me1) as a histone modification not previously described in plants. H3K23me1 is an evolutionarily conserved mark in diverse species of flowering plants. Chromatin immunoprecipitation followed by high-throughput sequencing in Arabidopsis thaliana showed that H3K23me1 was highly enriched in pericentromeric regions and depleted from chromosome arms. In transposable elements it co-localized with CG, CHG and CHH DNA methylation as well as with the heterochromatic histone mark H3K9me2. Transposable elements are often rich in H3K23me1 but different families vary in their enrichment: LTR-Gypsy elements are most enriched and RC/Helitron elements are least enriched. The histone methyltransferase KRYPTONITE and normal DNA methylation were required for normal levels of H3K23me1 on transposable elements. Immunostaining experiments confirmed the pericentromeric localization and also showed mild enrichment in less condensed regions. Accordingly, gene bodies of protein-coding genes had intermediate H3K23me1 levels, which coexisted with CG DNA methylation. Enrichment of H3K23me1 along gene bodies did not correlate with transcription levels. Together, this work establishes H3K23me1 as a so far undescribed component of the plant histone code. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  11. Site-specific Acetylation of Histone H3 Decreases Polymerase β Activity on Nucleosome Core Particles in Vitro.

    Science.gov (United States)

    Rodriguez, Yesenia; Hinz, John M; Laughery, Marian F; Wyrick, John J; Smerdon, Michael J

    2016-05-20

    Histone posttranslational modifications have been associated with changes in chromatin structure necessary for transcription, replication, and DNA repair. Acetylation is one of the most studied and best characterized histone posttranslational modifications, but it is not known if histone acetylation modulates base excision repair of DNA lesions in chromatin. To address this question, we generated nucleosome core particles (NCPs) containing site-specifically acetylated H3K14 or H3K56 and measured repair of uracil and single-nucleotide gaps. We find that H3K56Ac and H3K14Ac do not significantly contribute to removal of uracils by uracil DNA glycosylase regardless of the translational or rotational position of the lesions within NCPs. In repair of single-nucleotide gaps, however, the presence of H3K56Ac or H3K14Ac in NCPs decreases the gap-filling activity of DNA polymerase β near the dyad center, with H3K14Ac exhibiting stronger inhibition. To a lesser extent, H3K56Ac induces a similar effect near the DNA ends. Moreover, using restriction enzyme accessibility, we detect no changes in NCP structure or dynamics between H3K14Ac-NCPs and WT-NCPs containing single-nucleotide gaps. Thus, acetylation at H3K56 and H3K14 in nucleosomes may promote alternative gap-filling pathways by inhibiting DNA polymerase β activity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Dynamics of a novel centromeric histone variant CenH3 reveals the evolutionary ancestral timing of centromere biogenesis.

    Science.gov (United States)

    Dubin, Manu; Fuchs, Jörg; Gräf, Ralph; Schubert, Ingo; Nellen, Wolfgang

    2010-11-01

    The centromeric histone H3 variant (CenH3) serves to target the kinetochore to the centromeres and thus ensures correct chromosome segregation during mitosis and meiosis. The Dictyostelium H3-like variant H3v1 was identified as the CenH3 ortholog. Dictyostelium CenH3 has an extended N-terminal domain with no similarity to any other known proteins and a histone fold domain at its C-terminus. Within the histone fold, α-helix 2 (α2) and an extended loop 1 (L1) have been shown to be required for targeting CenH3 to centromeres. Compared to other known and putative CenH3 histones, Dictyostelium CenH3 has a shorter L1, suggesting that the extension is not an obligatory feature. Through ChIP analysis and fluorescence microscopy of live and fixed cells, we provide here the first survey of centromere structure in amoebozoa. The six telocentric centromeres were found to mostly consist of all the DIRS-1 elements and to associate with H3K9me3. During interphase, the centromeres remain attached to the centrosome forming a single CenH3-containing cluster. Loading of Dictyostelium CenH3 onto centromeres occurs at the G2/prophase transition, in contrast to the anaphase/telophase loading of CenH3 observed in metazoans. This suggests that loading during G2/prophase is the ancestral eukaryotic mechanism and that anaphase/telophase loading of CenH3 has evolved more recently after the amoebozoa diverged from the animal linage.

  13. Dynamic changes of histone H3 marks during Caenorhabditis elegans lifecycle revealed by middle-down proteomics

    DEFF Research Database (Denmark)

    Sidoli, Simone; Vandamme, Julien; Elisabetta Salcini, Anna

    2016-01-01

    that is transmitted during dauer formation. Collectively, our data describe the dynamics of histone H3 combinatorial code during C. elegans lifecycle and demonstrate the feasibility of using middle-down proteomics to study in vivo development of multicellular organisms. This article is protected by copyright. All......We applied a middle-down proteomics strategy for large scale protein analysis during in vivo development of Caenorhabditis elegans. We characterized post-translational modifications (PTMs) on histone H3 N-terminal tails at eight time points during the C. elegans lifecycle, including embryo, larval...... stages (L1 to L4), dauer and L1/L4 post dauer. Histones were analyzed by our optimized middle-down protein sequencing platform using high mass accuracy tandem mass spectrometry. This allows quantification of intact histone tails and detailed characterization of distinct histone tails carrying co...

  14. Steroid receptor coactivator-3 is a pivotal target of gambogic acid in B-cell Non-Hodgkin lymphoma and an inducer of histone H3 deacetylation.

    Science.gov (United States)

    Zhao, Zichu; Zhang, Xia; Wen, Lu; Yi, Sha; Hu, Jingyu; Ruan, Jun; Zhao, Fei; Cui, Guohui; Fang, Jun; Chen, Yan

    2016-10-15

    Gambogic acid (GA), the active ingredient from gamboges, has been verified as a potent anti-tumor agent in many cancer cells. Nevertheless, its function in lymphoma, especially in B-cell Non-Hodgkin lymphoma (NHL), remains unclear. Amplification and/or overexpression of steroid receptor coactivator-3 (SRC-3) have been detected in multiple tumors and have confirmed its critical roles in carcinogenesis, progression, metastasis and therapy resistance in these cancers. However, no clinical data have revealed the overexpression of SRC-3 and its role in B-cell NHL. In this study, we demonstrated the anti-tumor effects of GA, which included cell growth inhibition, G1/S phase cell cycle arrest and apoptosis in B-cell NHL. We also verified that SRC-3 was overexpressed in B-cell NHL in both cell lines and lymph node samples from patients. The overexpressed SRC-3 was a central drug target of GA, and its down-regulation subsequently modulated down-stream gene expression, ultimately contributing to apoptosis. Silencing SRC-3 decreased the expression of Bcl-2, Bcl-6 and cyclin D3, but not of NF-κB and IκB-α. GA treatment did not inhibit the activation of AKT signaling pathway, but induced the deacetylation of histone H3 at lysine 9 and lysine 27. Down-regulated SRC-3 was observed to interact with more HDAC1 to mediate the deacetylation of H3. As the component of E3 ligase, Cullin3 was up-regulated and mediated the degradation of SRC-3. Our results demonstrate that GA is a potent anti-tumor agent that can be used for therapy against B-cell NHL, especially against those with an abundance of SRC-3. Copyright © 2016. Published by Elsevier B.V.

  15. [Correlation between histone H3-K9 methylation, DNA methylation and expression of gene MGMT in Hep-2 cell line].

    Science.gov (United States)

    Yang, Jing; He, Liria; Ji, Wenyue; Jin, Mingzhu; Zhao, Xudong

    2012-11-01

    To explore the correlation between histone H3-K9 methylation, DNA methylation and expression of carcinoma suppressor gene MGMT in laryngeal carcinoma Hep-2 cell line. 5-Aza-dC was used to deal with Hep-2 cell cultured in vitro. ChIP, MSP and Realtime-PCR were used to detect H3-K9 methylation, DNA methylation, of MGMT gene promoter region and MGMT gene expression before and after treatment with drugs. (1) In Hep-2 cell line, gene MGMT was characterized by DNA methylation and histone H3-K9 hypermethylation. (2) 5-Aza-dC was able to reduce H3-K9 methylation of MGMT gene histone in Hep-2 cell line, 5-Aza-dC was able to reverse DNA methylation of MGMT gene histone in Hep-2 cell line, 5-Aza-dC was able to upregulate the down-regulated gene expression of tumor suppressor genes MGMT. Promoter methylation of cancer suppressor gene MGMT may induce the gene inactivity. DNA methylation may increase H3-K9 methylation. 5-Aza-dC can reduce H3-K9 methylation of tumor suppressor gene MGMT histone by reversing DNA methylation of tumor suppressor gene MGMT, and then the expression of tumor suppressor genes is increased and tumor development is inhibited.

  16. Expression and significance of histone H3K27 demethylases in renal cell carcinoma

    Directory of Open Access Journals (Sweden)

    Shen Yongqing

    2012-10-01

    Full Text Available Abstract Background The histone H3K27 demethylases UTX and JMJD3 are important regulatory factors that modulate gene expression by altering the physical state of chromatin. Previous studies have indicated an abnormal H3K27 methylation status in carcinogenesis. We therefore investigated the expression patterns of UTX and JMJD3 in renal cell carcinoma (RCC and their roles in cancer development. Methods The mRNA expression levels of the UTX and JMJD3 genes were determined in cancer tissues and adjacent normal tissues in 36 patients with primary RCC, using quantitative real-time-polymerase chain reaction. The UTX and JMJD3 protein contents were measured by western blotting and immunohistochemical analysis. Results UTX and JMJD3 transcripts were significantly increased in cancer tissues compared to normal tissues (P p16INK4a were also increased in cancer tissues (P  Conclusions This study demonstrated that UTX and JMJD3 were upregulated in cancer tissues, suggesting that they may be involved in the development of primary RCC. The potential roles of H3K27 demethylases as biomarkers in the early diagnosis of RCC need to be further explored.

  17. Dangerous R loops form in the absence of H3K9 methylation

    DEFF Research Database (Denmark)

    Salcini, Anna Elisabetta

    2016-01-01

    Methylation of histone H3 on lysine 9 (H3K9) is a hallmark of transcriptionally inactive heterochromatin that is deregulated in pathological conditions. A new study shows that complete loss of H3K9 methylation in Caenorhabditis elegans leads to derepression of repetitive elements and formation of...... of DNA:RNA hybrids (R loops), resulting in increased rates of repeat-specific mutation....

  18. Contribution of the two genes encoding histone variant h3.3 to viability and fertility in mice.

    Directory of Open Access Journals (Sweden)

    Michelle C W Tang

    2015-02-01

    Full Text Available Histones package DNA and regulate epigenetic states. For the latter, probably the most important histone is H3. Mammals have three near-identical H3 isoforms: canonical H3.1 and H3.2, and the replication-independent variant H3.3. This variant can accumulate in slowly dividing somatic cells, replacing canonical H3. Some replication-independent histones, through their ability to incorporate outside S-phase, are functionally important in the very slowly dividing mammalian germ line. Much remains to be learned of H3.3 functions in germ cell development. Histone H3.3 presents a unique genetic paradigm in that two conventional intron-containing genes encode the identical protein. Here, we present a comprehensive analysis of the developmental effects of null mutations in each of these genes. H3f3a mutants were viable to adulthood. Females were fertile, while males were subfertile with dysmorphic spermatozoa. H3f3b mutants were growth-deficient, dying at birth. H3f3b heterozygotes were also growth-deficient, with males being sterile because of arrest of round spermatids. This sterility was not accompanied by abnormalities in sex chromosome inactivation in meiosis I. Conditional ablation of H3f3b at the beginning of folliculogenesis resulted in zygote cleavage failure, establishing H3f3b as a maternal-effect gene, and revealing a requirement for H3.3 in the first mitosis. Simultaneous ablation of H3f3a and H3f3b in folliculogenesis resulted in early primary oocyte death, demonstrating a crucial role for H3.3 in oogenesis. These findings reveal a heavy reliance on H3.3 for growth, gametogenesis, and fertilization, identifying developmental processes that are particularly susceptible to H3.3 deficiency. They also reveal partial redundancy in function of H3f3a and H3f3b, with the latter gene being generally the most important.

  19. A sequence in the Drosophila H3-H4 promoter triggers 1 Histone Locus Body assembly and biosynthesis of replication-coupled histone mRNAs

    Science.gov (United States)

    Salzler, Harmony R.; Tatomer, Deirdre C.; Malek, Pamela Y.; McDaniel, Stephen L.; Orlando, Anna N.; Marzluff, William F.; Duronio, Robert J.

    2013-01-01

    Compartmentalization of RNA biosynthetic factors into nuclear bodies (NBs) is a ubiquitous feature of eukaryotic cells. How NBs initially assemble and ultimately affect gene expression remains unresolved. The histone locus body (HLB) contains factors necessary for replication-coupled histone mRNA transcription and processing and associates with histone gene clusters. Using a transgenic assay for ectopic Drosophila HLB assembly, we show that a sequence located between, and transcription from, the divergently transcribed H3-H4 genes nucleates HLB formation and activates other histone genes in the histone gene cluster. In the absence of transcription from the H3-H4 promoter, “proto-HLBs”, containing only a subset of HLB components, form and the adjacent histone H2a-H2b genes are not expressed. Proto-HLBs also transiently form in mutant embryos with the histone locus deleted. We conclude that HLB assembly occurs through a stepwise process involving stochastic interactions of individual components that localize to a specific sequence in the H3-H4 promoter. PMID:23537633

  20. Structure-based nuclear import mechanism of histones H3 and H4 mediated by Kap123

    Energy Technology Data Exchange (ETDEWEB)

    An, Sojin [Department of Biological Chemistry, University of Michigan Medical School, Michigan, United States; Yoon, Jungmin [Structural Biology Laboratory of Epigenetics, Department of Biological Sciences, Graduate school of Nanoscience and Technology (World Class University), KI for the BioCentury, Korea Advanced Institute of Science and Technology, Daejeon, South Korea; Kim, Hanseong [Department of Biological Chemistry, University of Michigan Medical School, Michigan, United States; Song, Ji-Joon [Structural Biology Laboratory of Epigenetics, Department of Biological Sciences, Graduate school of Nanoscience and Technology (World Class University), KI for the BioCentury, Korea Advanced Institute of Science and Technology, Daejeon, South Korea; Cho, Uhn-soo [Department of Biological Chemistry, University of Michigan Medical School, Michigan, United States

    2017-10-16

    Kap123, a major karyopherin protein of budding yeast, recognizes the nuclear localization signals (NLSs) of cytoplasmic histones H3 and H4 and translocates them into the nucleus during DNA replication. Mechanistic questions include H3- and H4-NLS redundancy toward Kap123 and the role of the conserved diacetylation of cytoplasmic H4 (K5ac and K12ac) in Kap123-mediated histone nuclear translocation. Here, we report crystal structures of full-length Kluyveromyces lactis Kap123 alone and in complex with H3- and H4-NLSs. Structures reveal the unique feature of Kap123 that possesses two discrete lysine-binding pockets for NLS recognition. Structural comparison illustrates that H3- and H4-NLSs share at least one of two lysine-binding pockets, suggesting that H3- and H4-NLSs are mutually exclusive. Additionally, acetylation of key lysine residues at NLS, particularly H4-NLS diacetylation, weakens the interaction with Kap123. These data support that cytoplasmic histone H4 diacetylation weakens the Kap123-H4-NLS interaction thereby facilitating histone Kap123-H3-dependent H3:H4/Asf1 complex nuclear translocation.

  1. Squamocin modulates histone H3 phosphorylation levels and induces G1 phase arrest and apoptosis in cancer cells.

    Science.gov (United States)

    Lee, Chien-Chih; Lin, Yi-Hsiung; Chang, Wen-Hsin; Lin, Pei-Chin; Wu, Yang-Chang; Chang, Jan-Gowth

    2011-02-08

    Histone modifications in tumorigenesis are increasingly recognized as important epigenetic factors leading to cancer. Increased phosphorylation levels of histone H3 as a result of aurora B and pMSK1 overexpression were observed in various tumors. We selected aurora B and MSK1 as representatives for testing various compounds and drugs, and found that squamocin, a bis-tetrahydrofuran annonaceous acetogenin, exerted a potent effect on histone H3 phosphorylation. GBM8401, Huh-7, and SW620 cells were incubated with 15, 30, and 60 μM squamocin for 24 h. The expressions of mRNA and proteins were analyzed by qRT-PCR and Western blotting, respectively. The cell viability was determined by an MTT assay. Cell cycle distribution and apoptotic cells were analyzed by flow cytometry. Our results showed that squamocin inhibited the proliferation of GBM8401, Huh-7, and SW620 cells, arrested the cell cycle at the G1 phase, and activated both intrinsic and extrinsic pathways to apoptosis. In addition, we demonstrated that squamocin had the ability to modulate the phosphorylation levels of H3S10 (H3S10p) and H3S28 (H3S28p) in association with the downregulation of aurora B and pMSK1 expressions. This study is the first to show that squamocin affects epigenetic alterations by modulating histone H3 phosphorylation at S10 and S28, providing a novel view of the antitumor mechanism of squamocin.

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

    Directory of Open Access Journals (Sweden)

    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.

  3. Histone H3K9 trimethylase Eggless controls germline stem cell maintenance and differentiation.

    Directory of Open Access Journals (Sweden)

    Xiaoxi Wang

    2011-12-01

    study has revealed the essential roles of histone H3K9 trimethylation in controlling stem cell maintenance and differentiation through distinct mechanisms.

  4. Histone crosstalk between H3S10ph and H4K16ac generates a histone code that mediates transcription elongation.

    Science.gov (United States)

    Zippo, Alessio; Serafini, Riccardo; Rocchigiani, Marina; Pennacchini, Susanna; Krepelova, Anna; Oliviero, Salvatore

    2009-09-18

    The phosphorylation of the serine 10 at histone H3 has been shown to be important for transcriptional activation. Here, we report the molecular mechanism through which H3S10ph triggers transcript elongation of the FOSL1 gene. Serum stimulation induces the PIM1 kinase to phosphorylate the preacetylated histone H3 at the FOSL1 enhancer. The adaptor protein 14-3-3 binds the phosphorylated nucleosome and recruits the histone acetyltransferase MOF, which triggers the acetylation of histone H4 at lysine 16 (H4K16ac). This histone crosstalk generates the nucleosomal recognition code composed of H3K9acS10ph/H4K16ac determining a nucleosome platform for the bromodomain protein BRD4 binding. The recruitment of the positive transcription elongation factor b (P-TEFb) via BRD4 induces the release of the promoter-proximal paused RNA polymerase II and the increase of its processivity. Thus, the single phosphorylation H3S10ph at the FOSL1 enhancer triggers a cascade of events which activate transcriptional elongation.

  5. Molecular pathways: deregulation of histone h3 lysine 27 methylation in cancer-different paths, same destination.

    Science.gov (United States)

    Ezponda, Teresa; Licht, Jonathan D

    2014-10-01

    Methylation of lysine 27 on histone H3 (H3K27me), a modification associated with gene repression, plays a critical role in regulating the expression of genes that determine the balance between cell differentiation and proliferation. Alteration of the level of this histone modification has emerged as a recurrent theme in many types of cancer, demonstrating that either excess or lack of H3K27 methylation can have oncogenic effects. Cancer genome sequencing has revealed the genetic basis of H3K27me deregulation, including mutations of the components of the H3K27 methyltransferase complex PRC2 and accessory proteins, and deletions and inactivating mutations of the H3K27 demethylase UTX in a wide variety of neoplasms. More recently, mutations of lysine 27 on histone H3 itself were shown to prevent H3K27me in pediatric glioblastomas. Aberrant expression or mutations in proteins that recognize H3K27me3 also occur in cancer and may result in misinterpretation of this mark. In addition, due to the cross-talk between different epigenetic modifications, alterations of chromatin modifiers controlling H3K36me, or even mutations of this residue, can ultimately regulate H3K27me levels and distribution across the genome. The significance of mutations altering H3K27me is underscored by the fact that many tumors harboring such lesions often have a poor clinical outcome. New therapeutic approaches targeting aberrant H3K27 methylation include small molecules that block the action of mutant EZH2 in germinal center-derived lymphoma. Understanding the biologic consequences and gene expression pathways affected by aberrant H3K27 methylation may also lead to other new therapeutic strategies. ©2014 American Association for Cancer Research.

  6. Social exclusion changes histone modifications H3K4me3 and H3K27ac in liver tissue of wild house mice.

    Science.gov (United States)

    Krause, Linda; Haubold, Bernhard; Börsch-Haubold, Angelika G

    2015-01-01

    Wild house mice form social hierarchies with aggressive males defending territories, in which females, young mice and submissive adult males share nests. In contrast, socially excluded males are barred from breeding groups, have numerous bite wounds and patches of thinning fur. Since their feeding times are often disrupted, we investigated whether social exclusion leads to changes in epigenetic marks of metabolic genes in liver tissue. We used chromatin immunoprecipitation and quantitative PCR to measure enrichment of two activating histone marks at 15 candidate loci. The epigenetic profiles of healthy males sampled from nest boxes differed significantly from the profiles of ostracized males caught outside of nests and showing bite wounds indicative of social exclusion. Enrichment of histone-3 lysine-4 trimethylation (H3K4me3) changed significantly at genes Cyp4a14, Gapdh, Nr3c1, Pck1, Ppara, and Sqle. Changes at histone-3 lysine-27 acetylation (H3K27ac) marks were detected at genes Fasn, Nr3c1, and Plin5. A principal components analysis separated the socialized from the ostracized mice. This was independent of body weight for the H3K4me3 mark, and partially dependent for H3K27ac. There was no separation, however, between healthy males that had been sampled from two different nests. A hierarchical cluster analysis also separated the two phenotypes, which was independent of body weight for both markers. Our study shows that a period of social exclusion during adult life leads to quantitative changes in histone modification patterns in mouse liver tissue. Similar epigenetic changes might occur during the development of stress-induced metabolic disorders in humans.

  7. Systems Level Analysis of Histone H3 Post-translational Modifications (PTMs) Reveals Features of PTM Crosstalk in Chromatin Regulation

    DEFF Research Database (Denmark)

    Schwämmle, Veit; Sidoli, Simone; Ruminowicz, Chrystian

    2016-01-01

    molecules contain multiple coexisting PTMs, some of which exhibit crosstalk, i.e. coordinated or mutually exclusive activities. Here, we present an integrated experimental and computational systems level molecular characterization of histone PTMs and PTM crosstalk. Using wild type and engineered mouse....... We characterized combinatorial PTM features across the four mESC lines and then applied statistical data analysis to predict crosstalk between histone H3 PTMs. We detected an overrepresentation of positive crosstalk (codependent marks) between adjacent mono-methylated and acetylated marks......, and negative crosstalk (mutually exclusive marks) among most of the seven characterized di- and tri-methylated lysine residues in the H3 tails. We report novel features of PTM interplay involving hitherto poorly characterized arginine methylation and lysine methylation sites, including H3R2me, H3R8me and H3K37...

  8. Timing of transcriptional quiescence during gametogenesis is controlled by global histone H3K4 demethylation.

    Science.gov (United States)

    Xu, Mengshu; Soloveychik, Maria; Ranger, Mathieu; Schertzberg, Michael; Shah, Zarna; Raisner, Ryan; Venkatasubrahmanyan, Shivkumar; Tsui, Kyle; Gebbia, Marinella; Hughes, Tim; van Bakel, Harm; Nislow, Corey; Madhani, Hiten D; Meneghini, Marc D

    2012-11-13

    Gametes are among the most highly specialized cells produced during development. Although gametogenesis culminates in transcriptional quiescence in plants and animals, regulatory mechanisms controlling this are unknown. Here, we confirm that gamete differentiation in the single-celled yeast Saccharomyces cerevisiae is accompanied by global transcriptional shutoff following the completion of meiosis. We show that Jhd2, a highly conserved JARID1-family histone H3K4 demethylase, activates protein-coding gene transcription in opposition to this programmed transcriptional shutoff, sustaining the period of productive transcription during spore differentiation. Moreover, using genome-wide nucleosome, H3K4me, and transcript mapping experiments, we demonstrate that JHD2 globally represses intergenic noncoding transcription during this period. The widespread transcriptional defects of JHD2 mutants are associated with precocious differentiation and the production of stress-sensitive spores, demonstrating that Jhd2 regulation of the global postmeiotic transcriptional program is critical for the production of healthy meiotic progeny. Copyright © 2012 Elsevier Inc. All rights reserved.

  9. UTX and JMJD3 are histone H3K27 demethylases involved in HOX gene regulation and development

    DEFF Research Database (Denmark)

    Agger, Karl; Cloos, Paul A C; Christensen, Jesper

    2007-01-01

    -methylation of Lys 27 on histone H3 (H3K27me2/me3). Owing to the essential role of the PRC2 complex in repressing a large number of genes involved in somatic processes, the H3K27me3 mark is associated with the unique epigenetic state of stem cells. The rapid decrease of the H3K27me3 mark during specific stages...... of embryogenesis and stem-cell differentiation indicates that histone demethylases specific for H3K27me3 may exist. Here we show that the human JmjC-domain-containing proteins UTX and JMJD3 demethylate tri-methylated Lys 27 on histone H3. Furthermore, we demonstrate that ectopic expression of JMJD3 leads...... to a strong decrease of H3K27me3 levels and causes delocalization of polycomb proteins in vivo. Consistent with the strong decrease in H3K27me3 levels associated with HOX genes during differentiation, we show that UTX directly binds to the HOXB1 locus and is required for its activation. Finally mutation of F...

  10. Dynamic changes of histone H3 marks during Caenorhabditis elegans lifecycle revealed by middle-down proteomics.

    Science.gov (United States)

    Sidoli, Simone; Vandamme, Julien; Salcini, Anna Elisabetta; Jensen, Ole N

    2016-02-01

    We applied a middle-down proteomics strategy for large-scale protein analysis during in vivo development of Caenorhabditis elegans. We characterized PTMs on histone H3 N-terminal tails at eight time points during the C. elegans lifecycle, including embryo, larval stages (L1-L4), dauer, and L1/L4 postdauer. Histones were analyzed by our optimized middle-down protein sequencing platform using high mass accuracy MS/MS. This allows quantification of intact histone tails and detailed characterization of distinct histone tails carrying cooccurring PTMs. We measured temporally distinct combinatorial PTM profiles during C. elegans development. We show that the doubly modified form H3K23me3K27me3, which is rare or nonexistent in mammals, is the most abundant PTM in all stages of C. elegans lifecycle. The abundance of H3K23me3 increased during development and it was mutually exclusive of the active marks H3K18ac, R26me1, and R40me1, suggesting a role for H3K23me3 in silent chromatin. We observed distinct PTM profiles for normal L1 larvae and for L1-postdauer larvae, or L4 and L4 postdauer, suggesting that histone PTMs mediate an epigenetic memory that is transmitted during dauer formation. Collectively, our data describe the dynamics of histone H3 combinatorial code during C. elegans lifecycle and demonstrate the feasibility of using middle-down proteomics to study in vivo development of multicellular organisms. All MS data have been deposited in the ProteomeXchange with identifier PXD002525 (http://proteomecentral.proteomexchange.org/dataset/PXD002525). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Distinct chromatin environment associated with phosphorylated H3S10 histone during pollen mitosis I in orchids.

    Science.gov (United States)

    Sharma, Santosh Kumar; Yamamoto, Maki; Mukai, Yasuhiko

    2017-01-01

    Pollen developmental pathway in plants involving synchronized transferal of cellular divisions from meiosis (microsporogenesis) to mitosis (pollen mitosis I/II) eventually offers a unique "meiosis-mitosis shift" at pollen mitosis I. Since the cell type (haploid microspore) and fate of pollen mitosis I differ from typical mitosis (in meristem cells), it is immensely important to analyze the chromosomal distribution of phosphorylated H3S10 histone during atypical pollen mitosis I to comprehend the role of histone phosphorylation in pollen development. We investigated the chromosomal phosphorylation of H3S10 histone during pollen mitosis I in orchids using immunostaining technique. The chromosomal distribution of H3S10ph during pollen mitosis I revealed differential pattern than that of typical mitosis in plants, however, eventually following the similar trends of mitosis in animals where H3S10 phosphorylation begins in the pericentromeric regions first, later extending to the whole chromosomes, and finally declining at anaphase/early cytokinesis (differentiation of vegetative and generative cells). The study suggests that the chromosomal distribution of H3S10ph during cell division is not universal and can be altered between different cell types encoded for diverse cellular processes. During pollen development, phosphorylation of histone might play a critical role in chromosome condensation events throughout pollen mitosis I in plants.

  12. Dual modified antiphospho (Ser10)-acetyl (Lys14)-histone H3 ...

    Indian Academy of Sciences (India)

    SANTOSH KUMAR SHARMA

    histone may serve as an additional cytogenetic landmark to identify pericentromeric chromatin during mitosis in plants. The plausible role of histone cross talk and future perspectives of combinatorial histone modification marks in plant cytogenetics with special reference to chromatin dynamics have been discussed.

  13. Accumulation of histone variant H3.3 with age is associated with profound changes in the histone methylation landscape

    DEFF Research Database (Denmark)

    Tvardovskiy, Andrey; Schwämmle, Veit; Kempf, Stefan J

    2017-01-01

    a causal relationship between H3 variant replacement and age-dependent changes in H3 methylation. Furthermore, the H3.3 level is drastically reduced in human hepatocarcinoma cells as compared to nontumoral hepatocytes, suggesting the potential utility of the H3.3 relative abundance as a biomarker...

  14. 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.

  15. Both H4K20 mono-methylation and H3K56 acetylation mark transcription-dependent histone turnover in fission yeast

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Hanna [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 34141 (Korea, Republic of); Kwon, Chang Seob [Department of Chemistry and Biology, Korea Science Academy of KAIST, Busan, 614-822 (Korea, Republic of); Choi, Yoonjung, E-mail: jjungii@kaist.ac.kr [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 34141 (Korea, Republic of); Lee, Daeyoup, E-mail: daeyoup@kaist.ac.kr [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 34141 (Korea, Republic of)

    2016-08-05

    Nucleosome dynamics facilitated by histone turnover is required for transcription as well as DNA replication and repair. Histone turnover is often associated with various histone modifications such as H3K56 acetylation (H3K56Ac), H3K36 methylation (H3K36me), and H4K20 methylation (H4K20me). In order to correlate histone modifications and transcription-dependent histone turnover, we performed genome wide analyses for euchromatic regions in G2/M-arrested fission yeast. The results show that transcription-dependent histone turnover at 5′ promoter and 3′ termination regions is directly correlated with the occurrence of H3K56Ac and H4K20 mono-methylation (H4K20me1) in actively transcribed genes. Furthermore, the increase of H3K56Ac and H4K20me1 and antisense RNA production was observed in the absence of the histone H3K36 methyltransferase Set2 and histone deacetylase complex (HDAC) that are involved in the suppression of histone turnover within the coding regions. These results together indicate that H4K20me1 as well as H3K56Ac are bona fide marks for transcription-dependent histone turnover in fission yeast.

  16. Structure–function studies of histone H3/H4 tetramer maintenance during transcription by chaperone Spt2

    Science.gov (United States)

    Chen, Shoudeng; Rufiange, Anne; Huang, Hongda; Rajashankar, Kanagalaghatta R.; Nourani, Amine; Patel, Dinshaw J.

    2015-01-01

    Cells use specific mechanisms such as histone chaperones to abrogate the inherent barrier that the nucleosome poses to transcribing polymerases. The current model postulates that nucleosomes can be transiently disrupted to accommodate passage of RNA polymerases and that histones H3 and H4 possess their own chaperones dedicated to the recovery of nucleosomes. Here, we determined the crystal structure of the conserved C terminus of human Suppressors of Ty insertions 2 (hSpt2C) chaperone bound to an H3/H4 tetramer. The structural studies demonstrate that hSpt2C is bound to the periphery of the H3/H4 tetramer, mimicking the trajectory of nucleosomal-bound DNA. These structural studies have been complemented with in vitro binding and in vivo functional studies on mutants that disrupt key intermolecular contacts involving two acidic patches and hydrophobic residues on Spt2C. We show that contacts between both human and yeast Spt2C with the H3/H4 tetramer are required for the suppression of H3/H4 exchange as measured by H3K56ac and new H3 deposition. These interactions are also crucial for the inhibition of spurious transcription from within coding regions. Together, our data indicate that Spt2 interacts with the periphery of the H3/H4 tetramer and promotes its recycling in the wake of RNA polymerase. PMID:26109053

  17. Transcription recovery after DNA damage requires chromatin priming by the H3.3 histone chaperone HIRA.

    Science.gov (United States)

    Adam, Salomé; Polo, Sophie E; Almouzni, Geneviève

    2013-09-26

    Understanding how to recover fully functional and transcriptionally active chromatin when its integrity has been challenged by genotoxic stress is a critical issue. Here, by investigating how chromatin dynamics regulate transcriptional activity in response to DNA damage in human cells, we identify a pathway involving the histone chaperone histone regulator A (HIRA) to promote transcription restart after UVC damage. Our mechanistic studies reveal that HIRA accumulates at sites of UVC irradiation upon detection of DNA damage prior to repair and deposits newly synthesized H3.3 histones. This local action of HIRA depends on ubiquitylation events associated with damage recognition. Furthermore, we demonstrate that the early and transient function of HIRA in response to DNA damage primes chromatin for later reactivation of transcription. We propose that HIRA-dependent histone deposition serves as a chromatin bookmarking system to facilitate transcription recovery after genotoxic stress. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. Squamocin modulates histone H3 phosphorylation levels and induces G1 phase arrest and apoptosis in cancer cells

    Directory of Open Access Journals (Sweden)

    Wu Yang-Chang

    2011-02-01

    Full Text Available Abstract Background Histone modifications in tumorigenesis are increasingly recognized as important epigenetic factors leading to cancer. Increased phosphorylation levels of histone H3 as a result of aurora B and pMSK1 overexpression were observed in various tumors. We selected aurora B and MSK1 as representatives for testing various compounds and drugs, and found that squamocin, a bis-tetrahydrofuran annonaceous acetogenin, exerted a potent effect on histone H3 phosphorylation. Methods GBM8401, Huh-7, and SW620 cells were incubated with 15, 30, and 60 μM squamocin for 24 h. The expressions of mRNA and proteins were analyzed by qRT-PCR and Western blotting, respectively. The cell viability was determined by an MTT assay. Cell cycle distribution and apoptotic cells were analyzed by flow cytometry. Results Our results showed that squamocin inhibited the proliferation of GBM8401, Huh-7, and SW620 cells, arrested the cell cycle at the G1 phase, and activated both intrinsic and extrinsic pathways to apoptosis. In addition, we demonstrated that squamocin had the ability to modulate the phosphorylation levels of H3S10 (H3S10p and H3S28 (H3S28p in association with the downregulation of aurora B and pMSK1 expressions. Conclusions This study is the first to show that squamocin affects epigenetic alterations by modulating histone H3 phosphorylation at S10 and S28, providing a novel view of the antitumor mechanism of squamocin.

  19. High expression of trimethylated histone H3 at lysine 27 predicts better prognosis in non-small cell lung cancer.

    Science.gov (United States)

    Chen, Xiaohui; Song, Ning; Matsumoto, Keitaro; Nanashima, Atsushi; Nagayasu, Takeshi; Hayashi, Tomayoshi; Ying, Mingang; Endo, Daisuke; Wu, Zhiren; Koji, Takehiko

    2013-11-01

    Epigenetic parameters such as DNA methylation and histone modifications play pivotal roles in carcinogenesis. Global histone modification patterns have been implicated as possible predictors of cancer recurrence and prognoses in a great variety of tumor entities. Our study was designed to evaluate the association among trimethylated histone H3 at lysine 27 (H3K27me3), clinicopathological variables and outcome in early-stage non-small cell lung cancer (NSCLC). The expression of H3K27me3 and its methyl-transferase, enhancer of zeste homolog 2 (EZH2) together with proliferating cell nuclear antigen (PCNA) were evaluated by immunohistochemistry in normal lung tissue (n=5) and resected NSCLC patients (n=42). In addition, the specificity of antibody for H3K27me3 was tested by western blot analysis. The optimal cut-off point of H3K27me3 expression for prognosis was determined by the X-tile program. The prognostic significance was determined by means of Kaplan-Meier survival estimates and log-rank tests. As a result, enhanced trimethylation of H3K27me3 was correlated with longer overall survival (OS) and better prognosis (P<0.05). Moreover, both univariate and multivariate analyses indicated that H3K27me3 level was a significant and independent predictor of better survival (hazard ratio, 0.187; 95% confidence interval, 0.066-0.531, P=0.002). Furthermore, H3K27me3 expression was positively correlated with DNA methylation level at CCGG sites while reversely related to EZH2 expression (P<0.05). In conclusion, H3K27me3 level defines unrecognized subgroups of NSCLC patients with distinct epigenetic phenotype and clinical outcome, and can probably be used as a novel predictor for better prognosis in NSCLC patients.

  20. 3-Deoxyglucosone: a potential glycating agent accountable for structural alteration in H3 histone protein through generation of different AGEs.

    Directory of Open Access Journals (Sweden)

    Jalaluddin M Ashraf

    Full Text Available Advanced glycation end-products (AGEs are heterogeneous group of compounds, known to be implicated in diabetic complications. One of the consequences of the Maillard reaction is attributed to the production of reactive intermediate products such as α-oxoaldehydes. 3-deoxyglucosone (3-DG, an α-oxoaldehyde has been found to be involved in accelerating vascular damage during diabetes. In the present study, calf thymus histone H3 was treated with 3-deoxyglucosone to investigate the generation of AGEs (Nε-carboxymethyllysine, pentosidine, by examining the degree of side chain modifications and formation of different intermediates and employing various physicochemical techniques. The results clearly indicate the formation of AGEs and structural changes upon glycation of H3 by 3-deoxyglucosone, which may hamper the normal functioning of H3 histone, that may compromise the veracity of chromatin structures and function in secondary complications of diabetes.

  1. SET DOMAIN GROUP 708, a histone H3 lysine 36-specific methyltransferase, controls flowering time in rice (Oryza sativa).

    Science.gov (United States)

    Liu, Bing; Wei, Gang; Shi, Jinlei; Jin, Jing; Shen, Ting; Ni, Ting; Shen, Wen-Hui; Yu, Yu; Dong, Aiwu

    2016-04-01

    As a key epigenetic modification, the methylation of histone H3 lysine 36 (H3K36) modulates chromatin structure and is involved in diverse biological processes. To better understand the language of H3K36 methylation in rice (Oryza sativa), we chose potential histone methylation enzymes for functional exploration. In particular, we characterized rice SET DOMAIN GROUP 708 (SDG708) as an H3K36-specific methyltransferase possessing the ability to deposit up to three methyl groups on H3K36. Compared with the wild-type, SDG708-knockdown rice mutants displayed a late-flowering phenotype under both long-day and short-day conditions because of the down-regulation of the key flowering regulatory genes Heading date 3a (Hd3a), RICE FLOWERING LOCUS T1 (RFT1), and Early heading date 1 (Ehd1). Chromatin immunoprecipitation experiments indicated that H3K36me1, H3K36me2, and H3K36me3 levels were reduced at these loci in SDG708-deficient plants. More importantly, SDG708 was able to directly target and effect H3K36 methylation on specific flowering genes. In fact, knockdown of SDG708 led to misexpression of a set of functional genes and a genome-wide decrease in H3K36me1/2/3 levels during the early growth stages of rice. SDG708 is a methyltransferase that catalyses genome-wide deposition of all three methyl groups on H3K36 and is involved in many biological processes in addition to flowering promotion. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

  2. SETD2-Dependent Histone H3K36 Trimethylation Is Required for Homologous Recombination Repair and Genome Stability

    Directory of Open Access Journals (Sweden)

    Sophia X. Pfister

    2014-06-01

    Full Text Available Modulating chromatin through histone methylation orchestrates numerous cellular processes. SETD2-dependent trimethylation of histone H3K36 is associated with active transcription. Here, we define a role for H3K36 trimethylation in homologous recombination (HR repair in human cells. We find that depleting SETD2 generates a mutation signature resembling RAD51 depletion at I-SceI-induced DNA double-strand break (DSB sites, with significantly increased deletions arising through microhomology-mediated end-joining. We establish a presynaptic role for SETD2 methyltransferase in HR, where it facilitates the recruitment of C-terminal binding protein interacting protein (CtIP and promotes DSB resection, allowing Replication Protein A (RPA and RAD51 binding to DNA damage sites. Furthermore, reducing H3K36me3 levels by overexpressing KDM4A/JMJD2A, an oncogene and H3K36me3/2 demethylase, or an H3.3K36M transgene also reduces HR repair events. We propose that error-free HR repair within H3K36me3-decorated transcriptionally active genomic regions promotes cell homeostasis. Moreover, these findings provide insights as to why oncogenic mutations cluster within the H3K36me3 axis.

  3. ZMYND11 links histone H3.3 K36 trimethylation to transcription elongation and tumor suppression

    Science.gov (United States)

    Wen, Hong; Li, Yuanyuan; Xi, Yuanxin; Jiang, Shiming; Stratton, Sabrina; Peng, Danni; Tanaka, Kaori; Ren, Yongfeng; Xia, Zheng; Wu, Jun; Li, Bing; Barton, Michelle C.; Li, Wei; Li, Haitao; Shi, Xiaobing

    2014-01-01

    Recognition of modified histones by “reader” proteins plays a critical role in the regulation of chromatin1. H3K36 trimethylation (H3K36me3) is deposited onto the nucleosomes in the transcribed regions following RNA polymerase II (Pol II) elongation. In yeast, this mark in turn recruits epigenetic regulators to reset the chromatin to a relatively repressive state thus suppressing cryptic transcription2. However, much less is known about the role of H3K36me3 in transcription regulation in mammals. This is further complicated by the transcription-coupled incorporation of the histone variant H3.3 in gene bodies3. Here we show that the candidate tumor suppressor ZMYND11 specifically recognizes H3K36me3 on H3.3 (H3.3K36me3) and regulates Pol II elongation. Structural studies reveal that in addition to the trimethyl-lysine binding by an aromatic cage within the PWWP domain, the H3.3-dependent recognition is mediated by the encapsulation of the H3.3-specific “Ser31” residue in a composite pocket formed by the tandem bromo-PWWP domains of ZMYND11. ChIP-sequencing analyses reveal a genome-wide colocalization of ZMYND11 with H3K36me3 and H3.3 in gene bodies, and its occupancy requires the pre-deposition of H3.3K36me3. Although ZMYND11 is associated with highly expressed genes, it functions as an unconventional transcription corepressor via modulating Pol II at the elongation stage. ZMYND11 is critical for the repression of a transcriptional program that is essential for tumor cell growth; low expression level of ZMYND11 in breast cancer patients correlates with worse prognosis. Consistently, overexpression of ZMYND11 suppresses cancer cell growth in vitro and tumor formation in mice. Together, this study identifies ZMYND11 as an H3.3-specific reader of H3K36me3 that links the histone variant-mediated transcription elongation control to tumor suppression. PMID:24590075

  4. ZMYND11 links histone H3.3K36me3 to transcription elongation and tumour suppression.

    Science.gov (United States)

    Wen, Hong; Li, Yuanyuan; Xi, Yuanxin; Jiang, Shiming; Stratton, Sabrina; Peng, Danni; Tanaka, Kaori; Ren, Yongfeng; Xia, Zheng; Wu, Jun; Li, Bing; Barton, Michelle C; Li, Wei; Li, Haitao; Shi, Xiaobing

    2014-04-10

    Recognition of modified histones by 'reader' proteins plays a critical role in the regulation of chromatin. H3K36 trimethylation (H3K36me3) is deposited onto the nucleosomes in the transcribed regions after RNA polymerase II elongation. In yeast, this mark in turn recruits epigenetic regulators to reset the chromatin to a relatively repressive state, thus suppressing cryptic transcription. However, much less is known about the role of H3K36me3 in transcription regulation in mammals. This is further complicated by the transcription-coupled incorporation of the histone variant H3.3 in gene bodies. Here we show that the candidate tumour suppressor ZMYND11 specifically recognizes H3K36me3 on H3.3 (H3.3K36me3) and regulates RNA polymerase II elongation. Structural studies show that in addition to the trimethyl-lysine binding by an aromatic cage within the PWWP domain, the H3.3-dependent recognition is mediated by the encapsulation of the H3.3-specific 'Ser 31' residue in a composite pocket formed by the tandem bromo-PWWP domains of ZMYND11. Chromatin immunoprecipitation followed by sequencing shows a genome-wide co-localization of ZMYND11 with H3K36me3 and H3.3 in gene bodies, and its occupancy requires the pre-deposition of H3.3K36me3. Although ZMYND11 is associated with highly expressed genes, it functions as an unconventional transcription co-repressor by modulating RNA polymerase II at the elongation stage. ZMYND11 is critical for the repression of a transcriptional program that is essential for tumour cell growth; low expression levels of ZMYND11 in breast cancer patients correlate with worse prognosis. Consistently, overexpression of ZMYND11 suppresses cancer cell growth in vitro and tumour formation in mice. Together, this study identifies ZMYND11 as an H3.3-specific reader of H3K36me3 that links the histone-variant-mediated transcription elongation control to tumour suppression.

  5. Plasticity of fission yeast CENP-A chromatin driven by relative levels of histone H3 and H4.

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    Araceli G Castillo

    2007-07-01

    Full Text Available The histone H3 variant CENP-A assembles into chromatin exclusively at centromeres. The process of CENP-A chromatin assembly is epigenetically regulated. Fission yeast centromeres are composed of a central kinetochore domain on which CENP-A chromatin is assembled, and this is flanked by heterochromatin. Marker genes are silenced when placed within kinetochore or heterochromatin domains. It is not known if fission yeast CENP-A(Cnp1 chromatin is confined to specific sequences or whether histone H3 is actively excluded. Here, we show that fission yeast CENP-A(Cnp1 can assemble on noncentromeric DNA when it is inserted within the central kinetochore domain, suggesting that in fission yeast CENP-A(Cnp1 chromatin assembly is driven by the context of a sequence rather than the underlying DNA sequence itself. Silencing in the central domain is correlated with the amount of CENP-A(Cnp1 associated with the marker gene and is also affected by the relative level of histone H3. Our analyses indicate that kinetochore integrity is dependent on maintaining the normal ratio of H3 and H4. Excess H3 competes with CENP-A(Cnp1 for assembly into central domain chromatin, resulting in less CENP-A(Cnp1 and other kinetochore proteins at centromeres causing defective kinetochore function, which is manifest as aberrant mitotic chromosome segregation. Alterations in the levels of H3 relative to H4 and CENP-A(Cnp1 influence the extent of DNA at centromeres that is packaged in CENP-A(Cnp1 chromatin and the composition of this chromatin. Thus, CENP-A(Cnp1 chromatin assembly in fission yeast exhibits plasticity with respect to the underlying sequences and is sensitive to the levels of CENP-A(Cnp1 and other core histones.

  6. Acidic domains differentially read histone H3 lysine 4 methylation status and are widely present in chromatin-associated proteins.

    Science.gov (United States)

    Wu, Meng; Wei, Wei; Chen, Jiwei; Cong, Rong; Shi, Tieliu; Bouvet, Philippe; Li, Jiwen; Wong, Jiemin; Du, James X

    2017-02-01

    Histone methylation is believed to provide binding sites for specific reader proteins, which translate histone code into biological function. Here we show that a family of acidic domain-containing proteins including nucleophosmin (NPM1), pp32, SET/TAF1β, nucleolin (NCL) and upstream binding factor (UBF) are novel H3K4me2-binding proteins. These proteins exhibit a unique pattern of interaction with methylated H3K4, as their binding is stimulated by H3K4me2 and inhibited by H3K4me1 and H3K4me3. These proteins contain one or more acidic domains consisting mainly of aspartic and/or glutamic residues that are necessary for preferential binding of H3K4me2. Furthermore, we demonstrate that the acidic domain with sufficient length alone is capable of binding H3K4me2 in vitro and in vivo. NPM1, NCL and UBF require their acidic domains for association with and transcriptional activation of rDNA genes. Interestingly, by defining acidic domain as a sequence with at least 20 acidic residues in 50 continuous amino acids, we identified 655 acidic domain-containing protein coding genes in the human genome and Gene Ontology (GO) analysis showed that many of the acidic domain proteins have chromatin-related functions. Our data suggest that acidic domain is a novel histone binding motif that can differentially read the status of H3K4 methylation and is broadly present in chromatin-associated proteins.

  7. Broad histone H3K4me3 domains in mouse oocytes modulate maternal-to-zygotic transition

    DEFF Research Database (Denmark)

    Dahl, John Arne; Jung, Inkyung; Aanes, Håvard

    2016-01-01

    device that is not readily available. We developed a micro-scale chromatin immunoprecipitation and sequencing (μChIP-seq) method, which we used to profile genome-wide histone H3 lysine methylation (H3K4me3) and acetylation (H3K27ac) in mouse immature and metaphase II oocytes and in 2-cell and 8-cell...... embryos. Notably, we show that ~22% of the oocyte genome is associated with broad H3K4me3 domains that are anti-correlated with DNA methylation. The H3K4me3 signal becomes confined to transcriptional-start-site regions in 2-cell embryos, concomitant with the onset of major zygotic genome activation...

  8. A reversible histone H3 acetylation cooperates with mismatch repair and replicative polymerases in maintaining genome stability.

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    Lyudmila Y Kadyrova

    2013-10-01

    Full Text Available Mutations are a major driving force of evolution and genetic disease. In eukaryotes, mutations are produced in the chromatin environment, but the impact of chromatin on mutagenesis is poorly understood. Previous studies have determined that in yeast Saccharomyces cerevisiae, Rtt109-dependent acetylation of histone H3 on K56 is an abundant modification that is introduced in chromatin in S phase and removed by Hst3 and Hst4 in G2/M. We show here that the chromatin deacetylation on histone H3 K56 by Hst3 and Hst4 is required for the suppression of spontaneous gross chromosomal rearrangements, base substitutions, 1-bp insertions/deletions, and complex mutations. The rate of base substitutions in hst3Δ hst4Δ is similar to that in isogenic mismatch repair-deficient msh2Δ mutant. We also provide evidence that H3 K56 acetylation by Rtt109 is important for safeguarding DNA from small insertions/deletions and complex mutations. Furthermore, we reveal that both the deacetylation and acetylation on histone H3 K56 are involved in mutation avoidance mechanisms that cooperate with mismatch repair and the proofreading activities of replicative DNA polymerases in suppressing spontaneous mutagenesis. Our results suggest that cyclic acetylation and deacetylation of chromatin contribute to replication fidelity and play important roles in the protection of nuclear DNA from diverse spontaneous mutations.

  9. The HIRA complex that deposits the histone H3.3 is conserved in Arabidopsis and facilitates transcriptional dynamics

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    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.

  10. Increased phosphorylation of histone H3 at serine 10 is involved in Epstein-Barr virus latent membrane protein-1-induced carcinogenesis of nasopharyngeal carcinoma.

    Science.gov (United States)

    Li, Binbin; Huang, Guoliang; Zhang, Xiangning; Li, Rong; Wang, Jian; Dong, Ziming; He, Zhiwei

    2013-03-18

    Increased histone H3 phosphorylation is an essential regulatory mechanism for neoplastic cell transformation. We aimed to explore the role of histone H3 phosphorylation at serine10 (p-H3Ser10) in Epstein-Barr virus (EBV) latent membrane protein-1 (LMP1)-induced carcinogenesis of nasopharyngeal carcinoma (NPC). The expression of p-H3Ser10 was detected by the immunohistochemical analysis in NPC, chronic nasopharyngitis and normal nasopharynx tissues, and its correlation with LMP1 was analyzed in NPC tissues and cell lines. Using the small interfering RNA (siRNA)-H3 and histone H3 mutant (S10A), the effect of histone H3 Ser10 motif on LMP1-induced CNE1 cell proliferation, transformation and activator protein-1 (AP-1) activation were evaluated by CCK-8, focus-forming and reporter gene assay respectively. Mitogen- and stress-activated kinase 1 (MSK1) kinase activity and phosphorylation were detected by in vitro kinase assay and western blot. Using MSK1 inhibitor H89 or siRNA-MSK1, the regulatory role of MSK1 on histone H3 phosphorylation and AP-1 activation were analyzed. Immunohistochemical analysis revealed that the expression of p-H3Ser10 was significantly higher in the poorly differentiated NPC tissues than that in chronic nasopharyngitis (p histone H3 suppressed LMP1-induced CNE1 cell proliferation, foci formation and AP-1 activation. In addition, LMP1 could increase MSK1 kinase activity and phosphorylation. MSK1 inhibitor H89 or knockdown of MSK1 by siRNA blocked LMP1-induced phosphorylation of histone H3 at Ser10 and AP-1 activation. EBV-LMP1 can induce phosphorylation of histone H3 at Ser10 via MSK1. Increased phosphorylation of histone H3 at Ser10 is likely a crucial regulatory mechanism involved in LMP1-induced carcinogenesis of NPC.

  11. Histone H3 lysine 23 acetylation is associated with oncogene TRIM24 expression and a poor prognosis in breast cancer.

    Science.gov (United States)

    Ma, Li; Yuan, Lili; An, Jing; Barton, Michelle C; Zhang, Qingyuan; Liu, Zhaoliang

    2016-11-01

    Acetylated H3 lysine 23 (H3K23ac) is a specific histone post-translational modification recognized by oncoprotein TRIM24. However, it is not clear whether H3K23ac levels are correlated with TRIM24 expression and what role H3K23ac may have in cancer. In this study, we collected breast carcinoma samples from 121 patients and conducted immunohistochemistry to determine the levels of TRIM24 and H3K23ac in breast cancer. Our results demonstrated that TRIM24 expression is positively correlated with H3K23ac levels, and high levels of both TRIM24 and H3K23ac predict shorter overall survival of breast cancer patients. We also showed that both TRIM24 and H3K23ac are higher in HER2-positive patients, and their levels were positively correlated with HER2 levels in breast cancer. Moreover, TRIM24 expression is associated with estrogen receptor (ER) and progesterone receptor (PR) statuses in both our cohort and The Cancer Genome Atlas (TCGA) breast carcinoma. In summary, our results revealed an important role of TRIM24 and H3K23ac in breast cancer and provided further evidence that TRIM24 small-molecule inhibitors may benefit ER- and PR-negative or HER2-positive breast cancer patients.

  12. Balancing of Histone H3K4 Methylation States by the Kdm5c/SMCX Histone Demethylase Modulates Promoter and Enhancer Function

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    Nikolay S. Outchkourov

    2013-04-01

    Full Text Available The functional organization of eukaryotic genomes correlates with specific patterns of histone methylations. Regulatory regions in genomes such as enhancers and promoters differ in their extent of methylation of histone H3 at lysine-4 (H3K4, but it is largely unknown how the different methylation states are specified and controlled. Here, we show that the Kdm5c/Jarid1c/SMCX member of the Kdm5 family of H3K4 demethylases can be recruited to both enhancer and promoter elements in mouse embryonic stem cells and in neuronal progenitor cells. Knockdown of Kdm5c deregulates transcription via local increases in H3K4me3. Our data indicate that by restricting H3K4me3 modification at core promoters, Kdm5c dampens transcription, but at enhancers Kdm5c stimulates their activity. Remarkably, an impaired enhancer function activates the intrinsic promoter activity of Kdm5c-bound distal elements. Our results demonstrate that the Kdm5c demethylase plays a crucial and dynamic role in the functional discrimination between enhancers and core promoters.

  13. H3K36 Histone Methyltransferase Setd2 Is Required for Murine Embryonic Stem Cell Differentiation toward Endoderm

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    Yuanliang Zhang

    2014-09-01

    Full Text Available Setd2 is known as a histone-H3K36-specific methyltransferase. However, its role in physiological function remains unclear. In this study, we show that Setd2 mainly regulates differentiation of murine embryonic stem cells (mESCs toward primitive endoderm. Furthermore, we show that downregulated endoderm-related genes in Setd2−/− mESCs are associated with an aberrantly low level of Erk activity and that enforced expression of Fgfr3 can rescue the defective Erk pathway in Setd2−/− mESCs. Interestingly, the transcriptional initiation of Fgfr3 is directly regulated through histone H3K36me3 modification in its distal promoter region by Setd2. These results indicate that Setd2 controls the primitive endoderm differentiation of mESCs by regulating the Fgfr3-Erk signaling.

  14. The Histone H3 Acetylase dGcn5 Is a Key Player in Drosophila melanogaster Metamorphosis

    Science.gov (United States)

    Carré, Clément; Szymczak, Dimitri; Pidoux, Josette; Antoniewski, Christophe

    2005-01-01

    Although it has been well established that histone acetyltransferases (HATs) are involved in the modulation of chromatin structure and gene transcription, there is only little information on their developmental role in higher organisms. Gcn5 was the first transcription factor with HAT activity identified in eukaryotes. Here we report the isolation and characterization of Drosophila melanogaster dGcn5 mutants. Null dGcn5 alleles block the onset of both oogenesis and metamorphosis, while hypomorphic dGcn5 alleles impair the formation of adult appendages and cuticle. Strikingly, the dramatic loss of acetylation of the K9 and K14 lysine residues of histone H3 in dGcn5 mutants has no noticeable effect on larval tissues. In contrast, strong cell proliferation defects in imaginal tissues are observed. In vivo complementation experiments revealed that dGcn5 integrates specific functions in addition to chromosome binding and acetylation. Surprisingly, a dGcn5 variant protein with a deletion of the bromodomain, which has been shown to recognize acetylated histones, appears to be fully functional. Our results establish dGcn5 as a major histone H3 acetylase in Drosophila which plays a key role in the control of specific morphogenetic cascades during developmental transitions. PMID:16135811

  15. Transcriptional activities of histone H3, cyclin D1 and claudin 7 encoding genes in laryngeal cancer.

    Science.gov (United States)

    Kapral, Malgorzata; Strzalka-Mrozik, Barbara; Kowalczyk, Malgorzata; Paluch, Jaroslaw; Gola, Joanna; Gierek, Tatiana; Weglarz, Ludmila

    2011-05-01

    Uncontrolled proliferation and a decrease in cell-cell adhesion are one of the most important characteristics of malignancy. Determination of replication-dependent histone H3 can be applied as a proliferative marker. Cyclin D1 (CCND1) regulates the cell cycle by participating in the control of the G1/S phase transition. Claudins (CLDN) are components of tight junctions and may play an essential role in the loss of tissue cohesion. The aim of the study was to assess the mRNA expression of histone H3, cyclin D1, and claudin 7 genes in laryngeal squamous cell carcinoma (LSCC) and adjacent nonneoplastic tissues. The study group consisted of 32 patients with LSCC. Adjacent nonneoplastic tissues of incision lines were used as controls. Quantification of H3, CCND1 and CLDN7 mRNAs was performed by the use of real-time QRT-PCR assay. Molecular analysis showed a significantly higher expression of CCND1 (P = 0.0001; Wilcoxon test) and H3 (P = 0.0141) genes in tumor tissues than in surrounding nonneoplastic tissues. On the contrary, transcriptional activity of claudin 7 gene was higher in histologically normal tissues; however, this difference was not statistically significant (P = 0.1499). The data obtained indicate that laryngeal cancer is characterized by high proliferative potential mediated by increase in cyclin D1 and H3 mRNAs expression. © Springer-Verlag 2010

  16. Quantitative analysis of histone H3 and H4 post-translational modifications in doxorubicin-resistant leukemia cells.

    Science.gov (United States)

    Liu, Tao; Guo, Qingcheng; Guo, Huaizu; Hou, Sheng; Li, Jing; Wang, Hao

    2016-04-01

    The epigenetic remodeling of chromatin through histone modifications has been widely implicated in drug resistance of cancer cells. However, whether epigenetic mechanisms contribute specifically to doxorubicin resistance in leukemia has not been carefully examined. Using a stable and sensitive workflow based on LC-MS, we quantitatively compared the extents of methylation and acetylation of histone H3 and H4 in acute leukemia cell line HL60 and its doxorubicin-resistant derivative, HL60/ADR, as well as the chronic leukemia cell line K562 and its doxorubicin-resistant derivative, K562/ADR. We found that increased levels of H3K9 methylation, H3K14, H3K18 and H3K23 acetylation, and potentially H4K20 methylation, are associated with drug resistance in both cells. Our results demonstrated that the doxorubicin-resistant acute and chronic leukemia cell lines may share a common epigenetic mechanism that involves a combination of transcriptional activation and silencing. Copyright © 2015 John Wiley & Sons, Ltd.

  17. Histone H3 Lys79 methylation is required for efficient nucleotide excision repair in a silenced locus of Saccharomyces cerevisiae

    Science.gov (United States)

    Chaudhuri, Shubho; Wyrick, John J.; Smerdon, Michael J.

    2009-01-01

    Methylation of specific histone lysine residues regulates gene expression and heterochromatin function, but little is known about its role in DNA repair. To examine how changes in conserved methylated residues of histone H3 affect nucleotide excision repair (NER), viable H3K4R and H3K79R mutants were generated in Saccharomyces cerevisiae. These mutants show decreased UV survival and impaired NER at the transcriptionally silent HML locus, while maintaining normal NER in the constitutively expressed RPB2 gene and transcriptionally repressed, nucleosome loaded GAL10 gene. Moreover, the HML chromatin in these mutants has reduced accessibility to Micrococcal nuclease (MNase). Importantly, chromatin immunoprecipitation analysis demonstrates there is enhanced recruitment of the Sir complex at the HML locus of these mutants, and deletion of the SIR2 or SIR3 genes restores the MNase accessibility and DNA repair efficiency at this locus. Furthermore, following UV irradiation expression of NER genes in these mutants remains at wild type levels, with the exception of RAD16 which decreases by more than 2-fold. These results indicate that impaired NER occurs in the silenced chromatin of H3K79R and H3K4,79R mutants as a result of increased binding of Sir complexes, which may reduce DNA lesion accessibility to repair enzymes. PMID:19155276

  18. Arabidopsis COMPASS-like complexes mediate histone H3 lysine-4 trimethylation to control floral transition and plant development.

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    Danhua Jiang

    2011-03-01

    Full Text Available Histone H3 lysine-4 (H3K4 methylation is associated with transcribed genes in eukaryotes. In Drosophila and mammals, both di- and tri-methylation of H3K4 are associated with gene activation. In contrast to animals, in Arabidopsis H3K4 trimethylation, but not mono- or di-methylation of H3K4, has been implicated in transcriptional activation. H3K4 methylation is catalyzed by the H3K4 methyltransferase complexes known as COMPASS or COMPASS-like in yeast and mammals. Here, we report that Arabidopsis homologs of the COMPASS and COMPASS-like complex core components known as Ash2, RbBP5, and WDR5 in humans form a nuclear subcomplex during vegetative and reproductive development, which can associate with multiple putative H3K4 methyltransferases. Loss of function of ARABIDOPSIS Ash2 RELATIVE (ASH2R causes a great decrease in genome-wide H3K4 trimethylation, but not in di- or mono-methylation. Knockdown of ASH2R or the RbBP5 homolog suppresses the expression of a crucial Arabidopsis floral repressor, FLOWERING LOCUS C (FLC, and FLC homologs resulting in accelerated floral transition. ASH2R binds to the chromatin of FLC and FLC homologs in vivo and is required for H3K4 trimethylation, but not for H3K4 dimethylation in these loci; overexpression of ASH2R causes elevated H3K4 trimethylation, but not H3K4 dimethylation, in its target genes FLC and FLC homologs, resulting in activation of these gene expression and consequent late flowering. These results strongly suggest that H3K4 trimethylation in FLC and its homologs can activate their expression, providing concrete evidence that H3K4 trimethylation accumulation can activate eukaryotic gene expression. Furthermore, our findings suggest that there are multiple COMPASS-like complexes in Arabidopsis and that these complexes deposit trimethyl but not di- or mono-methyl H3K4 in target genes to promote their expression, providing a molecular explanation for the observed coupling of H3K4 trimethylation (but not H3

  19. Ash1l methylates Lys36 of histone H3 independently of transcriptional elongation to counteract polycomb silencing.

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    Hitomi Miyazaki

    2013-11-01

    Full Text Available Molecular mechanisms for the establishment of transcriptional memory are poorly understood. 5,6-dichloro-1-D-ribofuranosyl-benzimidazole (DRB is a P-TEFb kinase inhibitor that artificially induces the poised RNA polymerase II (RNAPII, thereby manifesting intermediate steps for the establishment of transcriptional activation. Here, using genetics and DRB, we show that mammalian Absent, small, or homeotic discs 1-like (Ash1l, a member of the trithorax group proteins, methylates Lys36 of histone H3 to promote the establishment of Hox gene expression by counteracting Polycomb silencing. Importantly, we found that Ash1l-dependent Lys36 di-, tri-methylation of histone H3 in a coding region and exclusion of Polycomb group proteins occur independently of transcriptional elongation in embryonic stem (ES cells, although both were previously thought to be consequences of transcription. Genome-wide analyses of histone H3 Lys36 methylation under DRB treatment have suggested that binding of the retinoic acid receptor (RAR to a certain genomic region promotes trimethylation in the RAR-associated gene independent of its ongoing transcription. Moreover, DRB treatment unveils a parallel response between Lys36 methylation of histone H3 and occupancy of either Tip60 or Mof in a region-dependent manner. We also found that Brg1 is another key player involved in the response. Our results uncover a novel regulatory cascade orchestrated by Ash1l with RAR and provide insights into mechanisms underlying the establishment of the transcriptional activation that counteracts Polycomb silencing.

  20. Alcohol induces parallel changes in hippocampal histone H3 phosphorylation and c-fos protein expression in male rats

    Science.gov (United States)

    McClain, Justin A.; Nixon, Kimberly

    2015-01-01

    Background Changes in gene expression associated with alcohol-induced neuroadaptations are controlled in part by post-translational histone modifications. Serine 10 phosphorylation of histone H3 (H3S10ph) has been implicated in drug-induced changes in gene expression; however, ethanol’s effects on H3S10ph have yet to be examined in brain. Therefore, hippocampal H3S10ph was examined after acute alcohol exposure and alcohol dependence. Methods Adult male Sprague-Dawley rats received an acute exposure of ethanol (0–5 g/kg) via gavage. Or, rats were made alcohol dependent by administering 25% w/v ethanol every 8 hours for 4 days following a modified Majchrowicz protocol. In both cases, rats were perfused transcardially and paraformaldehyde-fixed brains were collected and processed for immunohistochemistry for H3S10ph or the immediate early gene, c-fos. Results Acute ethanol exposure dose-dependently altered the number of H3S10ph-positive (+) cells in the hippocampus. Specifically, 1 g/kg ethanol increased the number of H3S10ph+ cells in all neuronal layers, while 2.5 and 5 g/kg ethanol reduced the number of H3S10ph+ cells, an effect that was confined to the granule cell layer (GCL). In ethanol dependent rats, the number of H3S10ph+ cells in the GCL was reduced by 66% during intoxication; however, H3S10ph+ cells were increased in all neuronal layers during peak withdrawal. Subsequent examination of c-fos, a gene known to be regulated by H3S10ph, revealed that ethanol and withdrawal-associated changes in c-fos closely paralleled changes in H3S10ph. Conclusions These results suggest that H3S10ph regulates ethanol-mediated changes in c-fos expression, effects that likely have important implications for ethanol-induced changes in hippocampal neuronal plasticity. PMID:26727528

  1. GmPHD5 acts as an important regulator for crosstalk between histone H3K4 di-methylation and H3K14 acetylation in response to salinity stress in soybean

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    Wu Tao

    2011-12-01

    Full Text Available Abstract Background Accumulated evidence suggest that specific patterns of histone posttranslational modifications (PTMs and their crosstalks may determine transcriptional outcomes. However, the regulatory mechanisms of these "histone codes" in plants remain largely unknown. Results In this study, we demonstrate for the first time that a salinity stress inducible PHD (plant homeodomain finger domain containing protein GmPHD5 can read the "histone code" underlying the methylated H3K4. GmPHD5 interacts with other DNA binding proteins, including GmGNAT1 (an acetyl transferase, GmElongin A (a transcription elongation factor and GmISWI (a chromatin remodeling protein. Our results suggest that GmPHD5 can recognize specific histone methylated H3K4, with preference to di-methylated H3K4. Here, we illustrate that the interaction between GmPHD5 and GmGNAT1 is regulated by the self-acetylation of GmGNAT1, which can also acetylate histone H3. GmGNAT1 exhibits a preference toward acetylated histone H3K14. These results suggest a histone crosstalk between methylated H3K4 and acetylated H3K14. Consistent to its putative roles in gene regulation under salinity stress, we showed that GmPHD5 can bind to the promoters of some confirmed salinity inducible genes in soybean. Conclusion Here, we propose a model suggesting that the nuclear protein GmPHD5 is capable of regulating the crosstalk between histone methylation and histone acetylation of different lysine residues. Nevertheless, GmPHD5 could also recruit chromatin remodeling factors and transcription factors of salt stress inducible genes to regulate their expression in response to salinity stress.

  2. Different Patterns of Acetylation and Dimethylation of Histone H3 between Young and Aged Cases with Chronic Tonsillitis: Influences of Inflammation and Aging.

    Science.gov (United States)

    Saito, Akihiko; Watanabe, Ken-Ichi; Egawa, Seiko; Okubo, Kimihiro

    2016-01-01

    Epigenetics is now considered to be crucially involved in normal genetics and differentiation and in pathological conditions, such as cancer, aging, and inflammation. Epigenetic mechanisms involve DNA methylation and histone modifications. The purpose of this study was to investigate the effects of inflammation on epigenetics in young subjects and the effect of aging. The palatine tonsils were extracted from child and adult patients with chronic tonsillitis. Hematoxylin-eosin staining was performed to examine the morphology of the palatine tonsils. A fluorescence immunological examination was also performed to detect acetyl-histone H3 or dimethyl-histone H3. Confocal scanning microscopy was used for observations. Acetylated histone H3 was detected in tonsils from child patients but not from adult patients. Dimethylated histone H3 was not detected in tonsils from either group of patients. Degeneration of the tonsillar structures was apparent in tonsils from adult patients. The differential expression of acetylated histone H3 Lys9 may reflect immunological differences between young and aged tonsils. The decrease observed in the activity of histone methyltransferase induced the down-regulated expression of methylated histone H3. Our results suggest that epigenetic changes participate in chronic inflammation and aging in the palatine tonsils. Although the results do not lead to a direct treatment, the epigenetic pathogenesis of chronic inflammation, such as immunoglobulin A nephropathy, by focal infections will be described in greater detail in future studies, which will lead to new treatments being developed.

  3. Whole-genome regulation analysis of histone H3 lysin 27 trimethylation in subclinical mastitis cows infected by Staphylococcus aureus.

    Science.gov (United States)

    He, Yanghua; Song, Minyan; Zhang, Yi; Li, Xizhi; Song, Jiuzhou; Zhang, Yuan; Yu, Ying

    2016-08-08

    S. aureus is one of the major etiological agents causing bovine subclinical mastitis. The regulatory effects of H3K27me3 on gene expression in subclinical S. aureus mastitis cows are unknown. This study aimed to profile genome-wide transcriptional changes regulated by H3K27me3 in bovine lymphocytes applied in subclinical S. aureus mastitis cows and healthy controls. A total of 61 differentially expressed genes (DEGs) were detected in subclinical S. aureus mastitis cows compared to the healthy controls, of which 25 DEGs are up-regulated and the rest are down-regulated genes in subclinical S.aureus mastitis cows. The up-regulated genes are mainly involved in the Jak-STAT signaling pathway, cytokine-cytokine receptor interaction, and T cell receptor-signaling pathway, while the down-regulated genes are related to metabolism pathways. Combination analysis of histone methylation and gene expression revealed that H3K27 trimethylation levels in silent genes were higher in subclinical S. aureus mastitis cattle than in healthy cows. The key regions of H3K27me3 target genes related to subclinical S. aureus mastitis were the upstream 2 kb regions of the DEGs relative to transcription start site (TSS). The current study provides a novel insight into the interaction between S. aureus and lymphocytes in lactating cows by histone H3 methylation regulation. The differentially expressed genes in bovine lymphocytes regulated by H3K27me3 on upstream 2 kb regions (IL10, PTX3 and etc.) may relate to S. aureus mastitis susceptibility and could be considered as key candidate genes for anti- S. aureus mastitis study and breeding.

  4. Dual modified antiphospho (Ser10)-acetyl (Lys14)-histone H3 ...

    Indian Academy of Sciences (India)

    2016-12-01

    Dec 1, 2016 ... Epigenetic regulatory posttranslational histone modification marks not only function individually but also capable to act in combination as a unique pattern. A total of 16 plant species belonging to 11 genera of eight families (five dicots and threemonocots) including land plants, epiphytes (orchids) and the ...

  5. Dual modified antiphospho (Ser10)-acetyl (Lys14)-histone H3 ...

    Indian Academy of Sciences (India)

    2016-12-01

    Dec 1, 2016 ... Abstract. Epigenetic regulatory posttranslational histone modification marks not only function individually but also capable to act in combination as a unique pattern. A total of 16 plant species belonging to 11 genera of eight families (five dicots and three monocots) including land plants, epiphytes (orchids) ...

  6. Genome-wide mapping of histone H3 lysine 4 trimethylation in Eucalyptus grandis developing xylem

    Science.gov (United States)

    Steven G Hussey; Eshchar Mizrachi; Andrew Groover; Dave K Berger; Alexander A Myburg

    2015-01-01

    Background: Histone modifications play an integral role in plant development, but have been poorly studied inwoody plants. Investigating chromatin organization in wood-forming tissue and its role in regulating gene expression allows us to understand the mechanisms underlying cellular differentiation during xylogenesis (wood...

  7. AMG 900, a potent inhibitor of aurora kinases causes pharmacodynamic changes in p-Histone H3 immunoreactivity in human tumor xenografts and proliferating mouse tissues.

    Science.gov (United States)

    Juan, Gloria; Bush, Tammy L; Ma, Connie; Manoukian, Raffi; Chung, Grace; Hawkins, Jennifer M; Zoog, Stephen; Kendall, Richard; Radinsky, Robert; Loberg, Robert; Friberg, Greg; Payton, Marc

    2014-11-04

    The Aurora family of serine-threonine kinases are essential regulators of cell division in mammalian cells. Aurora-A and -B expression and kinase activity is elevated in a variety of human cancers and is associated with high proliferation rates and poor prognosis. AMG 900 is a highly potent and selective pan-aurora kinase inhibitor that has entered clinical evaluation in adult patients with advanced cancers. In mice, oral administration of AMG 900 blocks the phosphorylation of histone H3 on serine-10 (p-Histone H3), a proximal substrate of aurora-B and inhibits the growth of multiple human tumor xenografts, including multidrug-resistant models. In order to establish a preclinical pharmacokinetic-pharmacodynamic (PK-PD) relationship for AMG 900 that could be translated to the clinic, we used flow cytometry and laser scanning cytometry detection platforms to assess the effects on p-Histone H3 inhibition in terms of sensitivity, precision, and specificity, in human tumor xenografts in conjunction with mouse skin and bone marrow tissues. Mice with established COLO 205 tumors were administered AMG 900 at 3.75, 7.5, and 15 mg/kg and assessed after 3 hours. Significant suppression of p-Histone H3 in mouse skin was only observed at 15 mg/kg (p Histone H3 in tumors and surrogate tissues (although tissues such as skin may be less sensitive for assessing PD effects). To further extend our work, we evaluated the feasibility of measuring p-Histone H3 using fine-needle aspirate (FNA) tumor xenograft biopsies. Treatment with AMG 900 significantly inhibited p-Histone H3 (>99% inhibition, p Histone H3 positive cells using mock FNAs from primary human breast tumor tissues. Phosphorylation of histone H3 is a useful biomarker to determine the pharmacodynamics (PD) activity of AMG 900. FNA biopsies may be a viable approach for assessing AMG 900 PD effects in the clinic.

  8. Histone H3K9 methyltransferase G9a represses PPARγ expression and adipogenesis

    National Research Council Canada - National Science Library

    Wang, Lifeng; Xu, Shiliyang; Lee, Ji‐Eun; Baldridge, Anne; Grullon, Sean; Peng, Weiqun; Ge, Kai

    ...‐mediated repressive epigenetic mark H3K9me2 is selectively enriched on the entire PPAR γ locus. H3K9me2 and G9a levels decrease during adipogenesis, which correlates inversely with induction of PPAR...

  9. CXCL8 histone H3 acetylation is dysfunctional in airway smooth muscle in asthma: regulation by BET.

    Science.gov (United States)

    Clifford, Rachel L; Patel, Jamie K; John, Alison E; Tatler, Amanda L; Mazengarb, Lisa; Brightling, Christopher E; Knox, Alan J

    2015-05-01

    Asthma is characterized by airway inflammation and remodeling and CXCL8 is a CXC chemokine that drives steroid-resistant neutrophilic airway inflammation. We have shown that airway smooth muscle (ASM) cells isolated from asthmatic individuals secrete more CXCL8 than cells from nonasthmatic individuals. Here we investigated chromatin modifications at the CXCL8 promoter in ASM cells from nonasthmatic and asthmatic donors to further understand how CXCL8 is dysregulated in asthma. ASM cells from asthmatic donors had increased histone H3 acetylation, specifically histone H3K18 acetylation, and increased binding of histone acetyltransferase p300 compared with nonasthmatic donors but no differences in CXCL8 DNA methylation. The acetylation reader proteins Brd3 and Brd4 were bound to the CXCL8 promoter and Brd inhibitors inhibited CXCL8 secretion from ASM cells by disrupting Brd4 and RNA polymerase II binding to the CXCL8 promoter. Our results show a novel dysregulation of CXCL8 transcriptional regulation in asthma characterized by a promoter complex that is abnormal in ASM cells isolated from asthmatic donors and can be modulated by Brd inhibitors. Brd inhibitors may provide a new therapeutic strategy for steroid-resistant inflammation. Copyright © 2015 the American Physiological Society.

  10. Identification and characterization of Nek6 protein kinase, a potential human homolog of NIMA histone H3 kinase.

    Science.gov (United States)

    Hashimoto, Yoshihiro; Akita, Hidetoshi; Hibino, Mitsunobu; Kohri, Kenjiro; Nakanishi, Makoto

    2002-05-03

    In Aspergillus nidulans, the kinase activity of NIMA (never in mitosis, gene A) is critical for the initiation of mitosis. NIMA regulates mitotic chromatin condensation through phosphorylation of histone H3 at serine 10. In the present study, we identified human Nek6 (hNek6), a member of the mammalian NIMA-related kinases. The predicted hNek6 protein is comprised of 338 amino acids. Northern blot analysis revealed that hNek6 transcripts are ubiquitously expressed with the highest expression found in the heart and skeletal muscle. Lower cell cycle-dependent expression of hNek6 transcripts was observed in the early G1 phase. GFP-fused hNek6 protein showed both nuclear and cytoplasmic localizations in HeLa cells. Fluorescence in situ hybridization using full-length hNek6 cDNA as a probe showed that the hNek6 gene is localized to human chromosome 9q33-34, a region at which the loss of heterozygosity is associated with transitional cell carcinomas. Importantly, recombinant hNek6 protein produced in insect cells effectively phosphorylated histones H1 and H3, but not casein. Thus, these results suggest that, unlike other mammalian NIMA-related kinases, Nek6 is a mitotic histone kinase which regulates chromatin condensation in mammalian cells. Copyright 2002 Elsevier Science (USA).

  11. Acetylation of Chromatin-Associated Histone H3 Lysine 56 Inhibits the Development of Encysted Artemia Embryos.

    Directory of Open Access Journals (Sweden)

    Rong Zhou

    Full Text Available As a response to harsh environments, the crustacean artemia produces diapause gastrula embryos (cysts, in which cell division and embryonic development are totally arrested. This dormant state can last for very long periods but be terminated by specific environmental stimuli. Thus, artemia is an ideal model organism in which to study cell cycle arrest and embryonic development.Our study focuses on the roles of H3K56ac in the arrest of cell cycle and development during artemia diapause formation and termination. We found that the level of H3K56ac on chromatin increased during diapause formation, and decreased upon diapause termination, remaining basal level throughout subsequent embryonic development. In both HeLa cells and artemia, blocking the deacetylation with nicotinamide, a histone deacetylase inhibitor, increased the level of H3K56ac on chromatin and induced an artificial cell cycle arrest. Furthermore, we found that this arrest of the cell cycle and development was induced by H3K56ac and dephosphorylation of the checkpoint protein, retinoblastoma protein.These results have revealed the dynamic change in H3K56ac on chromatin during artemia diapause formation and termination. Thus, our findings provide insight into the regulation of cell division during arrest of artemia embryonic development and provide further insight into the functions of H3K56ac.

  12. Different expression patterns of histone H3K27 demethylases in renal cell carcinoma and bladder cancer.

    Science.gov (United States)

    Hong, Zehui; Li, Hui; Li, Lili; Wang, Weilong; Xu, Ting

    2017-01-01

    UTX and JMJD3 are recently identified histone H3 lysine 27 (H3K27) demethylases. Many studies have shown aberrant H3K27 trimethylation (H3K27me3) levels widely exist in multiple cancers, and that altered H3K27me3 levels are correlated with tumorigenesis and tumor progression. To investigate expression patterns of UTX and JMJD3 genes in renal cell carcinoma (RCC) and bladder cancer and the relationship between gene expression and tumor development. Samples were collected from 35 patients with RCC and 21 patients with bladder cancer and qRT-PCR was performed. By comparing with adjacent normal tissues, the expression of JMJD3 (10/21 = 47.62%) and UTX (10/21 = 47.62%) were significantly upregulated in bladder cancer tissues and the expression of JMJD3 (15/35 = 42.86%) was significantly downregulated in RCC tissues. Stratified analyses revealed that upregulated expression of JMJD3 was significantly associated with poorly differentiated tumor nuclear grade (p= 0.005) and advanced clinical stage (p= 0.043) in the bladder cancer group, while downregulated expression of JMJD3 was significantly associated with advanced clinical stage (p= 0.045) and poorly differentiated tumor nuclear grade (p= 0.011) in the RCC group. These results suggest JMJD3 could be a hallmark and is involved in the development of RCC and bladder cancers. The potential role of H3K27 demethylases as biomarkers needs further investigations.

  13. Acetylation of Chromatin-Associated Histone H3 Lysine 56 Inhibits the Development of Encysted Artemia Embryos.

    Science.gov (United States)

    Zhou, Rong; Yang, Fan; Chen, Dian-Fu; Sun, Yu-Xia; Yang, Jin-Shu; Yang, Wei-Jun

    2013-01-01

    As a response to harsh environments, the crustacean artemia produces diapause gastrula embryos (cysts), in which cell division and embryonic development are totally arrested. This dormant state can last for very long periods but be terminated by specific environmental stimuli. Thus, artemia is an ideal model organism in which to study cell cycle arrest and embryonic development. Our study focuses on the roles of H3K56ac in the arrest of cell cycle and development during artemia diapause formation and termination. We found that the level of H3K56ac on chromatin increased during diapause formation, and decreased upon diapause termination, remaining basal level throughout subsequent embryonic development. In both HeLa cells and artemia, blocking the deacetylation with nicotinamide, a histone deacetylase inhibitor, increased the level of H3K56ac on chromatin and induced an artificial cell cycle arrest. Furthermore, we found that this arrest of the cell cycle and development was induced by H3K56ac and dephosphorylation of the checkpoint protein, retinoblastoma protein. These results have revealed the dynamic change in H3K56ac on chromatin during artemia diapause formation and termination. Thus, our findings provide insight into the regulation of cell division during arrest of artemia embryonic development and provide further insight into the functions of H3K56ac.

  14. Over-expression of histone H3K4 demethylase gene JMJ15 enhances salt tolerance in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Yuan eShen

    2014-06-01

    Full Text Available Histone H3 lysine 4 trimethylation (H3K4me3 has been shown to be involved in stress-responsive gene expression and gene priming in plants. However, the role of H3K4me3 resetting in the processes is not clear. In this work we studied the expression and function of Arabidopsis H3K4 demethylase gene JMJ15. We show that the expression of JMJ15 was relatively low and was limited to a number of tissues during vegetative growth but was higher in young floral organs. Over-expression of the gene in gain-of-function mutants reduced the plant height with accumulation of lignin in stems, while the loss-of-function mutation did not produce any visible phenotype. The gain-of-function mutants showed enhanced salt tolerance, whereas the loss-of-function mutant was more sensitive to salt compared to the wild type. Transcriptomic analysis revealed that over-expression of JMJ15 down-regulated many genes which are preferentially marked by H3K4me3 and H3K4me2. Many of the down-regulated genes encode transcription regulators involved in stress responses. The data suggest that increased JMJ15 levels may regulate the gene expression program that enhances stress tolerance.

  15. Histone H3K56 acetylation, CAF1, and Rtt106 coordinate nucleosome assembly and stability of advancing replication forks.

    Directory of Open Access Journals (Sweden)

    Marta Clemente-Ruiz

    2011-11-01

    Full Text Available Chromatin assembly mutants accumulate recombinogenic DNA damage and are sensitive to genotoxic agents. Here we have analyzed why impairment of the H3K56 acetylation-dependent CAF1 and Rtt106 chromatin assembly pathways, which have redundant roles in H3/H4 deposition during DNA replication, leads to genetic instability. We show that the absence of H3K56 acetylation or the simultaneous knock out of CAF1 and Rtt106 increases homologous recombination by affecting the integrity of advancing replication forks, while they have a minor effect on stalled replication fork stability in response to the replication inhibitor hydroxyurea. This defect in replication fork integrity is not due to defective checkpoints. In contrast, H3K56 acetylation protects against replicative DNA damaging agents by DNA repair/tolerance mechanisms that do not require CAF1/Rtt106 and are likely subsequent to the process of replication-coupled nucleosome deposition. We propose that the tight connection between DNA synthesis and histone deposition during DNA replication mediated by H3K56ac/CAF1/Rtt106 provides a mechanism for the stabilization of advancing replication forks and the maintenance of genome integrity, while H3K56 acetylation has an additional, CAF1/Rtt106-independent function in the response to replicative DNA damage.

  16. Vliv PRDM9 histon H3K4 tri-metyl transferázy na chromozomální sterilitu

    OpenAIRE

    Svatoňová, Martina

    2015-01-01

    The influence of PRDM9 histon H3K4 tri-methyltransferase on chromosomal sterility The Prdm9 gene encodes a meiotic histone methyltransferase, which enables tri- methylation of lysine 4 on histon H3. H3K4 tri-methylation is associated with gene expression activation. The same gene was identified in the mouse as the hybrid sterility 1gene. The regulation of hotspots of meiotic recombination in mouse and human is its other function. The aim of diploma thesis was be to verify the influence of sup...

  17. Saturated fatty acid palmitate induces extracellular release of histone H3: A possible mechanistic basis for high-fat diet-induced inflammation and thrombosis

    Energy Technology Data Exchange (ETDEWEB)

    Shrestha, Chandan [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Department of Laboratory and Vascular Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Ito, Takashi [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Kawahara, Ko-ichi [Department of Biomedical Engineering, Osaka Institute of Technology, Osaka (Japan); Shrestha, Binita; Yamakuchi, Munekazu; Hashiguchi, Teruto [Department of Laboratory and Vascular Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Maruyama, Ikuro, E-mail: rinken@m3.kufm.kagoshima-u.ac.jp [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan)

    2013-08-09

    Highlights: •High-fat diet feeding and palmitate induces the release of nuclear protein histone H3. •ROS production and JNK signaling mediates the release of histone H3. •Extracellular histones induces proinflammatory and procoagulant response. -- Abstract: Chronic low-grade inflammation is a key contributor to high-fat diet (HFD)-related diseases, such as type 2 diabetes, non-alcoholic steatohepatitis, and atherosclerosis. The inflammation is characterized by infiltration of inflammatory cells, particularly macrophages, into obese adipose tissue. However, the molecular mechanisms by which a HFD induces low-grade inflammation are poorly understood. Here, we show that histone H3, a major protein component of chromatin, is released into the extracellular space when mice are fed a HFD or macrophages are stimulated with the saturated fatty acid palmitate. In a murine macrophage cell line, RAW 264.7, palmitate activated reactive oxygen species (ROS) production and JNK signaling. Inhibitors of these pathways dampened palmitate-induced histone H3 release, suggesting that the extracellular release of histone H3 was mediated, in part, through ROS and JNK signaling. Extracellular histone activated endothelial cells toexpress the adhesion molecules ICAM-1 and VCAM-1 and the procoagulant molecule tissue factor, which are known to contribute to inflammatory cell recruitment and thrombosis. These results suggest the possible contribution of extracellular histone to the pathogenesis of HFD-induced inflammation and thrombosis.

  18. Cloning and Functional Analysis of Histones H3 and H4 in Nuclear Shaping during Spermatogenesis of the Chinese Mitten Crab, Eriocheir sinensis.

    Directory of Open Access Journals (Sweden)

    Jiang-Li Wu

    Full Text Available During spermatogenesis in most animals, the basic proteins associated with DNA are continuously changing and somatic-typed histones are partly replaced by sperm-specific histones, which are then successively replaced by transition proteins and protamines. With the replacement of sperm nuclear basic proteins, nuclei progressively undergo chromatin condensation. The Chinese Mitten Crab (Eriocheir sinensis is also known as the hairy crab or river crab (phylum Arthropoda, subphylum Crustacea, order Decapoda, and family Grapsidae. The spermatozoa of this species are aflagellate, and each has a spherical acrosome surrounded by a cup-shaped nucleus, peculiar to brachyurans. An interesting characteristic of the E. sinensis sperm nucleus is its lack of electron-dense chromatin. However, its formation is not clear. In this study, sequences encoding histones H3 and H4 were cloned by polymerase chain reaction amplification. Western blotting indicated that H3 and H4 existed in the sperm nuclei. Immunofluorescence and ultrastructural immunocytochemistry demonstrated that histones H3 and H4 were both present in the nuclei of spermatogonia, spermatocytes, spermatids and mature spermatozoa. The nuclear labeling density of histone H4 decreased in sperm nuclei, while histone H3 labeling was not changed significantly. Quantitative real-time PCR showed that the mRNA expression levels of histones H3 and H4 were higher at mitotic and meiotic stages than in later spermiogenesis. Our study demonstrates that the mature sperm nuclei of E. sinensis contain histones H3 and H4. This is the first report that the mature sperm nucleus of E. sinensis contains histones H3 and H4. This finding extends the study of sperm histones of E. sinensis and provides some basic data for exploring how decapod crustaceans form uncondensed sperm chromatin.

  19. Histone methyltransferase MMSET/NSD2 alters EZH2 binding and reprograms the myeloma epigenome through global and focal changes in H3K36 and H3K27 methylation.

    Directory of Open Access Journals (Sweden)

    Relja Popovic

    2014-09-01

    Full Text Available Overexpression of the histone methyltransferase MMSET in t(4;14+ multiple myeloma patients is believed to be the driving factor in the pathogenesis of this subtype of myeloma. MMSET catalyzes dimethylation of lysine 36 on histone H3 (H3K36me2, and its overexpression causes a global increase in H3K36me2, redistributing this mark in a broad, elevated level across the genome. Here, we demonstrate that an increased level of MMSET also induces a global reduction of lysine 27 trimethylation on histone H3 (H3K27me3. Despite the net decrease in H3K27 methylation, specific genomic loci exhibit enhanced recruitment of the EZH2 histone methyltransferase and become hypermethylated on this residue. These effects likely contribute to the myeloma phenotype since MMSET-overexpressing cells displayed increased sensitivity to EZH2 inhibition. Furthermore, we demonstrate that such MMSET-mediated epigenetic changes require a number of functional domains within the protein, including PHD domains that mediate MMSET recruitment to chromatin. In vivo, targeting of MMSET by an inducible shRNA reversed histone methylation changes and led to regression of established tumors in athymic mice. Together, our work elucidates previously unrecognized interplay between MMSET and EZH2 in myeloma oncogenesis and identifies domains to be considered when designing inhibitors of MMSET function.

  20. An Autoregulatory Mechanism Imposes Allosteric Control on the V(DJ Recombinase by Histone H3 Methylation

    Directory of Open Access Journals (Sweden)

    Chao Lu

    2015-01-01

    Full Text Available V(DJ recombination is initiated by a specialized transposase consisting of the subunits RAG-1 and RAG-2. The susceptibility of gene segments to DNA cleavage by the V(DJ recombinase is correlated with epigenetic modifications characteristic of active chromatin, including trimethylation of histone H3 on lysine 4 (H3K4me3. Engagement of H3K4me3 by a plant homeodomain (PHD in RAG-2 promotes recombination in vivo and stimulates DNA cleavage by RAG in vitro. We now show that H3K4me3 acts allosterically at the PHD finger to relieve autoinhibition imposed by a separate domain within RAG-2. Disruption of this autoinhibitory domain was associated with constitutive increases in recombination frequency, DNA cleavage activity, substrate binding affinity, and catalytic rate, thus mimicking the stimulatory effects of H3K4me3. Our observations support a model in which allosteric control of RAG is enforced by an autoinhibitory domain whose action is relieved by engagement of active chromatin.

  1. Alcohol Exposure Causes Overexpression of Heart Development-Related Genes by Affecting the Histone H3 Acetylation via BMP Signaling Pathway in Cardiomyoblast Cells.

    Science.gov (United States)

    Shi, Jin; Zhao, Weian; Pan, Bo; Zheng, Min; Si, Lina; Zhu, Jing; Liu, Lingjuan; Tian, Jie

    2017-01-01

    Abusive alcohol utilization of pregnant woman may cause congenital heart disease (CHD) of fetus, where alcohol ignites histone H3 hyperacetylation leading to abnormal development of heart morphogenesis and associated genes. Knowledge about the regularized upstream genes is little, but bone morphogenetic protein (BMP) signaling may actively and prominently take part in alteration in acetylation of histone H3. The supreme objective of this study was to unearth the involvement of BMP signaling pathway in alcohol-driven hyperacetylation of histone H3 in cardiomyoblast cells. Cardiomyoblast cells (H9c2 cells) were addicted with alcohol (100 mM) for 24 hours. Dorsomorphin (5 μM) was used for the inhibition of BMP signaling pathway. We detected the phosphorylation activity of SMAD1/5/8, mRNA expression, histone acetyltransferases (HAT)/histone deacetylase (HDAC) activity, and acetylation of histone H3. Following alcohol exposure, phosphorylation of SMAD1/5/8 and HAT activities was increased to a significant extent, while histone H3 acetylation and expression of heart development-related genes were also increased. The said phenomenon influenced by alcohol was reverted upon dorsomorphin treatment to the cells without effecting HDAC activity. The data clearly identified that BMP-mediated histone H3 acetylation of heart development-related genes might be one of the possible cellular mechanisms to control alcohol-induced expression of heart development-related genes. Dorsomorphin, on the other hand, may modulate alcohol-induced hyperacetylation of histone H3 through BMP targeting, which could be a potential way to block CHD. Copyright © 2016 by the Research Society on Alcoholism.

  2. H3 Histone Tail Conformation within the Nucleosome and the Impact of K14 Acetylation Studied Using Enhanced Sampling Simulation

    Science.gov (United States)

    Ikebe, Jinzen; Sakuraba, Shun; Kono, Hidetoshi

    2016-01-01

    Acetylation of lysine residues in histone tails is associated with gene transcription. Because histone tails are structurally flexible and intrinsically disordered, it is difficult to experimentally determine the tail conformations and the impact of acetylation. In this work, we performed simulations to sample H3 tail conformations with and without acetylation. The results show that irrespective of the presence or absence of the acetylation, the H3 tail remains in contact with the DNA and assumes an α-helix structure in some regions. Acetylation slightly weakened the interaction between the tail and DNA and enhanced α-helix formation, resulting in a more compact tail conformation. We inferred that this compaction induces unwrapping and exposure of the linker DNA, enabling DNA-binding proteins (e.g., transcription factors) to bind to their target sequences. In addition, our simulation also showed that acetylated lysine was more often exposed to the solvent, which is consistent with the fact that acetylation functions as a post-translational modification recognition site marker. PMID:26967163

  3. Histone H3 phosphorylation in GBM: a new rational to guide the use of kinase inhibitors in anti-GBM therapy.

    Science.gov (United States)

    Pacaud, Romain; Cheray, Mathilde; Nadaradjane, Arulraj; Vallette, François M; Cartron, Pierre-François

    2015-01-01

    Histones post-translational modifications (PTMs) are crucial components of diverse processes that modulate chromatin. Among the histones PTMs, the histones phosphorylation appears such crucial since it plays a significant role into DNA repair structure, transcription and chromatin compaction during cell division and apoptosis. However, little is known about the prognostic value of the histone phosphorylation in human cancer. This point could be considerate such as an important gap in anti-cancer therapy since the use of adequate kinase inhibitors could remedy to the aberrant histone phosphorylation associated with a poor prognosis factor. To remedy at this situation, we analyzed the phosphorylation level of histone H3 at the residues T3, T6, S10, S28, Y41 and T45 in a collection of 42 glioblastoma multiformes (GBM). Our data indicated that the high level of pH3T6, pH3S10 and pH3Y41 are signatures associated with a poor prognosis of overall survival (OS) of GBM treated with the "temozolomide and irradiation standard" treatment of GBM (named TMZ+Irad treatment). Our data also showed that these signatures are correlated with the high activity of kinases already described as writers of the pH3T6, pH3S10 and pH3Y41 i.e. the PKC, Aurora-B and JAK2, respectively. Finally, our analysis revealed that the use of Enzastaurin, AZD1152, and AZD1480 abrogated the high level of pH3T6, pH3S10 and pH3Y41 while increasing the sensitivity to the "temozolomide and irradiation"-induced cell death. To conclude, it appears that this work provides biomarkers for patient stratification for a therapy including kinase inhibitors.

  4. Genome-wide alteration of histone H3K9 acetylation pattern in mouse offspring prenatally exposed to arsenic.

    Directory of Open Access Journals (Sweden)

    Andrea A Cronican

    Full Text Available Chronic exposure to arsenic in drinking water, especially in utero or perinatal exposure, can initiate neurological and cognitive dysfunction, as well as memory impairment. Several epidemiological studies have demonstrated cognitive and learning deficits in children with early exposure to low to moderate levels of arsenic, but pathogenic mechanisms or etiology for these deficits are poorly understood. Since in vivo studies show a role for histone acetylation in cognitive performance and memory formation, we examined if prenatal exposure to arsenic causes changes in the epigenomic landscape. We exposed C57Bl6/J mice to 100 μg/L arsenic in the drinking water starting 1 week before conception till birth and applied chromatin immunoprecipitation followed by high-throughput massive parallel sequencing (ChIP-seq to evaluate H3K9 acetylation pattern in the offspring of exposed and control mice. Arsenic exposure during embryonic life caused global hypo-acetylation at H3K9 and changes in functional annotation with highly significant representation of Krüppel associated box (KRAB transcription factors in brain samples from exposed pups. We also found that arsenic exposure of adult mice impaired spatial and episodic memory, as well as fear conditioning performance. This is the first study to demonstrate: a genome wide changes in H3K9 acetylation pattern in an offspring prenatally exposed to arsenic, and b a connection between moderate arsenic exposure and cognitive impairment in adult mice. The results also emphasize the applicability of Next Generation Sequencing methodology in studies aiming to reveal the role of environmental factors, other than dietary restriction, in developmental reprogramming through histone modifications during embryonic development.

  5. Multilocus loss of DNA methylation in individuals with mutations in the histone H3 Lysine 4 Demethylase KDM5C

    Directory of Open Access Journals (Sweden)

    Grafodatskaya Daria

    2013-01-01

    Full Text Available Abstract Background A number of neurodevelopmental syndromes are caused by mutations in genes encoding proteins that normally function in epigenetic regulation. Identification of epigenetic alterations occurring in these disorders could shed light on molecular pathways relevant to neurodevelopment. Results Using a genome-wide approach, we identified genes with significant loss of DNA methylation in blood of males with intellectual disability and mutations in the X-linked KDM5C gene, encoding a histone H3 lysine 4 demethylase, in comparison to age/sex matched controls. Loss of DNA methylation in such individuals is consistent with known interactions between DNA methylation and H3 lysine 4 methylation. Further, loss of DNA methylation at the promoters of the three top candidate genes FBXL5, SCMH1, CACYBP was not observed in more than 900 population controls. We also found that DNA methylation at these three genes in blood correlated with dosage of KDM5C and its Y-linked homologue KDM5D. In addition, parallel sex-specific DNA methylation profiles in brain samples from control males and females were observed at FBXL5 and CACYBP. Conclusions We have, for the first time, identified epigenetic alterations in patient samples carrying a mutation in a gene involved in the regulation of histone modifications. These data support the concept that DNA methylation and H3 lysine 4 methylation are functionally interdependent. The data provide new insights into the molecular pathogenesis of intellectual disability. Further, our data suggest that some DNA methylation marks identified in blood can serve as biomarkers of epigenetic status in the brain.

  6. PfSETvs methylation of histone H3K36 represses virulence genes in Plasmodium falciparum

    DEFF Research Database (Denmark)

    Jiang, Lubin; Mu, Jianbing; Zhang, Qingfeng

    2013-01-01

    The variant antigen Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1), which is expressed on the surface of P. falciparum-infected red blood cells, is a critical virulence factor for malaria. Each parasite has 60 antigenically distinct var genes that each code for a different PfEMP1...... parasite nuclei and their expression as proteins on the surface of individual infected red blood cells. PfSETvs-dependent H3K36me3 is present along the entire gene body, including the transcription start site, to silence var genes. With low occupancy of PfSETvs at both the transcription start site of var...

  7. Pre-analytical variables of circulating cell-free nucleosomes containing 5-methylcytosine DNA or histone modification H3K9Me3

    DEFF Research Database (Denmark)

    Rasmussen, Louise; Herzog, Marielle; Rømer, Eva

    2016-01-01

    Aim: To evaluate pre-analytical variables of circulating cell-free nucleosomes containing 5-methylcytosine DNA (5mC) or histone modification H3K9Me3 (H3K9Me3). Materials and methods: Six studies were designed to assess the possible influence of pre-analytical variables. Study 1: influence of stas...

  8. Genome-wide analysis of histone H3 acetylation patterns in AML identifies PRDX2 as an epigenetically silenced tumor suppressor gene

    DEFF Research Database (Denmark)

    Agrawal-Singh, Shuchi; Isken, Fabienne; Agelopoulos, Konstantin

    2012-01-01

    With the use of ChIP on microarray assays in primary leukemia samples, we report that acute myeloid leukemia (AML) blasts exhibit significant alterations in histone H3 acetylation (H3Ac) levels at > 1000 genomic loci compared with CD34+ progenitor cells. Importantly, core promoter regions tended ...

  9. [Ash2, a subunit of histone H3K4 methyltransferase complex, is involved in the sporulation in Schizosaccharomyces pombe].

    Science.gov (United States)

    Wang, Wenchao; Zhou, Huan; Yu, Yao; Lv, Hong

    2014-09-01

    Schizosaccharomyces pombe undergoes meiosis instead of mitosis under conditions of nitrogen starvation and pheromone signalling, which results in conjugation and sporulation. During this progress, the pheromone-responsive MAPK(Mitogen-activated protein kinases) pathway plays an important role in regulating the conjuation and the transcriptional activation of genes required for meiosis. Spk1, a key component of MAPK pathway, activates Ste11 through protein phosphorylation and then induced the transcriptions of several genes requied for meiosis, including mei2(+), mam2(+) and map3(+). Methylation of histone H3K4 is involved in several important biological processes, including transcriptional activation and chromatin remodeling. However, its role in the sporualtion of fission yeast is poorly understood. Ash2 is a subunit of COMPASS, a conserved H3K4 methyltransferase complex. Sequence alignment analysis revealed that Ash2 in pombe shares two conserved domain with other homologues. Ash2 is localized in nucleus and contributes to methylation of H3K4. Deletion of ash2(+) resulted in a delay of sporulation and a substantial drop of sporulation efficiency. ChIP and qPCR analysis showed that deletion of ash2(+) caused a reduction of H3K4me2 level in the coding region of spk1(+), as well as a reduction of its mRNA level. Although the mRNA level of ste11(+) kept unchanged, the levels of Ste11-targetted genes, such as mei2(+), mam2(+) and map3(+), all reduced in ash2Δ cells. The results suggest that Ash2 regulates MAPK pathway and sporulation through H3K4 methylation. This might provide a new clue to elucidate the link between meiosis and epigenetic regulation.

  10. The ASH1 HOMOLOG 2 (ASHH2 histone H3 methyltransferase is required for ovule and anther development in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Paul E Grini

    Full Text Available BACKGROUND: SET-domain proteins are histone lysine (K methyltransferases (HMTase implicated in defining transcriptionally permissive or repressive chromatin. The Arabidopsis ASH1 HOMOLOG 2 (ASHH2 protein (also called SDG8, EFS and CCR1 has been suggested to methylate H3K4 and/or H3K36 and is similar to Drosophila ASH1, a positive maintainer of gene expression, and yeast Set2, a H3K36 HMTase. Mutation of the ASHH2 gene has pleiotropic developmental effects. Here we focus on the role of ASHH2 in plant reproduction. METHODOLOGY/PRINCIPAL FINDINGS: A slightly reduced transmission of the ashh2 allele in reciprocal crosses implied involvement in gametogenesis or gamete function. However, the main requirement of ASHH2 is sporophytic. On the female side, close to 80% of mature ovules lack embryo sac. On the male side, anthers frequently develop without pollen sacs or with specific defects in the tapetum layer, resulting in reduction in the number of functional pollen per anther by up to approximately 90%. In consistence with the phenotypic findings, an ASHH2 promoter-reporter gene was expressed at the site of megaspore mother cell formation as well as tapetum layers and pollen. ashh2 mutations also result in homeotic changes in floral organ identity. Transcriptional profiling identified more than 300 up-regulated and 600 down-regulated genes in ashh2 mutant inflorescences, whereof the latter included genes involved in determination of floral organ identity, embryo sac and anther/pollen development. This was confirmed by real-time PCR. In the chromatin of such genes (AP1, AtDMC1 and MYB99 we observed a reduction of H3K36 trimethylation (me3, but not H3K4me3 or H3K36me2. CONCLUSIONS/SIGNIFICANCE: The severe distortion of reproductive organ development in ashh2 mutants, argues that ASHH2 is required for the correct expression of genes essential to reproductive development. The reduction in the ashh2 mutant of H3K36me3 on down-regulated genes relevant to

  11. Mitotic accumulation of dimethylated lysine 79 of histone H3 is important for maintaining genome integrity during mitosis in human cells.

    Science.gov (United States)

    Guppy, Brent J; McManus, Kirk J

    2015-02-01

    The loss of genome stability is an early event that drives the development and progression of virtually all tumor types. Recent studies have revealed that certain histone post-translational modifications exhibit dynamic and global increases in abundance that coincide with mitosis and exhibit essential roles in maintaining genomic stability. Histone H2B ubiquitination at lysine 120 (H2Bub1) is regulated by RNF20, an E3 ubiquitin ligase that is altered in many tumor types. Through an evolutionarily conserved trans-histone pathway, H2Bub1 is an essential prerequisite for subsequent downstream dimethylation events at lysines 4 (H3K4me2) and 79 (H3K79me2) of histone H3. Although the role that RNF20 plays in tumorigenesis has garnered much attention, the downstream components of the trans-histone pathway, H3K4me2 and H3K79me2, and their potential contributions to genome stability remain largely overlooked. In this study, we employ single-cell imaging and biochemical approaches to investigate the spatial and temporal patterning of RNF20, H2Bub1, H3K4me2, and H3K79me2 throughout the cell cycle, with a particular focus on mitosis. We show that H2Bub1, H3K4me2, and H3K79me2 exhibit distinct temporal progression patterns throughout the cell cycle. Most notably, we demonstrate that H3K79me2 is a highly dynamic histone post-translational modification that reaches maximal abundance during mitosis in an H2Bub1-independent manner. Using RNAi and chemical genetic approaches, we identify DOT1L as a histone methyltransferase required for the mitotic-associated increases in H3K79me2. We also demonstrate that the loss of mitotic H3K79me2 levels correlates with increases in chromosome numbers and increases in mitotic defects. Collectively, these data suggest that H3K79me2 dynamics during mitosis are normally required to maintain genome stability and further implicate the loss of H3K79me2 during mitosis as a pathogenic event that contributes to the development and progression of tumors

  12. Unabridged Analysis of Human Histone H3 by Differential Top-Down Mass Spectrometry Reveals Hypermethylated Proteoforms from MMSET/NSD2 Overexpression.

    Science.gov (United States)

    Zheng, Yupeng; Fornelli, Luca; Compton, Philip D; Sharma, Seema; Canterbury, Jesse; Mullen, Christopher; Zabrouskov, Vlad; Fellers, Ryan T; Thomas, Paul M; Licht, Jonathan D; Senko, Michael W; Kelleher, Neil L

    2016-03-01

    Histones, and their modifications, are critical components of cellular programming and epigenetic inheritance. Recently, cancer genome sequencing has uncovered driver mutations in chromatin modifying enzymes spurring high interest how such mutations change histone modification patterns. Here, we applied Top-Down mass spectrometry for the characterization of combinatorial modifications (i.e. methylation and acetylation) on full length histone H3 from human cell lines derived from multiple myeloma patients with overexpression of the histone methyltransferase MMSET as the result of a t(4;14) chromosomal translocation. Using the latest in Orbitrap-based technology for clean isolation of isobaric proteoforms containing up to 10 methylations and/or up to two acetylations, we provide extensive characterization of histone H3.1 and H3.3 proteoforms. Differential analysis of modifications by electron-based dissociation recapitulated antagonistic crosstalk between K27 and K36 methylation in H3.1, validating that full-length histone H3 (15 kDa) can be analyzed with site-specific assignments for multiple modifications. It also revealed K36 methylation in H3.3 was affected less by the overexpression of MMSET because of its higher methylation levels in control cells. The co-occurrence of acetylation with a minimum of three methyl groups in H3K9 and H3K27 suggested a hierarchy in the addition of certain modifications. Comparative analysis showed that high levels of MMSET in the myeloma-like cells drove the formation of hypermethyled proteoforms containing H3K36me2 co-existent with the repressive marks H3K9me2/3 and H3K27me2/3. Unique histone proteoforms with such "trivalent hypermethylation" (K9me2/3-K27me2/3-K36me2) were not discovered when H3.1 peptides were analyzed by Bottom-Up. Such disease-correlated proteoforms could link tightly to aberrant transcription programs driving cellular proliferation, and their precise description demonstrates that Top-Down mass spectrometry can

  13. Inhalable Metal-Rich Air Particles and Histone H3K4 Dimethylation and H3K9 Acetylation in a Cross-sectional Study of Steel Workers

    National Research Council Canada - National Science Library

    Laura Cantone; Francesco Nordio; Lifang Hou; Pietro Apostoli; Matteo Bonzini; Letizia Tarantini; Laura Angelici; Valentina Bollati; Antonella Zanobetti; Joel Schwartz; Pier A. Bertazzi; Andrea Baccarelli

    2011-01-01

    ...) with increased risk of lung cancer. PM contains carcinogenic and toxic metals, including arsenic and nickel, which have been shown in in vitro studies to induce histone modifications that activate gene expression by inducing open-chromatin states...

  14. Histone Crosstalk between H3S10ph and H4K16ac Generates a Histone Code that Mediates Transcription Elongation

    National Research Council Canada - National Science Library

    Zippo, Alessio; Serafini, Riccardo; Rocchigiani, Marina; Pennacchini, Susanna; Krepelova, Anna; Oliviero, Salvatore

    2009-01-01

    .... This model is based on the histone code hypothesis, where specific histone modifications, acting alone or in combination, can promote or inhibit the binding of a protein (the “reader”) to the nucleosome ( Strahl and Allis, 2000 ). The reader can promote a second histone modification, determining a histone crosstalk which generates a different binding platform ...

  15. Quantitative mass spectrometry of histones H3.2 and H3.3 in Suz12-deficient mouse embryonic stem cells reveals distinct, dynamic post-translational modifications at Lys-27 and Lys-36

    DEFF Research Database (Denmark)

    Jung, Hye Ryung; Pasini, Diego; Helin, Kristian

    2010-01-01

    SUZ12 is a core component of the polycomb repressive complex 2 (PRC2) and is required for the differentiation of mouse embryonic stem cells (ESCs). PRC2 is associated with transcriptional repression via methylation of H3 Lys-27. We applied quantitative mass spectrometry to investigate the effects....... The combined use of ETD and CID MS/MS increased the total number of identified modified peptides. Comparative quantitative analysis of histones from wild type and Suz12-deficient ESCs using stable isotope labeling with amino acids in cell culture and LC-MS/MS revealed a dramatic reduction of H3K27me2 and H3K27...

  16. Dual modified antiphospho (Ser10)-acetyl (Lys14)-histone H3 predominantly mark the pericentromeric chromatin during mitosis in monokinetic plants.

    Science.gov (United States)

    Sharma, Santosh Kumar; Yamamoto, Maki; Mukai, Yasuhiko

    2016-12-01

    Epigenetic regulatory posttranslational histone modification marks not only function individually but also capable to act in combination as a unique pattern. A total of 16 plant species belonging to 11 genera of eight families (five dicots and three monocots) including land plants, epiphytes (orchids) and the holokinetic taxa (Drosera spp.) were analysed for chromosomal distribution of dual modified antiphospho (Ser10)-acetyl (K14)-histone H3 (H3S10phK14ac) to understand the combinatorial chromatin dynamics during mitotic cell division in plants. The anti-H3S10phK14ac evidently mark the pericentromeric chromatin on mitotic chromosomes of the plants excluding the holokinetic Drosera species, which revealed the immunolabelling of whole chromosomes all along the arms. The dual modified immunosignals were absent during early stages of mitosis, appeared intensively at metaphase and remained visible until late-anaphase/telophase however, labelled the whole chromosomes during meiotic metaphase I. Colocalization of anti-H3S10phK14ac with an onion's CENH3 antibody on mitotic chromosomes of Allium revealed the chromosomal location of anti-H3S10phK14ac in the region between signals for CENH3 detection. Overall analysis suggests that the unique localization of combinatorial histone modification mark at pericentromeric chromatin might have attributed through 'phospho-acetyl' cross talk that ultimately facilitate the sister chromatid cohesion at pericentromeres following condensation events in mitotic chromosomes. Here, we propose that dual modified H3S10phK14ac histone may serve as an additional cytogenetic landmark to identify pericentromeric chromatin during mitosis in plants. The plausible role of histone cross talk and future perspectives of combinatorial histone modification marks in plant cytogenetics with special reference to chromatin dynamics have been discussed.

  17. The Histone-H3K4-Specific Demethylase KDM5B Binds to Its Substrate and Product through Distinct PHD Fingers

    Directory of Open Access Journals (Sweden)

    Brianna J. Klein

    2014-01-01

    Full Text Available The histone lysine demethylase KDM5B regulates gene transcription and cell differentiation and is implicated in carcinogenesis. It contains multiple conserved chromatin-associated domains, including three PHD fingers of unknown function. Here, we show that the first and third, but not the second, PHD fingers of KDM5B possess histone binding activities. The PHD1 finger is highly specific for unmodified histone H3 (H3K4me0, whereas the PHD3 finger shows preference for the trimethylated histone mark H3K4me3. RNA-seq analysis indicates that KDM5B functions as a transcriptional repressor for genes involved in inflammatory responses, cell proliferation, adhesion, and migration. Biochemical analysis reveals that KDM5B associates with components of the nucleosome remodeling and deacetylase (NuRD complex and may cooperate with the histone deacetylase 1 (HDAC1 in gene repression. KDM5B is downregulated in triple-negative breast cancer relative to estrogen-receptor-positive breast cancer. Overexpression of KDM5B in the MDA-MB 231 breast cancer cells suppresses cell migration and invasion, and the PHD1-H3K4me0 interaction is essential for inhibiting migration. These findings highlight tumor-suppressive functions of KDM5B in triple-negative breast cancer cells and suggest a multivalent mechanism for KDM5B-mediated transcriptional regulation.

  18. A role for WDR5 in integrating threonine 11 phosphorylation to lysine 4 methylation on histone H3 during androgen signaling and in prostate cancer.

    Science.gov (United States)

    Kim, Ji-Young; Banerjee, Taraswi; Vinckevicius, Aurimas; Luo, Qianyi; Parker, J Brandon; Baker, Mairead R; Radhakrishnan, Ishwar; Wei, Jian-Jun; Barish, Grant D; Chakravarti, Debabrata

    2014-05-22

    Upon androgen stimulation, PKN1-mediated histone H3 threonine 11 phosphorylation (H3T11P) promotes AR target gene activation. However, the underlying mechanism is not completely understood. Here, we show that WDR5, a subunit of the SET1/MLL complex, interacts with H3T11P, and this interaction facilitates the recruitment of the MLL1 complex and subsequent H3K4 tri-methylation (H3K4me3). Using ChIP-seq, we find that androgen stimulation results in a 6-fold increase in the number of H3T11P-marked regions and induces WDR5 colocalization to one third of H3T11P-enriched promoters, thus establishing a genome-wide relationship between H3T11P and recruitment of WDR5. Accordingly, PKN1 knockdown or chemical inhibition severely blocks WDR5 chromatin association and H3K4me3 on AR target genes. Finally, WDR5 is critical in prostate cancer cell proliferation and is hyperexpressed in human prostate cancers. Together, these results identify WDR5 as a critical epigenomic integrator of histone phosphorylation and methylation and as a major driver of androgen-dependent prostate cancer cell proliferation. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. The amino-terminal tails of histones H2A and H3 coordinate efficient base excision repair, DNA damage signaling and postreplication repair in Saccharomyces cerevisiae

    Science.gov (United States)

    Meas, Rithy; Smerdon, Michael J.; Wyrick, John J.

    2015-01-01

    Histone amino-terminal tails (N-tails) are required for cellular resistance to DNA damaging agents; therefore, we examined the role of histone N-tails in regulating DNA damage response pathways in Saccharomyces cerevisiae. Combinatorial deletions reveal that the H2A and H3 N-tails are important for the removal of MMS-induced DNA lesions due to their role in regulating the basal and MMS-induced expression of DNA glycosylase Mag1. Furthermore, overexpression of Mag1 in a mutant lacking the H2A and H3 N-tails rescues base excision repair (BER) activity but not MMS sensitivity. We further show that the H3 N-tail functions in the Rad9/Rad53 DNA damage signaling pathway, but this function does not appear to be the primary cause of MMS sensitivity of the double tailless mutants. Instead, epistasis analyses demonstrate that the tailless H2A/H3 phenotypes are in the RAD18 epistasis group, which regulates postreplication repair. We observed increased levels of ubiquitylated PCNA and significantly lower mutation frequency in the tailless H2A/H3 mutant, indicating a defect in postreplication repair. In summary, our data identify novel roles of the histone H2A and H3 N-tails in (i) regulating the expression of a critical BER enzyme (Mag1), (ii) supporting efficient DNA damage signaling and (iii) facilitating postreplication repair. PMID:25897129

  20. Upregulation of KLF4 by methylseleninic acid in human esophageal squamous cell carcinoma cells: Modification of histone H3 acetylation through HAT/HDAC interplay.

    Science.gov (United States)

    Hu, Chenfei; Liu, Mei; Zhang, Wei; Xu, Qing; Ma, Kai; Chen, Lechuang; Wang, Zaozao; He, Shun; Zhu, Hongxia; Xu, Ningzhi

    2015-10-01

    Esophageal squamous cell carcinoma (ESCC) occurs at a very high frequency in certain areas of China. Supplementation with selenium-containing compounds was associated with a significantly lower cancer mortality rate in a study conducted in Linxia, China. Thus, selenium could be a potential anti-esophageal cancer agent. In this study, methylseleninic acid (MSA) could inhibit cell growth of ESCC cells in vitro and in vivo. Upon treated with MSA, the activity of histone deacetylases (HDACs) was decreased and general control nonrepressed protein 5 (GCN5) was upregulated in ESCC cells. Meanwhile, a significant increase of H3K9 acetylation (H3K9ac) was detected. Upregulation of Krüppel-like factor 4 (KLF4) was also observed after MSA treatment. Additionally, the acetylated histone H3 located more at KLF4 promoter region after MSA treatment, shown by chromatin immunoprecipitation (ChIP) assay. Moreover, knockdown of GCN5 decreased the protein level of both H3K9ac and KLF4, along with less cell growth inhibition. Taken all, our results indicated that MSA could inhibit ESCC cell growth, at least in part, by MSA-HDAC/GCN5-H3K9ac-KLF4 axis. To our best knowledge, this is the first report that MSA induced acetylation of histone H3 at Lys9, which might depend on the activities and the balance between HDACs and HATs. © 2014 Wiley Periodicals, Inc.

  1. Hypoxia and nickel inhibit histone demethylase JMJD1A and repress Spry2 expression in human bronchial epithelial BEAS-2B cells

    OpenAIRE

    Chen, Haobin; Kluz, Thomas; Zhang, Ronghe; Costa, Max

    2010-01-01

    Epigenetic silencing of tumor suppressor genes commonly occurs in human cancers via increasing DNA methylation and repressive histone modifications at gene promoters. However, little is known about how pathogenic environmental factors contribute to cancer development by affecting epigenetic regulatory mechanisms. Previously, we reported that both hypoxia and nickel (an environmental carcinogen) increased global histone H3 lysine 9 methylation in cells through inhibiting a novel class of iron-...

  2. The H3K4me3/2 histone demethylase RBR-2 controls axon guidance by repressing the actin-remodeling gene wsp-1

    DEFF Research Database (Denmark)

    Mariani, Luca; Lussi, Yvonne C.; Vandamme, Julien

    2016-01-01

    The dynamic regulation of histone modifications is important for modulating transcriptional programs during development. Aberrant H3K4 methylation is associated with neurological disorders, but how the levels and the recognition of this modification affect specific neuronal processes is unclear...

  3. Histone 3.3 hotspot mutations in conventional osteosarcomas: a comprehensive clinical and molecular characterization of six H3F3A mutated cases.

    NARCIS (Netherlands)

    Koelsche, C.; Schrimpf, D.; Tharun, L.; Roth, E.; Sturm, D.; Jones, D.T.; Renker, E.K.; Sill, M.; Baude, A.; Sahm, F.; Capper, D.; Bewerunge-Hudler, M.; Hartmann, W.; Kulozik, A.E.; Petersen, I.; Flucke, U.E.; Schreuder, H.W.B.; Buttner, R.; Weber, M.A.; Schirmacher, P.; Plass, C.; Pfister, S.M.; Deimling, A. von; Mechtersheimer, G.

    2017-01-01

    BACKGROUND: Histone 3.3 (H3.3) hotspot mutations in bone tumors occur in the vast majority of giant cell tumors of bone (GCTBs; 96%), chondroblastomas (95%) and in a few cases of osteosarcomas. However, clinical presentation, histopathological features, and additional molecular characteristics of

  4. The H3K4me3/2 histone demethylase RBR-2 controls axon guidance by repressing the actin-remodeling gene wsp-1

    DEFF Research Database (Denmark)

    Mariani, Luca; Lussi, Yvonne C.; Vandamme, Julien

    2016-01-01

    The dynamic regulation of histone modifications is important for modulating transcriptional programs during development. Aberrant H3K4 methylation is associated with neurological disorders, but how the levels and the recognition of this modification affect specific neuronal processes is unclear. ...

  5. Dimethylated H3K27 Is a Repressive Epigenetic Histone Mark in the Protist Entamoeba histolytica and Is Significantly Enriched in Genes Silenced via the RNAi Pathway.

    Science.gov (United States)

    Foda, Bardees M; Singh, Upinder

    2015-08-21

    RNA interference (RNAi) is a fundamental biological process that plays a crucial role in regulation of gene expression in many organisms. Transcriptional gene silencing (TGS) is one of the important nuclear roles of RNAi. Our previous data show that Entamoeba histolytica has a robust RNAi pathway that links to TGS via Argonaute 2-2 (Ago2-2) associated 27-nucleotide small RNAs with 5'-polyphosphate termini. Here, we report the first repressive histone mark to be identified in E. histolytica, dimethylation of H3K27 (H3K27Me2), and demonstrate that it is enriched at genes that are silenced by RNAi-mediated TGS. An RNAi-silencing trigger can induce H3K27Me2 deposits at both episomal and chromosomal loci, mediating gene silencing. Our data support two phases of RNAi-mediated TGS: an active silencing phase where the RNAi trigger is present and both H3K27Me2 and Ago2-2 concurrently enrich at chromosomal loci; and an established silencing phase in which the RNAi trigger is removed, but gene silencing with H3K27Me2 enrichment persist independently of Ago2-2 deposition. Importantly, some genes display resistance to chromosomal silencing despite induction of functional small RNAs. In those situations, the RNAi-triggering plasmid that is maintained episomally gets partially silenced and has H3K27Me2 enrichment, but the chromosomal copy displays no repressive histone enrichment. Our data are consistent with a model in which H3K27Me2 is a repressive histone modification, which is strongly associated with transcriptional repression. This is the first example of an epigenetic histone modification that functions to mediate RNAi-mediated TGS in the deep-branching eukaryote E. histolytica. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Dimethylated H3K27 Is a Repressive Epigenetic Histone Mark in the Protist Entamoeba histolytica and Is Significantly Enriched in Genes Silenced via the RNAi Pathway*

    Science.gov (United States)

    Foda, Bardees M.; Singh, Upinder

    2015-01-01

    RNA interference (RNAi) is a fundamental biological process that plays a crucial role in regulation of gene expression in many organisms. Transcriptional gene silencing (TGS) is one of the important nuclear roles of RNAi. Our previous data show that Entamoeba histolytica has a robust RNAi pathway that links to TGS via Argonaute 2-2 (Ago2-2) associated 27-nucleotide small RNAs with 5′-polyphosphate termini. Here, we report the first repressive histone mark to be identified in E. histolytica, dimethylation of H3K27 (H3K27Me2), and demonstrate that it is enriched at genes that are silenced by RNAi-mediated TGS. An RNAi-silencing trigger can induce H3K27Me2 deposits at both episomal and chromosomal loci, mediating gene silencing. Our data support two phases of RNAi-mediated TGS: an active silencing phase where the RNAi trigger is present and both H3K27Me2 and Ago2-2 concurrently enrich at chromosomal loci; and an established silencing phase in which the RNAi trigger is removed, but gene silencing with H3K27Me2 enrichment persist independently of Ago2-2 deposition. Importantly, some genes display resistance to chromosomal silencing despite induction of functional small RNAs. In those situations, the RNAi-triggering plasmid that is maintained episomally gets partially silenced and has H3K27Me2 enrichment, but the chromosomal copy displays no repressive histone enrichment. Our data are consistent with a model in which H3K27Me2 is a repressive histone modification, which is strongly associated with transcriptional repression. This is the first example of an epigenetic histone modification that functions to mediate RNAi-mediated TGS in the deep-branching eukaryote E. histolytica. PMID:26149683

  7. Comparative analysis of histone H3 and H4 post-translational modifications of esophageal squamous cell carcinoma with different invasive capabilities.

    Science.gov (United States)

    Zhang, Kai; Li, Liyan; Zhu, Mengxiao; Wang, Guojuan; Xie, Jianjun; Zhao, Yunlong; Fan, Enguo; Xu, Liyan; Li, Enmin

    2015-01-01

    Eukaryotic DNA is packaged into a chromatin with the help of four core histones (H2A, H2B, H3, and H4). Diverse histone post-translational modifications (PTMs) are hence involved in the regulation of gene transcription. However, how this regulation does work is still poorly understood and lacks details. Here we used the mass spectrometry-based proteomics approach to perform a comparative analysis of histone marks at a global level in two phenotypes of esophageal squamous cell carcinoma (ESCC) with different invasiveness. We obtained a comprehensive profiling of histone H3 and H4 PTMs including lysine methylation, acetylation and novel butyrylation. The correlation between histone marks and cancer invasive capabilities was further characterized and one distinguishable PTM, H4K79me2 was discovered and verified in this study. Immunohistochemistry analysis suggests that abnormal level of H4K79me2 may be related to poor survival of ESCC patients. Our results enrich the dataset of the feature pattern of global histone PTMs in ESCC cell lines. Core histone proteins, decorated by multiple biological significant protein post-translational modifications (PTMs) such as lysine acetylation and lysine methylation, are considered to regulate gene transcription and be associated with the development of cancer. Recent studies have further shown that global level of histone modifications is the potential hallmark of cancer to predict the clinical outcomes of human cancers. However, the regulation mechanism is largely unknown. Here we used the mass spectrometry based proteomics coupled with stable isotope labeling with amino acids in cell culture (SILAC) to characterize the global levels of histone marks in two phenotypes of esophageal squamous cell carcinoma (ESCC) cell lines with different invasive capabilities. To the best of our knowledge, it is the first report about the comparative analysis for histone marks of the different invasive ESCC cell lines. A significantly

  8. A time-series analysis of altered histone H3 acetylation and gene expression during the course of MMAIII-induced malignant transformation of urinary bladder cells.

    Science.gov (United States)

    Zhu, Jinqiu; Wang, Jie; Chen, Xushen; Tsompana, Maria; Gaile, Daniel; Buck, Michael; Ren, Xuefeng

    2017-04-01

    Our previous studies have shown that chronic exposure to low doses of monomethylarsonous acid (MMAIII) causes global histone acetylation dysregulation in urothelial cells (UROtsa cells) during the course of malignant transformation. To reveal the relationship between altered histone acetylation patterns and aberrant gene expression, more specifically, the carcinogenic relevance of these alterations, we performed a time-course analysis of the binding patterns of histone 3 lysine 18 acetylation (H3K18ac) across the genome and generated global gene-expression profiles from this UROtsa cell malignant transformation model. We showed that H3K18ac, one of the most significantly upregulated histone acetylation sites following MMAIII exposure, was enriched at gene promoter-specific regions across the genome and that MMAIII-induced upregulation of H3K18ac led to an altered binding pattern in a large number of genes that was most significant during the critical window for MMAIII-induced UROtsa cells' malignant transformation. Some genes identified as having a differential binding pattern with H3K18ac, acted as upstream regulators of critical gene networks with known functions in tumor development and progression. The altered H3K18ac binding patterns not only led to changes in expression of these directly affected upstream regulators but also resulted in gene-expression changes in their regulated networks. Collectively, our data suggest that MMAIII-induced alteration of histone acetylation patterns in UROtsa cells led to a time- and malignant stage-dependent aberrant gene-expression pattern, and that some gene regulatory networks were altered in accordance with their roles in carcinogenesis, probably contributing to MMAIII-induced urothelial cell malignant transformation and carcinogenesis. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  9. The histone demethylase Jarid1b ensures faithful mouse development by protecting developmental genes from aberrant H3K4me3.

    Directory of Open Access Journals (Sweden)

    Mareike Albert

    2013-04-01

    Full Text Available Embryonic development is tightly regulated by transcription factors and chromatin-associated proteins. H3K4me3 is associated with active transcription and H3K27me3 with gene repression, while the combination of both keeps genes required for development in a plastic state. Here we show that deletion of the H3K4me2/3 histone demethylase Jarid1b (Kdm5b/Plu1 results in major neonatal lethality due to respiratory failure. Jarid1b knockout embryos have several neural defects including disorganized cranial nerves, defects in eye development, and increased incidences of exencephaly. Moreover, in line with an overlap of Jarid1b and Polycomb target genes, Jarid1b knockout embryos display homeotic skeletal transformations typical for Polycomb mutants, supporting a functional interplay between Polycomb proteins and Jarid1b. To understand how Jarid1b regulates mouse development, we performed a genome-wide analysis of histone modifications, which demonstrated that normally inactive genes encoding developmental regulators acquire aberrant H3K4me3 during early embryogenesis in Jarid1b knockout embryos. H3K4me3 accumulates as embryonic development proceeds, leading to increased expression of neural master regulators like Pax6 and Otx2 in Jarid1b knockout brains. Taken together, these results suggest that Jarid1b regulates mouse development by protecting developmental genes from inappropriate acquisition of active histone modifications.

  10. A lesson learned from the H3.3K27M mutation found in pediatric glioma: a new approach to the study of the function of histone modifications in vivo?

    Science.gov (United States)

    Chan, Kui Ming; Han, Jing; Fang, Dong; Gan, Haiyun; Zhang, Zhiguo

    2013-08-15

    Glioblastoma (GBM) is the most aggressive primary brain tumor in human. Recent studies on high-grade pediatric GBM have identified two recurrent mutations (K27M and G34R/V) in genes encoding histone H3 (H3F3A for H3.3 and HIST1H3B for H3.1). The two histone H3 mutations are mutually exclusive and give rise to tumors in different brain compartments. Recently, we and others have shown that the histone H3 K27M mutation specifically altered the di- and tri-methylation of endogenous histone H3 at Lys27. Genome-wide studies using ChIP-seq on H3.3K27M patient samples indicate a global reduction of H3K27me3 on chromatin. Remarkably, we also found a dramatic enrichment of H3K27me3 and EZH2 (the catalytic subunit H3K27 methyltransferase) at hundreds of gene loci in H3.3K27M patient cells. Here, we discuss potential mechanisms whereby H3K27me3 is enriched at chromatin loci in cells expressing the H3.3K27M mutation and report effects of Lys-to-Met mutations of other well-studied lysine residues of histone H3.1/H3.3 and H4 on the corresponding endogenous lysine methylation. We suggest that mutation(s) on histones may be found in a variety of human diseases, and the expression of mutant histones may help to address the function of histone lysine methylation and possibly other modifications in mammalian cells.

  11. Differential regulation of the phosphorylation of Trimethyl-lysine27 histone H3 at serine 28 in distinct populations of striatal projection neurons

    DEFF Research Database (Denmark)

    Bonito-Oliva, Alessandra; Södersten, Erik; Spigolon, Giada

    2016-01-01

    Phosphorylation of histone H3 (H3) on serine 28 (S28) at genomic regions marked by trimethylation of lysine 27 (H3K27me3) often correlates with increased expression of genes normally repressed by Polycomb group proteins (PcG). We show that amphetamine, an addictive psychostimulant, and haloperidol...... of the protein phosphatase-1 inhibitor, dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32), reduces the phosphorylation of H3K27me3S28 produced by amphetamine and haloperidol. In contrast, knockout of the mitogen- and stress activated kinase 1 (MSK1), which is implicated in the phosphorylation...... of histone H3, decreases the effect of amphetamine, but not that of haloperidol. Chromatin immunoprecipitation analysis shows that amphetamine and haloperidol increase the phosphorylation of H3K27me3S28 at the promoter regions of Atf3, Npas4 and Lipg, three genes repressed by PcG. These results identify H3K...

  12. A TGFβ-PRMT5-MEP50 axis regulates cancer cell invasion through histone H3 and H4 arginine methylation coupled transcriptional activation and repression.

    Science.gov (United States)

    Chen, H; Lorton, B; Gupta, V; Shechter, D

    2017-01-19

    Protein arginine methyltransferase 5 (PRMT5) complexed with MEP50/WDR77 catalyzes arginine methylation on histones and other proteins. PRMT5-MEP50 activity is elevated in cancer cells and its expression is highly correlated with poor prognosis in many human tumors. We demonstrate that PRMT5-MEP50 is essential for transcriptional regulation promoting cancer cell invasive phenotypes in lung adenocarcinoma, lung squamous cell carcinoma and breast carcinoma cancer cells. RNA-Seq transcriptome analysis demonstrated that PRMT5 and MEP50 are required to maintain expression of metastasis and Epithelial-to-mesenchymal transition (EMT) markers and to potentiate an epigenetic mechanism of the TGFβ response. We show that PRMT5-MEP50 activity both positively and negatively regulates expression of a wide range of genes. Exogenous TGFβ promotes EMT in a unique pathway of PRMT5-MEP50 catalyzed histone mono- and dimethylation of chromatin at key metastasis suppressor and EMT genes, defining a new mechanism regulating cancer invasivity. PRMT5 methylation of histone H3R2me1 induced transcriptional activation by recruitment of WDR5 and concomitant H3K4 methylation at targeted genes. In parallel, PRMT5 methylation of histone H4R3me2s suppressed transcription at distinct genomic loci. Our decoding of histone methylarginine at key genes supports a critical role for complementary PRMT5-MEP50 transcriptional activation and repression in cancer invasion pathways and in response to TGFβ stimulation and therefore orients future chemotherapeutic opportunities.

  13. Chromatin binding of SRp20 and ASF/SF2 and dissociation from mitotic chromosomes is modulated by histone H3 serine 10 phosphorylation.

    Science.gov (United States)

    Loomis, Rebecca J; Naoe, Yoshinori; Parker, J Brandon; Savic, Velibor; Bozovsky, Matthew R; Macfarlan, Todd; Manley, James L; Chakravarti, Debabrata

    2009-02-27

    Histone H3 serine 10 phosphorylation is a hallmark of mitotic chromosomes, but its full function remains to be elucidated. We report here that two SR protein splicing factors, SRp20 and ASF/SF2, associate with interphase chromatin, are released from hyperphosphorylated mitotic chromosomes, but reassociate with chromatin late in M-phase. Inhibition of Aurora B kinase diminished histone H3 serine 10 phosphorylation and increased SRp20 and ASF/SF2 retention on mitotic chromosomes. Unexpectedly, we also found that HP1 proteins interact with ASF/SF2 in mitotic cells. Strikingly, siRNA-mediated knockdown of ASF/SF2 caused retention of HP1 proteins on mitotic chromatin. Finally, ASF/SF2-depleted cells released from a mitotic block displayed delayed G0/G1 entry, suggesting a functional consequence of these interactions. These findings underscore the evolving role of histone H3 phosphorylation and demonstrate a direct, functional, and histone-modification-regulated association of SRp20 and ASF/SF2 with chromatin.

  14. An Alternative Approach to ChIP-Seq Normalization Enables Detection of Genome-Wide Changes in Histone H3 Lysine 27 Trimethylation upon EZH2 Inhibition

    Science.gov (United States)

    Yuan, Chih-Chi; Craske, Madeleine Lisa; Labhart, Paul; Guler, Gulfem D.; Arnott, David; Maile, Tobias M.; Busby, Jennifer; Henry, Chisato; Kelly, Theresa K.; Tindell, Charles A.; Jhunjhunwala, Suchit; Zhao, Feng; Hatton, Charlie; Bryant, Barbara M.

    2016-01-01

    Chromatin immunoprecipitation and DNA sequencing (ChIP-seq) has been instrumental in inferring the roles of histone post-translational modifications in the regulation of transcription, chromatin compaction and other cellular processes that require modulation of chromatin structure. However, analysis of ChIP-seq data is challenging when the manipulation of a chromatin-modifying enzyme significantly affects global levels of histone post-translational modifications. For example, small molecule inhibition of the methyltransferase EZH2 reduces global levels of histone H3 lysine 27 trimethylation (H3K27me3). However, standard ChIP-seq normalization and analysis methods fail to detect a decrease upon EZH2 inhibitor treatment. We overcome this challenge by employing an alternative normalization approach that is based on the addition of Drosophila melanogaster chromatin and a D. melanogaster-specific antibody into standard ChIP reactions. Specifically, the use of an antibody that exclusively recognizes the D. melanogaster histone variant H2Av enables precipitation of D. melanogaster chromatin as a minor fraction of the total ChIP DNA. The D. melanogaster ChIP-seq tags are used to normalize the human ChIP-seq data from DMSO and EZH2 inhibitor-treated samples. Employing this strategy, a substantial reduction in H3K27me3 signal is now observed in ChIP-seq data from EZH2 inhibitor treated samples. PMID:27875550

  15. An Alternative Approach to ChIP-Seq Normalization Enables Detection of Genome-Wide Changes in Histone H3 Lysine 27 Trimethylation upon EZH2 Inhibition.

    Science.gov (United States)

    Egan, Brian; Yuan, Chih-Chi; Craske, Madeleine Lisa; Labhart, Paul; Guler, Gulfem D; Arnott, David; Maile, Tobias M; Busby, Jennifer; Henry, Chisato; Kelly, Theresa K; Tindell, Charles A; Jhunjhunwala, Suchit; Zhao, Feng; Hatton, Charlie; Bryant, Barbara M; Classon, Marie; Trojer, Patrick

    2016-01-01

    Chromatin immunoprecipitation and DNA sequencing (ChIP-seq) has been instrumental in inferring the roles of histone post-translational modifications in the regulation of transcription, chromatin compaction and other cellular processes that require modulation of chromatin structure. However, analysis of ChIP-seq data is challenging when the manipulation of a chromatin-modifying enzyme significantly affects global levels of histone post-translational modifications. For example, small molecule inhibition of the methyltransferase EZH2 reduces global levels of histone H3 lysine 27 trimethylation (H3K27me3). However, standard ChIP-seq normalization and analysis methods fail to detect a decrease upon EZH2 inhibitor treatment. We overcome this challenge by employing an alternative normalization approach that is based on the addition of Drosophila melanogaster chromatin and a D. melanogaster-specific antibody into standard ChIP reactions. Specifically, the use of an antibody that exclusively recognizes the D. melanogaster histone variant H2Av enables precipitation of D. melanogaster chromatin as a minor fraction of the total ChIP DNA. The D. melanogaster ChIP-seq tags are used to normalize the human ChIP-seq data from DMSO and EZH2 inhibitor-treated samples. Employing this strategy, a substantial reduction in H3K27me3 signal is now observed in ChIP-seq data from EZH2 inhibitor treated samples.

  16. MLL5 Orchestrates a Cancer Self-Renewal State by Repressing the Histone Variant H3.3 and Globally Reorganizing Chromatin.

    Science.gov (United States)

    Gallo, Marco; Coutinho, Fiona J; Vanner, Robert J; Gayden, Tenzin; Mack, Stephen C; Murison, Alex; Remke, Marc; Li, Ren; Takayama, Naoya; Desai, Kinjal; Lee, Lilian; Lan, Xiaoyang; Park, Nicole I; Barsyte-Lovejoy, Dalia; Smil, David; Sturm, Dominik; Kushida, Michelle M; Head, Renee; Cusimano, Michael D; Bernstein, Mark; Clarke, Ian D; Dick, John E; Pfister, Stefan M; Rich, Jeremy N; Arrowsmith, Cheryl H; Taylor, Michael D; Jabado, Nada; Bazett-Jones, David P; Lupien, Mathieu; Dirks, Peter B

    2015-12-14

    Mutations in the histone 3 variant H3.3 have been identified in one-third of pediatric glioblastomas (GBMs), but not in adult tumors. Here we show that H3.3 is a dynamic determinant of functional properties in adult GBM. H3.3 is repressed by mixed lineage leukemia 5 (MLL5) in self-renewing GBM cells. MLL5 is a global epigenetic repressor that orchestrates reorganization of chromatin structure by punctuating chromosomes with foci of compacted chromatin, favoring tumorigenic and self-renewing properties. Conversely, H3.3 antagonizes self-renewal and promotes differentiation. We exploited these epigenetic states to rationally identify two small molecules that effectively curb cancer stem cell properties in a preclinical model. Our work uncovers a role for MLL5 and H3.3 in maintaining self-renewal hierarchies in adult GBM. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Damaged DNA-binding protein down-regulates epigenetic mark H3K56Ac through histone deacetylase 1 and 2

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Qianzheng; Battu, Aruna; Ray, Alo; Wani, Gulzar; Qian, Jiang; He, Jinshan; Wang, Qi-en [Department of Radiology, The Ohio State University, Columbus, OH 43210 (United States); Wani, Altaf A., E-mail: wani.2@osu.edu [Department of Radiology, The Ohio State University, Columbus, OH 43210 (United States); Department of Molecular and Cellular Biochemistry, The Ohio State University, Columbus, OH 43210 (United States); James Cancer Hospital and Solove Research Institute, The Ohio State University, Columbus, OH 43210 (United States)

    2015-06-15

    Highlights: • HDAC1 and HDAC2 co-localize with UV radiation-induced DNA damage sites. • HDAC1 translocation to chromatin is dependent on DDB2 function. • HDAC1 and HDAC2 are involved in H3K56Ac deacetylation. • H3K56Ac deacetylation requires DDB1 and DDB2 but not XPA or XPC functions. • HDAC1/2 depletion decreases XPC ubiquitination and local γH2AX accumulation. - Abstract: Acetylated histone H3 lysine 56 (H3K56Ac) is one of the reversible histone post-translational modifications (PTMs) responsive to DNA damage. We previously described a biphasic decrease and increase of epigenetic mark H3K56Ac in response to ultraviolet radiation (UVR)-induced DNA damage. Here, we report a new function of UV damaged DNA-binding protein (DDB) in deacetylation of H3K56Ac through specific histone deacetylases (HDACs). We show that simultaneous depletion of HDAC1/2 compromises the deacetylation of H3K56Ac, while depletion of HDAC1 or HDAC2 alone has no effect on H3K56Ac. The H3K56Ac deacetylation does not require functional nucleotide excision repair (NER) factors XPA and XPC, but depends on the function of upstream factors DDB1 and DDB2. UVR enhances the association of DDB2 with HDAC1 and, enforced DDB2 expression leads to translocation of HDAC1 to UVR-damaged chromatin. HDAC1 and HDAC2 are recruited to UVR-induced DNA damage spots, which are visualized by anti-XPC immunofluorescence. Dual HDAC1/2 depletion decreases XPC ubiquitination, but does not affect the recruitment of DDB2 to DNA damage. By contrast, the local accumulation of γH2AX at UVR-induced DNA damage spots was compromised upon HDAC1 as well as dual HDAC1/2 depletions. Additionally, UVR-induced ATM activation decreased in H12899 cells expressing H3K56Ac-mimicing H3K56Q. These results revealed a novel role of DDB in H3K56Ac deacetylation during early step of NER and the existence of active functional cross-talk between DDB-mediated damage recognition and H3K56Ac deacetylation.

  18. Methylation of histone H3 lysine-79 by Dot1p plays multiple roles in the response to UV damage in Saccharomyces cerevisiae.

    Science.gov (United States)

    Bostelman, Lindsey J; Keller, Andrew M; Albrecht, Ashley M; Arat, Arzu; Thompson, Jeffrey S

    2007-03-01

    Various proteins have been found to play roles in both the repair of UV damaged DNA and heterochromatin-mediated silencing in the yeast Saccharomyces cerevisiae. In particular, factors that are involved in the methylation of lysine-79 of histone H3 by Dot1p have been implicated in both processes, suggesting a bipartite function for this modification. We find that a dot1 null mutation and a histone H3 point mutation at lysine-79 cause increased sensitivity to UV radiation, suggesting that lysine-79 methylation is important for efficient repair of UV damage. Epistasis analysis between dot1 and various UV repair genes indicates that lysine-79 methylation plays overlapping roles within the nucleotide excision, post-replication and recombination repair pathways, as well as RAD9-mediated checkpoint function. In contrast, epistasis analysis with the H3 lysine-79 point mutation indicates that the lysine-to-glutamic acid substitution exerts specific effects within the nucleotide excision repair and post-replication repair pathways, suggesting that this allele only disrupts a subset of the functions of lysine-79 methylation. The overall results indicate the existence of distinct and separable roles of histone H3 lysine-79 methylation in the response to UV damage, potentially serving to coordinate the various repair processes.

  19. The histone variant H3.3 G34W substitution in giant cell tumor of the bone link chromatin and RNA processing.

    Science.gov (United States)

    Lim, Jinyeong; Park, Joo Hyun; Baude, Annika; Yoo, Yeongran; Lee, Yeon Kyu; Schmidt, Christopher R; Park, Jong Bae; Fellenberg, Jörg; Zustin, Josef; Haller, Florian; Krücken, Irene; Kang, Hyun Guy; Park, Yoon Jung; Plass, Christoph; Lindroth, Anders M

    2017-10-18

    While transcription as regulated by histones and their post-translational modifications has been well described, the function of histone variants in this process remains poorly characterized. Potentially important insight into this process pertain to the frequently occurring mutations of H3.3, leading to G34 substitutions in childhood glioblastoma and giant cell tumor of the bone (GCTB). In this study, we have established primary cell lines from GCTB patients and used them to uncover the influence of H3.3 G34W substitutions on cellular growth behavior, gene expression, and chromatin compaction. Primary cell lines with H3.3 G34W showed increased colony formation, infiltration and proliferation, known hallmarks of tumor development. Isogenic cell lines with H3.3 G34W recapitulated the increased proliferation observed in primary cells. Transcriptomic analysis of primary cells and tumor biopsies revealed slightly more downregulated gene expression, perhaps by increased chromatin compaction. We identified components related to splicing, most prominently hnRNPs, by immunoprecipitation and mass spectrometry that specifically interact with H3.3 G34W in the isogenic cell lines. RNA-sequencing analysis and hybridization-based validations further enforced splicing aberrations. Our data uncover a role for H3.3 in RNA processing and chromatin modulation that is blocked by the G34W substitution, potentially driving the tumorigenic process in GCTB.

  20. Clipping of Flexible Tails of Histones H3 and H4 Affects the Structure and Dynamics of the Nucleosome

    Science.gov (United States)

    Nurse, Nathan P.; Jimenez-Useche, Isabel; Smith, Ian Tad; Yuan, Chongli

    2013-01-01

    Förster resonance energy transfer was used to monitor the dynamic conformations of mononucleosomes under different chromatin folding conditions to elucidate the role of the flexible N-terminal regions of H3 and H4 histones. The H3 tail was shown to partake in intranucleosomal interactions by restricting the DNA breathing motion and compacting the nucleosome. The H3 tail effects were mostly independent of the ionic strength and valency of the ions. The H4 tail was shown to not greatly affect the nucleosome conformation, but did slightly influence the relative population of the preferred conformation. The role of the H4 tail varied depending on the valency and ionic strength, suggesting that electrostatic forces play a primary role in H4 tail interactions. Interestingly, despite the H4 tail’s lack of influence, when H3 and H4 tails were simultaneously clipped, a more dramatic effect was seen than when only H3 or H4 tails were clipped. The combinatorial effect of H3 and H4 tail truncation suggests a potential mechanism by which various combinations of histone tail modifications can be used to control accessibility of DNA-binding proteins to nucleosomal DNA. PMID:23473491

  1. Posttranslational Modifications of the Histone 3 Tail and Their Impact on the Activity of Histone Lysine Demethylases In Vitro

    DEFF Research Database (Denmark)

    Lohse, Brian; Helgstrand, Charlotte; Andersson, Jan Legaard

    2013-01-01

    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...... 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 understanding of the dynamics of epigenetic marks....

  2. Cross-communication between histone H3 and H4 acetylation and Akt-mTOR signalling in prostate cancer cells.

    Science.gov (United States)

    Makarević, Jasmina; Tawanaie, Nassim; Juengel, Eva; Reiter, Michael; Mani, Jens; Tsaur, Igor; Bartsch, Georg; Haferkamp, Axel; Blaheta, Roman A

    2014-07-01

    Molecular tumour targeting has significantly improved anti-cancer protocols. Still, the addition of molecular targeting to the treatment regime has not led to a curative breakthrough. Combined mammalian target of Rapamycin (mTOR) and histone deacetylase (HDAC) inhibition has been shown not only to enhance anti-tumour potential, but also to prevent resistance development seen under mono-drug therapy. This investigation was designed to evaluate whether cross-communication exists between mTOR signalling and epigenetic events regulated by HDAC. DU-145 prostate cancer cells were treated with insulin-like growth factor (IGF) to activate the Akt-mTOR cascade or with the HDAC-inhibitor valproic acid (VPA) to induce histone H3 and H4 acetylation (aH3, aH4). Subsequently, mTOR, Rictor, Raptor, p70s6k, Akt (all: total and phosphorylated), H3 and H4 (total and acetylated) were analysed by western blotting. Both techniques revealed a link between mTOR and the epigenetic machinery. IGF activated mTOR, Rictor, Raptor, p70s6k and Akt, but also enhanced aH3 and aH4. Inversely, IGFr blockade and knock-down blocked the Akt-mTOR axis, but simultaneously diminished aH3 and aH4. VPA treatment up-regulated histone acetylation, but also activated mTOR-Akt signalling. HDAC1 and 2 knock-down revealed that the interaction with the mTOR system is initiated by histone H3 acetylation. HDAC-mTOR communication, therefore, is apparent whereby tumour-promoting (Akt/mTOR(high), aH3/aH4(low)) and tumour-suppressing signals (Akt/mTOR(low) , aH3/aH4(high)) are activated in parallel. Combined use of an HDAC- and mTOR inhibitor might then diminish pro-tumour effects triggered by the HDAC- (Akt/mTOR(high)) or mTOR inhibitor (aH3/aH4(low)) alone. © 2014 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  3. The amino-terminal tails of histones H2A and H3 coordinate efficient base excision repair, DNA damage signaling and postreplication repair in Saccharomyces cerevisiae.

    Science.gov (United States)

    Meas, Rithy; Smerdon, Michael J; Wyrick, John J

    2015-05-26

    Histone amino-terminal tails (N-tails) are required for cellular resistance to DNA damaging agents; therefore, we examined the role of histone N-tails in regulating DNA damage response pathways in Saccharomyces cerevisiae. Combinatorial deletions reveal that the H2A and H3 N-tails are important for the removal of MMS-induced DNA lesions due to their role in regulating the basal and MMS-induced expression of DNA glycosylase Mag1. Furthermore, overexpression of Mag1 in a mutant lacking the H2A and H3 N-tails rescues base excision repair (BER) activity but not MMS sensitivity. We further show that the H3 N-tail functions in the Rad9/Rad53 DNA damage signaling pathway, but this function does not appear to be the primary cause of MMS sensitivity of the double tailless mutants. Instead, epistasis analyses demonstrate that the tailless H2A/H3 phenotypes are in the RAD18 epistasis group, which regulates postreplication repair. We observed increased levels of ubiquitylated PCNA and significantly lower mutation frequency in the tailless H2A/H3 mutant, indicating a defect in postreplication repair. In summary, our data identify novel roles of the histone H2A and H3 N-tails in (i) regulating the expression of a critical BER enzyme (Mag1), (ii) supporting efficient DNA damage signaling and (iii) facilitating postreplication repair. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  4. Genome-wide profiling of histone h3 lysine 4 and lysine 27 trimethylation reveals an epigenetic signature in prostate carcinogenesis.

    Directory of Open Access Journals (Sweden)

    Xi-Song Ke

    Full Text Available BACKGROUND: Increasing evidence implicates the critical roles of epigenetic regulation in cancer. Very recent reports indicate that global gene silencing in cancer is associated with specific epigenetic modifications. However, the relationship between epigenetic switches and more dynamic patterns of gene activation and repression has remained largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: Genome-wide profiling of the trimethylation of histone H3 lysine 4 (H3K4me3 and lysine 27 (H3K27me3 was performed using chromatin immunoprecipitation coupled with whole genome promoter microarray (ChIP-chip techniques. Comparison of the ChIP-chip data and microarray gene expression data revealed that loss and/or gain of H3K4me3 and/or H3K27me3 were strongly associated with differential gene expression, including microRNA expression, between prostate cancer and primary cells. The most common switches were gain or loss of H3K27me3 coupled with low effect on gene expression. The least prevalent switches were between H3K4me3 and H3K27me3 coupled with much higher fractions of activated and silenced genes. Promoter patterns of H3K4me3 and H3K27me3 corresponded strongly with coordinated expression changes of regulatory gene modules, such as HOX and microRNA genes, and structural gene modules, such as desmosome and gap junction genes. A number of epigenetically switched oncogenes and tumor suppressor genes were found overexpressed and underexpressed accordingly in prostate cancer cells. CONCLUSIONS/SIGNIFICANCE: This work offers a dynamic picture of epigenetic switches in carcinogenesis and contributes to an overall understanding of coordinated regulation of gene expression in cancer. Our data indicate an H3K4me3/H3K27me3 epigenetic signature of prostate carcinogenesis.

  5. Chromatin H3K27me3/H3K4me3 histone marks define gene sets in high-grade serous ovarian cancer that distinguish malignant, tumour-sustaining and chemo-resistant ovarian tumour cells.

    Science.gov (United States)

    Chapman-Rothe, N; Curry, E; Zeller, C; Liber, D; Stronach, E; Gabra, H; Ghaem-Maghami, S; Brown, R

    2013-09-19

    In embryonic stem (ES) cells, bivalent chromatin domains containing H3K4me3 and H3K27me3 marks silence developmental genes, while keeping them poised for activation following differentiation. We have identified gene sets associated with H3K27me3 and H3K4me3 marks at transcription start sites in a high-grade ovarian serous tumour and examined their association with epigenetic silencing and malignant progression. This revealed novel silenced bivalent marked genes, not described previously for ES cells, which are significantly enriched for the PI3K (P<10(-7)) and TGF-β signalling pathways (P<10(-5)). We matched histone marked gene sets to gene expression sets of eight normal fallopian tubes and 499 high-grade serous malignant ovarian samples. This revealed a significant decrease in gene expression for the H3K27me3 and bivalent gene sets in malignant tissue. We then correlated H3K27me3 and bivalent gene sets to gene expression data of ovarian tumour 'stem cell-like' sustaining cells versus non-sustaining cells. This showed a significantly lower expression for the H3K27me3 and bivalent gene sets in the tumour-sustaining cells. Similarly, comparison of matched chemo-sensitive and chemo-resistant ovarian cell lines showed a significantly lower expression of H3K27me3/bivalent marked genes in the chemo-resistant compared with the chemo-sensitive cell line. Our analysis supports the hypothesis that bivalent marks are associated with epigenetic silencing in ovarian cancer. However it also suggests that additional tumour specific bivalent marks, to those known in ES cells, are present in tumours and may potentially influence the subsequent development of drug resistance and tumour progression.

  6. The Histone Demethylase Jarid1b Ensures Faithful Mouse Development by Protecting Developmental Genes from Aberrant H3K4me3

    DEFF Research Database (Denmark)

    Albert, Mareike; Schmitz, Sandra U; Kooistra, Susanne M

    2013-01-01

    Embryonic development is tightly regulated by transcription factors and chromatin-associated proteins. H3K4me3 is associated with active transcription and H3K27me3 with gene repression, while the combination of both keeps genes required for development in a plastic state. Here we show that deleti...... Pax6 and Otx2 in Jarid1b knockout brains. Taken together, these results suggest that Jarid1b regulates mouse development by protecting developmental genes from inappropriate acquisition of active histone modifications....

  7. Phosphorylation of histone H3 on Ser-10 by Aurora B is essential for chromosome condensation in porcine embryos during the first mitotic division.

    Science.gov (United States)

    Chen, Changchao; Zhang, Zixiao; Cui, Panpan; Liao, Yaya; Zhang, Yue; Yao, Lingyun; Rui, Rong; Ju, Shiqiang

    2017-07-01

    Phosphorylation of histone H3 on Ser-10 (H3S10ph) is involved in regulating mitotic chromosome condensation and decondensation, which plays an important regulatory role during mitotic cell cycle progression in mammalian cells. However, whether H3S10ph plays a similar role in early porcine embryos during the first mitotic division remains uncertain. In this study, the subcellular localization and possible roles of H3S10ph were evaluated in the first mitotic cell cycle progression of porcine embryos using western blot, indirect immunofluorescence and barasertib (H3S10ph upstream regulator Aurora-B inhibitor) treatments. H3S10ph exhibited a dynamic localization pattern and was localized to chromosomes from prometaphase to anaphase stages. Treatment of porcine embryos with barasertib inhibited mitotic division at the prophase stage and was associated with a defect in chromosome condensation accompanied by the reduction of H3S10ph. These results indicated that H3S10ph is involved in the first mitotic division in porcine embryos through its regulatory function in chromosome condensation, which further affects porcine embryo cell cycle progression during mitotic division.

  8. Chromatin methylation activity of Dnmt3a and Dnmt3a/3L is guided by interaction of the ADD domain with the histone H3 tail

    Science.gov (United States)

    Zhang, Yingying; Jurkowska, Renata; Soeroes, Szabolcs; Rajavelu, Arumugam; Dhayalan, Arunkumar; Bock, Ina; Rathert, Philipp; Brandt, Ole; Reinhardt, Richard; Fischle, Wolfgang; Jeltsch, Albert

    2010-01-01

    Using peptide arrays and binding to native histone proteins, we show that the ADD domain of Dnmt3a specifically interacts with the H3 histone 1–19 tail. Binding is disrupted by di- and trimethylation of K4, phosphorylation of T3, S10 or T11 and acetylation of K4. We did not observe binding to the H4 1–19 tail. The ADD domain of Dnmt3b shows the same binding specificity, suggesting that the distinct biological functions of both enzymes are not related to their ADD domains. To establish a functional role of the ADD domain binding to unmodified H3 tails, we analyzed the DNA methylation of in vitro reconstituted chromatin with Dnmt3a2, the Dnmt3a2/Dnmt3L complex, and the catalytic domain of Dnmt3a. All Dnmt3a complexes preferentially methylated linker DNA regions. Chromatin substrates with unmodified H3 tail or with H3K9me3 modification were methylated more efficiently by full-length Dnmt3a and full-length Dnmt3a/3L complexes than chromatin trimethylated at H3K4. In contrast, the catalytic domain of Dnmt3a was not affected by the H3K4me3 modification. These results demonstrate that the binding of the ADD domain to H3 tails unmethylated at K4 leads to the preferential methylation of DNA bound to chromatin with this modification state. Our in vitro results recapitulate DNA methylation patterns observed in genome-wide DNA methylation studies. PMID:20223770

  9. Heterochromatin and histone modifications in the germline-restricted chromosome of the zebra finch undergoing elimination during spermatogenesis.

    Science.gov (United States)

    Goday, Clara; Pigozzi, María Inés

    2010-06-01

    In the zebra finch (Taeniopygia guttata) a germline-restricted chromosome (GRC) is regularly present in males and females. While the GRC is euchromatic in oocytes, in spermatocytes this chromosome is cytologically seen as entirely heterochromatic and presumably inactive. At the end of male meiosis, the GRC is eliminated from the nucleus. By immunofluorescence on microspreads, we investigated HP1 proteins and histone modifications throughout male meiotic prophase, as well as in young spermatid stages after the GRC elimination. We found that in prophase spermatocytes the GRC chromatin differs from that of the regular chromosome complement. The GRC is highly enriched in HP1 beta and exhibits high levels of di- and tri-methylated histone H3 at lysine 9 and tri- and di-methylated histone H4 at lysine 20. The GRC does not exhibit neither detectable levels of di- and tri-methylated histone H3 at lysine 4 nor acetylated histone H4 at lysine 5 and 8. The results prove the heterochromatic organization of the GRC in male germline and strongly suggest its transcriptional inactive state during male prophase. Following elimination, in young spermatids the GRC lacks HP1 beta signals but maintains high levels of methylated histone H3 at lysine 9 and methylated histone H4 at lysine 20. The release of HP1 from the GRC with respect to its elimination is discussed.

  10. The PHD1 finger of KDM5B recognizes unmodified H3K4 during the demethylation of histone H3K4me2/3 by KDM5B.

    Science.gov (United States)

    Zhang, Yan; Yang, Huirong; Guo, Xue; Rong, Naiyan; Song, Yujiao; Xu, Youwei; Lan, Wenxian; Zhang, Xu; Liu, Maili; Xu, Yanhui; Cao, Chunyang

    2014-11-01

    KDM5B is a histone H3K4me2/3 demethylase. The PHD1 domain of KDM5B is critical for demethylation, but the mechanism underlying the action of this domain is unclear. In this paper, we observed that PHD1KDM5B interacts with unmethylated H3K4me0. Our NMR structure of PHD1KDM5B in complex with H3K4me0 revealed that the binding mode is slightly different from that of other reported PHD fingers. The disruption of this interaction by double mutations on the residues in the interface (L325A/D328A) decreases the H3K4me2/3 demethylation activity of KDM5B in cells by approximately 50% and increases the transcriptional repression of tumor suppressor genes by approximately twofold. These findings imply that PHD1KDM5B may help maintain KDM5B at target genes to mediate the demethylation activities of KDM5B.

  11. p38-MAPK/MSK1-mediated overexpression of histone H3 serine 10 phosphorylation defines distance-dependent prognostic value of negative resection margin in gastric cancer.

    Science.gov (United States)

    Khan, Shafqat Ali; Amnekar, Ramchandra; Khade, Bharat; Barreto, Savio George; Ramadwar, Mukta; Shrikhande, Shailesh V; Gupta, Sanjay

    2016-01-01

    Alterations in histone modifications are now well known to result in epigenetic heterogeneity in tumor tissues; however, their prognostic value and association with resection margins still remain poorly understood and controversial. Further, histopathologically negative resection margins in several cancers have been associated with better prognosis of the disease. However, in gastric cancer, despite a high rate of R0 resection, a considerably high incidence of loco-regional recurrence is observed. We believe alterations of global histone post-translational modifications could help in identifying molecular signatures for defining the true negative surgical resection margins and also the prognosis of gastric cancer patients. The present study compares the level of H3S10ph among paired tumor and histopathologically confirmed disease-free (R0) proximal and distal surgical resection margin (PRM and DRM) tissue samples of GC patients (n = 101). Immunoblotting and immune-histochemical analysis showed a significantly (p cancer cells confirmed p38-MAPK/MSK1 pathway-mediated regulation of H3S10ph in gastric cancer. Our study provides the first evidence that p38-MAPK/MSK1-regulated increase of H3S10ph in GC is predictive of a more aggressive cancer phenotype and could help in defining true negative surgical resection margin. Importantly, our data also gave a new rationale for exploration of the use of MSK1 inhibitor in gastric cancer therapy and the combination of histone post-translational modifications, H4K16ac and H4K20me3 along with H3S10ph as epigenetic prognostic markers.

  12. KAT6B Is a Tumor Suppressor Histone H3 Lysine 23 Acetyltransferase Undergoing Genomic Loss in Small Cell Lung Cancer.

    Science.gov (United States)

    Simó-Riudalbas, Laia; Pérez-Salvia, Montserrat; Setien, Fernando; Villanueva, Alberto; Moutinho, Catia; Martínez-Cardús, Anna; Moran, Sebastian; Berdasco, Maria; Gomez, Antonio; Vidal, Enrique; Soler, Marta; Heyn, Holger; Vaquero, Alejandro; de la Torre, Carolina; Barceló-Batllori, Silvia; Vidal, August; Roz, Luca; Pastorino, Ugo; Szakszon, Katalin; Borck, Guntram; Moura, Conceição S; Carneiro, Fátima; Zondervan, Ilse; Savola, Suvi; Iwakawa, Reika; Kohno, Takashi; Yokota, Jun; Esteller, Manel

    2015-09-15

    Recent efforts to sequence human cancer genomes have highlighted that point mutations in genes involved in the epigenetic setting occur in tumor cells. Small cell lung cancer (SCLC) is an aggressive tumor with poor prognosis, where little is known about the genetic events related to its development. Herein, we have identified the presence of homozygous deletions of the candidate histone acetyltransferase KAT6B, and the loss of the corresponding transcript, in SCLC cell lines and primary tumors. Furthermore, we show, in vitro and in vivo, that the depletion of KAT6B expression enhances cancer growth, while its restoration induces tumor suppressor-like features. Most importantly, we demonstrate that KAT6B exerts its tumor-inhibitory role through a newly defined type of histone H3 Lys23 acetyltransferase activity. ©2015 American Association for Cancer Research.

  13. Inhibition of histone acetylation by curcumin reduces alcohol-induced expression of heart development-related transcription factors in cardiac progenitor cells.

    Science.gov (United States)

    Wang, Linyi; Sun, Huichao; Pan, Bo; Zhu, Jing; Huang, Guoying; Huang, Xupei; Tian, Jie

    2012-08-03

    Alcohol exposure during pregnancy may cause congenital heart disease (CHD). In our previous studies, we found that alcohol selectively increased acetylation of histone H3 at lysine 9 (H3K9) and enhanced the expression of heart development-related genes in cardiac progenitor cells. The objective of this study is to investigate the protective effects of histone acetyltransferases (HATs) inhibitor, curcumin, on histone hyperacetylation and the over-expression of heart development genes induced by alcohol. Western blot analysis was employed to detect the acetylation levels of histone H3K9 and real-time PCR was applied to measure the expressions of heart development-related transcription factors, GATA4, Mef2c and Tbx5 (GMT). Our results showed that alcohol increased the acetylation of H3K9 by 2.76-fold (Palcohol plus 25 μM curcumin, the hyperacetylation of H3K9 and over-expression of GATA4 and Mef2c by alcohol was reversed. These data indicate that curcumin can correct the over-expression of cardiac genes by reversing the alcohol induced hyperacetylation of histone H3 at lysine 9 in cardiac progenitor cells, suggesting that curcumin is protective against alcohol-induced cardiac gene over-expression that may result in heart malformations. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. Effect of SPL (Spent Pot Liner) and its main components on root growth, mitotic activity and phosphorylation of Histone H3 in Lactuca sativa L.

    Science.gov (United States)

    Freitas, Aline Silva; Fontes Cunha, Isabela Martinez; Andrade-Vieira, Larissa Fonseca; Techio, Vânia Helena

    2016-02-01

    Spent Pot Liner (SPL) is a solid waste from the aluminum industry frequently disposed of in industrial landfills; it can be leached and contaminate the soil, sources of drinking water and plantations, and thus may pose a risk to human health and to ecosystems. Its composition is high variable, including cyanide, fluoride and aluminum salts, which are highly toxic and environmental pollutants. This study evaluated the effect of SPL and its main components on root growth and the mitosis of Lactuca sativa, by investigating the mechanisms of cellular and chromosomal alterations with the aid of immunolocalization. To this end, newly emerged roots of L. sativa were exposed to SPL and its main components (solutions of cyanide, fluoride and aluminum) and to calcium chloride (control) for 48h. After this, root length was measured and cell cycle was examined by means of conventional cytogenetics and immunolocalization. Root growth was inhibited in the treatments with SPL and aluminum; chromosomal and nuclear alterations were observed in all treatments. The immunolocalization evidenced normal dividing cells with regular temporal and spatial distribution of histone H3 phosphorylation at serine 10 (H3S10ph). However, SPL and its main components inhibited the phosphorylation of histone H3 at serine 10, inactivated pericentromeric regions and affected the cohesion of sister chromatids, thus affecting the arrangement of chromosomes in the metaphase plate and separation of chromatids in anaphase. In addition, these substances induced breaks in pericentromeric regions, characterized as fragile sites. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. The replication-independent histone H3-H4 chaperones HIR, ASF1, and RTT106 co-operate to maintain promoter fidelity.

    Science.gov (United States)

    Silva, Andrea C; Xu, Xiaomeng; Kim, Hyun-Soo; Fillingham, Jeffrey; Kislinger, Thomas; Mennella, Thomas A; Keogh, Michael-Christopher

    2012-01-13

    RNA polymerase II initiates from low complexity sequences so cells must reliably distinguish "real" from "cryptic" promoters and maintain fidelity to the former. Further, this must be performed under a range of conditions, including those found within inactive and highly transcribed regions. Here, we used genome-scale screening to identify those factors that regulate the use of a specific cryptic promoter and how this is influenced by the degree of transcription over the element. We show that promoter fidelity is most reliant on histone gene transactivators (Spt10, Spt21) and H3-H4 chaperones (Asf1, HIR complex) from the replication-independent deposition pathway. Mutations of Rtt106 that abrogate its interactions with H3-H4 or dsDNA permit extensive cryptic transcription comparable with replication-independent deposition factor deletions. We propose that nucleosome shielding is the primary means to maintain promoter fidelity, and histone replacement is most efficiently mediated in yeast cells by a HIR/Asf1/H3-H4/Rtt106 pathway.

  16. Characterization of an antagonistic switch between histone H3 lysine 27 methylation and acetylation in the transcriptional regulation of Polycomb group target genes.

    Science.gov (United States)

    Pasini, Diego; Malatesta, Martina; Jung, Hye Ryung; Walfridsson, Julian; Willer, Anton; Olsson, Linda; Skotte, Julie; Wutz, Anton; Porse, Bo; Jensen, Ole Nørregaard; Helin, Kristian

    2010-08-01

    Polycomb group (PcG) proteins are transcriptional repressors, which regulate proliferation and cell fate decisions during development, and their deregulated expression is a frequent event in human tumours. The Polycomb repressive complex 2 (PRC2) catalyzes trimethylation (me3) of histone H3 lysine 27 (K27), and it is believed that this activity mediates transcriptional repression. Despite the recent progress in understanding PcG function, the molecular mechanisms by which the PcG proteins repress transcription, as well as the mechanisms that lead to the activation of PcG target genes are poorly understood. To gain insight into these mechanisms, we have determined the global changes in histone modifications in embryonic stem (ES) cells lacking the PcG protein Suz12 that is essential for PRC2 activity. We show that loss of PRC2 activity results in a global increase in H3K27 acetylation. The methylation to acetylation switch correlates with the transcriptional activation of PcG target genes, both during ES cell differentiation and in MLL-AF9-transduced hematopoietic stem cells. Moreover, we provide evidence that the acetylation of H3K27 is catalyzed by the acetyltransferases p300 and CBP. Based on these data, we propose that the PcG proteins in part repress transcription by preventing the binding of acetyltransferases to PcG target genes.

  17. Mammalian target of rapamycin complex 2 (mTORC2) controls glycolytic gene expression by regulating Histone H3 Lysine 56 acetylation.

    Science.gov (United States)

    Vadla, Raghavendra; Haldar, Devyani

    2017-11-16

    Metabolic reprogramming is a hallmark of cancer cells, but the mechanisms are not well understood. The mammalian target of rapamycin complex 2 (mTORC2) controls cell growth and proliferation and plays a critical role in metabolic reprogramming in glioma. mTORC2 regulates cellular processes such as cell survival, metabolism, and proliferation by phosphorylation of AGC kinases. Components of mTORC2 are shown to localize to the nucleus, but whether mTORC2 modulates epigenetic modifications to regulate gene expression is not known. Here, we identified histone H3 lysine 56 acetylation (H3K56Ac) is regulated by mTORC2 and show that global H3K56Ac levels were downregulated on mTORC2 knockdown but not on mTORC1 knockdown. mTORC2 promotes H3K56Ac in a tuberous sclerosis complex 1/2 (TSC1/2) mediated signaling pathway. We show that knockdown of sirtuin6 (SIRT6) prevented H3K56 deacetylation in mTORC2 depleted cells. Using glioma model consisting of U87EGFRvIII cells, we established that mTORC2 promotes H3K56Ac in glioma. Finally, we show that mTORC2 regulates the expression of glycolytic genes by regulating H3K56Ac levels at the promoters of these genes in glioma cells and depletion of mTOR leads to increased recruitment of SIRT6 to these promoters. Collectively, these results identify mTORC2 signaling pathway positively promotes H3K56Ac through which it may mediate metabolic reprogramming in glioma.

  18. Supplementation of fetal bovine serum alters histone modification H3R26me2 during preimplantation development of in vitro produced bovine embryos

    Directory of Open Access Journals (Sweden)

    Daniel R. Arnold

    2015-07-01

    Full Text Available Abstract In vitro production (IVP of bovine embryos is not only of great economic importance to the cattle industry, but is also an important model for studying embryo development. The aim of this study was to evaluate the histone modification, H3R26me2 during pre-implantation development of IVP bovine embryos cultured with or without serum supplementation and how these in vitro treatments compared to in vivo embryos at the morula stage. After in vitro maturation and fertilization, bovine embryos were cultured with either 0 or 2.5% fetal bovine serum (FBS. Development was evaluated and embryos were collected and fixed at different stages during development (2-, 4-, 8-, 16-cell, morula and blastocyst. Fixed embryos were then used for immunofluorescence utilizing an antibody for H3R26me2. Images of stained embryos were analyzed as a percentage of total DNA. Embryos cultured with 2.5% FBS developed to blastocysts at a greater rate than 0%FBS groups (34.85±5.43% vs. 23.38±2.93%; P<0.05. Levels of H3R26me2 changed for both groups over development. In the 0%FBS group, the greatest amount of H3R26me2 staining was at the 4-cell (P<0.05, 16-cell (P<0.05 and morula (P<0.05 stages. In the 2.5%FBS group, only 4-cell stage embryos were significantly higher than all other stages (P<0.01. Morula stage in vivo embryos had similar levels as the 0%FBS group, and both were significantly higher than the 2.5%FBS group. These results suggest that the histone modification H3R26me2 is regulated during development of pre-implantation bovine embryos, and that culture conditions greatly alter this regulation.

  19. Hypermethylated in cancer 1 (HIC1), a tumor suppressor gene epigenetically deregulated in hyperparathyroid tumors by histone H3 lysine modification.

    Science.gov (United States)

    Svedlund, Jessica; Koskinen Edblom, Susanne; Marquez, Victor E; Åkerström, Göran; Björklund, Peyman; Westin, Gunnar

    2012-07-01

    Primary hyperparathyroidism (pHPT) resulting from parathyroid tumors is a common endocrine disorder with incompletely understood etiology. In renal failure, secondary hyperparathyroidism (sHPT) occurs with multiple tumor development as a result of calcium and vitamin D regulatory disturbance. The aim of the study was to investigate whether HIC1 may act as a tumor suppressor in the parathyroid glands and whether deregulated expression involves epigenetic mechanisms. Parathyroid tumors from patients with pHPT included single adenomas, multiple tumors from the same patient, and cancer. Hyperplastic parathyroid glands from patients with sHPT and hypercalcemia and normal parathyroid tissue specimens were included in the study. Quantitative RT-PCR, bisulfite pyrosequencing, colony formation assay, chromatin immunoprecipitation, and RNA interference was used. HIC1 was generally underexpressed regardless of the hyperparathyroid disease state including multiple parathyroid tumors from the same patient, and overexpression of HIC1 led to a decrease in clonogenic survival of parathyroid tumor cells. Only the carcinomas showed a high methylation level and reduced HIC1 expression. Cell culture experiments, including use of primary parathyroid tumor cells prepared directly after operation, the general histone methyltransferase inhibitor 3-deazaneplanocin A, chromatin immunoprecipitation, and RNA interference of DNA methyltransferases and EZH2 (enhancer of zeste homolog 2), supported a role of repressive histone H3 modifications (H3K27me2/3) rather than DNA methylation in repression of HIC1. The results strongly support a growth-regulatory role of HIC1 in the parathyroid glands and suggest that perturbed expression of HIC1 may represent an early event during tumor development. Repressive histone modification H3K27me2/3 is involved in repression of HIC1 expression in hyperparathyroid tumors.

  20. The histone H3K9 methylation and RNAi pathways regulate normalnucleolar and repeated DNA organization by inhibiting formation ofextrachromosomal DNAs

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Jamy C.; Karpen, Gary H.

    2006-06-15

    In order to identify regulators of nuclear organization, Drosophila mutants in the Su(var)3-9 histone H3K9 methyltransferase, RNAi pathway components, and other regulators of heterochromatin-mediated gene silencing were examined for altered nucleoli and positioning of repeated DNAs. Animals lacking components of the H3K9 methylation and RNAi pathways contained disorganized nucleoli, ribosomal DNA (rDNA) and satellite DNAs. The levels of H3K9 dimethylation (H3K9me2) in chromatin associated with repeated DNAs decreased dramatically in Su(var)3-9 and dcr-2 (dicer-2) mutant tissues compared to wild type. We also observed a substantial increase in extrachromosomal repeated DNAs in mutant tissues. The disorganized nucleolus phenotype depends on the presence of Ligase 4 (Lig4), and ecc DNA formation is not induced by removal of cohesin. We conclude that H3K9 methylation of rDNA and satellites, maintained by Su(var)3-9, HP1, and the RNAi pathway, is necessary for the structural stability of repeated DNAs, which is mediated through suppression of non-homologous end joining (NHEJ). These results suggest a mechanism for how local chromatin structure can regulate genome stability, and the organization of chromosomal elements and nuclear organelles.

  1. Characterization of an antagonistic switch between histone H3 lysine 27 methylation and acetylation in the transcriptional regulation of Polycomb group target genes

    DEFF Research Database (Denmark)

    Pasini, Diego; Malatesta, Martina; Jung, Hye Ryung

    2010-01-01

    are poorly understood. To gain insight into these mechanisms, we have determined the global changes in histone modifications in embryonic stem (ES) cells lacking the PcG protein Suz12 that is essential for PRC2 activity. We show that loss of PRC2 activity results in a global increase in H3K27 acetylation...... on these data, we propose that the PcG proteins in part repress transcription by preventing the binding of acetyltransferases to PcG target genes....

  2. Transgenerational programming of longevity through E(z)-mediated histone H3K27 trimethylation in Drosophila.

    Science.gov (United States)

    Xia, Brian; Gerstin, Ed; Schones, Dustin E; Huang, Wendong; Steven de Belle, J

    2016-11-25

    Transgenerational effects on health and development of early-life nutrition have gained increased attention recently. However, the underlying mechanisms of transgenerational transmission are only starting to emerge, with epigenetics as perhaps the most important mechanism. We recently reported the first animal model to study transgenerational programming of longevity after early-life dietary manipulations, enabling investigations to identify underlying epigenetic mechanisms. We report here that post-eclosion dietary manipulation (PDM) with a low-protein (LP) diet upregulates the protein level of E(z), an H3K27 specific methyltransferase, leading to higher levels of H3K27 trimethylation (H3K27me3). This PDM-mediated change in H3K27me3 corresponded with a shortened longevity of F0 flies as well as their F2 offspring. Specific RNAi-mediated post-eclosion knockdown of E(z) or pharmacological inhibition of its enzymatic function with EPZ-6438 in the F0 parents improved longevity while rendering H3K27me3 low across generations. Importantly, addition of EPZ-6438 to the LP diet fully alleviated the longevity-reducing effect of the LP PDM, supporting the increased level of E(z)-dependent H3K27me3 as the primary cause and immediate early-life period as the critical time to program longevity through epigenetic regulation. These observations establish E(z)-mediated H3K27me3 as one epigenetic mechanism underlying nutritional programming of longevity and support the use of EPZ-6438 to extend lifespan.

  3. Differential acetylation of histone H3 at the regulatory region of OsDREB1b promoter facilitates chromatin remodelling and transcription activation during cold stress.

    Directory of Open Access Journals (Sweden)

    Dipan Roy

    Full Text Available The rice ortholog of DREB1, OsDREB1b, is transcriptionally induced by cold stress and over-expression of OsDREB1b results in increase tolerance towards high salt and freezing stress. This spatio-temporal expression of OsDREB1b is preceded by the change in chromatin structure at the promoter and the upstream region for gene activation. The promoter and the upstream region of OsDREB1b genes appear to be arranged into a nucleosome array. Nucleosome mapping of ∼ 700 bp upstream region of OsDREB1b shows two positioned nucleosomes between -610 to -258 and a weakly positioned nucleosome at the core promoter and the TSS. Upon cold stress, there is a significant change in the nucleosome arrangement at the upstream region with increase in DNaseI hypersensitivity or MNase digestion in the vicinity of cis elements and TATA box at the core promoter. ChIP assays shows hyper-acetylation of histone H3K9 throughout the locus whereas region specific increase was observed in H3K14ac and H3K27ac. Moreover, there is an enrichment of RNA PolII occupancy at the promoter region during transcription activation. There is no significant change in the H3 occupancy in OsDREB1b locus negating the possibility of nucleosome loss during cold stress. Interestingly, cold induced enhanced transcript level of OsDREB1b as well as histone H3 acetylation at the upstream region was found to diminish when stressed plants were returned to normal temperature. The result indicates absolute necessity of changes in chromatin conformation for the transcription up-regulation of OsDREB1b gene in response to cold stress. The combined results show the existence of closed chromatin conformation at the upstream and promoter region of OsDREB1b in the transcription "off" state. During cold stress, changes in region specific histone modification marks promote the alteration of chromatin structure to facilitate the binding of transcription machinery for proper gene expression.

  4. ChIP-seq analysis of histone H3K9 trimethylation in peripheral blood mononuclear cells of membranous nephropathy patients

    Energy Technology Data Exchange (ETDEWEB)

    Sui, W.G. [Guangxi Key Laboratory of Metabolic Diseases Research, Nephrology Department, 181st Hospital, Guilin, Guangxi (China); He, H.Y. [The Life Science College, Guangxi Normal University, Guilin, Guangxi (China); Yan, Q.; Chen, J.J. [Guangxi Key Laboratory of Metabolic Diseases Research, Nephrology Department, 181st Hospital, Guilin, Guangxi (China); Zhang, R.H. [The Life Science College, Guangxi Normal University, Guilin, Guangxi (China); Dai, Y. [Clinical Medical Research Center, The Second Clinical Medical College, Shenzhen People’s Hospital, Jinan University, Shenzhen, Guangdong (China)

    2013-12-12

    Membranous nephropathy (MN), characterized by the presence of diffuse thickening of the glomerular basement membrane and subepithelial in situ immune complex disposition, is the most common cause of idiopathic nephrotic syndrome in adults, with an incidence of 5-10 per million per year. A number of studies have confirmed the relevance of several experimental insights to the pathogenesis of human MN, but the specific biomarkers of MN have not been fully elucidated. As a result, our knowledge of the alterations in histone methylation in MN is unclear. We used chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) to analyze the variations in a methylated histone (H3K9me3) in peripheral blood mononuclear cells from 10 MN patients and 10 healthy subjects. There were 108 genes with significantly different expression in the MN patients compared with the normal controls. In MN patients, significantly increased activity was seen in 75 H3K9me3 genes, and decreased activity was seen in 33, compared with healthy subjects. Five positive genes, DiGeorge syndrome critical region gene 6 (DGCR6), sorting nexin 16 (SNX16), contactin 4 (CNTN4), baculoviral IAP repeat containing 3 (BIRC3), and baculoviral IAP repeat containing 2 (BIRC2), were selected and quantified. There were alterations of H3K9me3 in MN patients. These may be candidates to help explain pathogenesis in MN patients. Such novel findings show that H3K9me3 may be a potential biomarker or promising target for epigenetic-based MN therapies.

  5. Intrabody-mediated diverting of HP1β to the cytoplasm induces co-aggregation of H3-H4 histones and lamin-B receptor.

    Science.gov (United States)

    Cardinale, Alessio; Filesi, Ilaria; Singh, Prim B; Biocca, Silvia

    2015-10-15

    Diverting a protein from its intracellular location is a unique property of intrabodies. To interfere with the intracellular traffic of heterochromatin protein 1β (HP1β) in living cells, we have generated a cytoplasmic targeted anti-HP1β intrabody, specifically directed against the C-terminal portion of the molecule. HP1β is a conserved component of mouse and human constitutive heterochromatin involved in diverse nuclear functions including gene silencing, DNA repair and nuclear membrane assembly. We found that the anti-HP1β intrabody sequesters HP1β into cytoplasmic aggregates, inhibiting its traffic to the nucleus. Lamin B receptor (LBR) and a subset of core histones (H3/H4) are also specifically co-sequestered in the cytoplasm of anti-HP1β intrabody-expressing cells. Methylated histone H3 at K9 (Me9H3), a marker of constitutive heterochromatin, is not affected by the anti-HP1β intrabody expression. Hyper-acetylating conditions completely dislodge H3 from HP1β:LBR containing aggregates. The expression of anti-HP1β scFv fragments induces apoptosis, associated with an alteration of nuclear morphology. Both these phenotypes are specifically rescued either by overexpression of recombinant full length HP1β or by HP1β mutant containing the chromoshadow domain, but not by recombinant LBR protein. The HP1β-chromodomain mutant, on the other hand, does not rescue the phenotypes, but does compete with LBR for binding to HP1β. These findings provide new insights into the mode of action of cytoplasmic-targeted intrabodies and the interaction between HP1β and its binding partners involved in peripheral heterochromatin organisation. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Precision mapping of coexisting modifications in histone H3 tails from embryonic stem cells by ETD-MS/MS

    DEFF Research Database (Denmark)

    Jung, Hye Ryung; Sidoli, Simone; Haldbo, Simon

    2013-01-01

    H3 N-terminal tail peptides (amino acids 1-50, 5-6 kDa) were separated by optimized weak cation exchange/hydrophilic interaction liquid chromatography (WCX/HILIC) and sequenced online by electron transfer dissociation (ETD) tandem mass spectrometry (MS/MS). High mass accuracy and near complete...

  7. Chromosomal Mapping of Repetitive DNAs in the Grasshopper Abracris flavolineata Reveal Possible Ancestry of the B Chromosome and H3 Histone Spreading.

    Directory of Open Access Journals (Sweden)

    Danilo Bueno

    Full Text Available Supernumerary chromosomes (B chromosomes occur in approximately 15% of eukaryote species. Although these chromosomes have been extensively studied, knowledge concerning their specific molecular composition is lacking in most cases. The accumulation of repetitive DNAs is one remarkable characteristic of B chromosomes, and the occurrence of distinct types of multigene families, satellite DNAs and some transposable elements have been reported. Here, we describe the organization of repetitive DNAs in the A complement and B chromosome system in the grasshopper species Abracris flavolineata using classical cytogenetic techniques and FISH analysis using probes for five multigene families, telomeric repeats and repetitive C0t-1 DNA fractions. The 18S rRNA and H3 histone multigene families are highly variable and well distributed in A. flavolineata chromosomes, which contrasts with the conservation of U snRNA genes and less variable distribution of 5S rDNA sequences. The H3 histone gene was an extensively distributed with clusters occurring in all chromosomes. Repetitive DNAs were concentrated in C-positive regions, including the pericentromeric region and small chromosomal arms, with some occurrence in C-negative regions, but abundance was low in the B chromosome. Finally, the first demonstration of the U2 snRNA gene in B chromosomes in A. flavolineata may shed light on its possible origin. These results provide new information regarding chromosomal variability for repetitive DNAs in grasshoppers and the specific molecular composition of B chromosomes.

  8. Chromosomal Mapping of Repetitive DNAs in the Grasshopper Abracris flavolineata Reveal Possible Ancestry of the B Chromosome and H3 Histone Spreading.

    Science.gov (United States)

    Bueno, Danilo; Palacios-Gimenez, Octavio Manuel; Cabral-de-Mello, Diogo Cavalcanti

    2013-01-01

    Supernumerary chromosomes (B chromosomes) occur in approximately 15% of eukaryote species. Although these chromosomes have been extensively studied, knowledge concerning their specific molecular composition is lacking in most cases. The accumulation of repetitive DNAs is one remarkable characteristic of B chromosomes, and the occurrence of distinct types of multigene families, satellite DNAs and some transposable elements have been reported. Here, we describe the organization of repetitive DNAs in the A complement and B chromosome system in the grasshopper species Abracris flavolineata using classical cytogenetic techniques and FISH analysis using probes for five multigene families, telomeric repeats and repetitive C0t-1 DNA fractions. The 18S rRNA and H3 histone multigene families are highly variable and well distributed in A. flavolineata chromosomes, which contrasts with the conservation of U snRNA genes and less variable distribution of 5S rDNA sequences. The H3 histone gene was an extensively distributed with clusters occurring in all chromosomes. Repetitive DNAs were concentrated in C-positive regions, including the pericentromeric region and small chromosomal arms, with some occurrence in C-negative regions, but abundance was low in the B chromosome. Finally, the first demonstration of the U2 snRNA gene in B chromosomes in A. flavolineata may shed light on its possible origin. These results provide new information regarding chromosomal variability for repetitive DNAs in grasshoppers and the specific molecular composition of B chromosomes.

  9. Dynamic link between histone H3 acetylation and an increase in the functional characteristics of human ESC/iPSC-derived cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Tomomi G Otsuji

    Full Text Available Cardiomyocytes (CMs derived from human embryonic stem cells (hESCs or human induced pluripotent stem cells (hiPSCs are functionally heterogeneous, display insufficient biological efficacy and generally possess the electrophysiological properties seen in fetal CMs. However, a homogenous population of hESC/hiPSC-CMs, with properties similar to those of adult human ventricular cells, is required for use in drug cardiotoxicity screening. Unfortunately, despite the requirement for the functional characteristics of post-mitotic beating cell aggregates to mimic the behavior of mature cardiomyocytes in vitro, few technological improvements have been made in this field to date. Previously, we showed that culturing hESC-CMs under low-adhesion conditions with cyclic replating confers continuous contractility on the cells, leading to a functional increase in cardiac gene expression and electrophysiological properties over time. The current study reveals that culturing hESC/hiPSC-CMs under non-adhesive culture conditions enhances the electrophysiological properties of the CMs through an increase in the acetylation of histone H3 lysine residues, as confirmed by western blot analyses. Histone H3 acetylation was induced chemically by treating primitive hESC/hiPSC-CMs with Trichostatin A (TSA, a histone deacetylase (HDAC inhibitor, resulting in an immediate increase in global cardiac gene expression. In functional analyses using multi-electrode array (MEA recordings, TSA-treated hESC/hiPSC-CM colonies showed appropriate responses to particular concentrations of known potassium ion channel inhibitors. Thus, the combination of a cell-autonomous functional increase in response to non-adhesive culture and short-term TSA treatment of hESC/hiPSC-CM colonies cultured on MEA electrodes will help to make cardiac toxicity tests more accurate and reproducible via genome-wide chromatin activation.

  10. Dynamics of RNA Polymerase II Pausing and Bivalent Histone H3 Methylation during Neuronal Differentiation in Brain Development

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    Jiancheng Liu

    2017-08-01

    Full Text Available During cellular differentiation, genes important for differentiation are expected to be silent in stem/progenitor cells yet can be readily activated. RNA polymerase II (Pol II pausing and bivalent chromatin marks are two paradigms suited for establishing such a poised state of gene expression; however, their specific contributions in development are not well understood. Here we characterized Pol II pausing and H3K4me3/H3K27me3 marks in neural progenitor cells (NPCs and their daughter neurons purified from the developing mouse cortex. We show that genes paused in NPCs or neurons are characteristic of respective cellular functions important for each cell type, although pausing and pause release were not correlated with gene activation. Bivalent chromatin marks poised the marked genes in NPCs for activation in neurons. Interestingly, we observed a positive correlation between H3K27me3 and paused Pol II. This study thus reveals cell type-specific Pol II pausing and gene activation-associated bivalency during mammalian neuronal differentiation.

  11. The Set1/COMPASS histone H3 methyltransferase helps regulate mitosis with the CDK1 and NIMA mitotic kinases in Aspergillus nidulans.

    Science.gov (United States)

    Govindaraghavan, Meera; Anglin, Sarah Lea; Osmani, Aysha H; Osmani, Stephen A

    2014-08-01

    Mitosis is promoted and regulated by reversible protein phosphorylation catalyzed by the essential NIMA and CDK1 kinases in the model filamentous fungus Aspergillus nidulans. Protein methylation mediated by the Set1/COMPASS methyltransferase complex has also been shown to regulate mitosis in budding yeast with the Aurora mitotic kinase. We uncover a genetic interaction between An-swd1, which encodes a subunit of the Set1 protein methyltransferase complex, with NIMA as partial inactivation of nimA is poorly tolerated in the absence of swd1. This genetic interaction is additionally seen without the Set1 methyltransferase catalytic subunit. Importantly partial inactivation of NIMT, a mitotic activator of the CDK1 kinase, also causes lethality in the absence of Set1 function, revealing a functional relationship between the Set1 complex and two pivotal mitotic kinases. The main target for Set1-mediated methylation is histone H3K4. Mutational analysis of histone H3 revealed that modifying the H3K4 target residue of Set1 methyltransferase activity phenocopied the lethality seen when either NIMA or CDK1 are partially functional. We probed the mechanistic basis of these genetic interactions and find that the Set1 complex performs functions with CDK1 for initiating mitosis and with NIMA during progression through mitosis. The studies uncover a joint requirement for the Set1 methyltransferase complex with the CDK1 and NIMA kinases for successful mitosis. The findings extend the roles of the Set1 complex to include the initiation of mitosis with CDK1 and mitotic progression with NIMA in addition to its previously identified interactions with Aurora and type 1 phosphatase in budding yeast. Copyright © 2014 by the Genetics Society of America.

  12. Chromosomal mapping of H3 histone and 5S rRNA genes in eight species of Astyanax (Pisces, Characiformes) with different diploid numbers: syntenic conservation of repetitive genes.

    Science.gov (United States)

    Piscor, Diovani; Parise-Maltempi, Patricia Pasquali

    2016-03-01

    The genus Astyanax is widely distributed from the southern United States to northern Patagonia, Argentina. While cytogenetic studies have been performed for this genus, little is known about the histone gene families. The aim of this study was to examine the chromosomal relationships among the different species of Astyanax. The chromosomal locations of the 5S rRNA and H3 histone genes were determined in A. abramis, A. asuncionensis, A. altiparanae, A. bockmanni, A. eigenmanniorum, A. mexicanus (all 2n = 50), A. fasciatus (2n = 46), and A. schubarti (2n = 36). All eight species exhibited H3 histone clusters on two chromosome pairs. In six species (A. abramis, A. asuncionensis, A. altiparanae, A. bockmanni, A. eigenmanniorum, and A. fasciatus), syntenic clusters of H3 histone and 5S rDNA were observed on metacentric (m) or submetacentric (sm) chromosomes. In seven species, clusters of 5S rDNA sequences were located on one or two chromosome pairs. In A. mexicanus, 5S rDNA clusters were located on four chromosome pairs. This study demonstrates that H3 histone clusters are conserved on two chromosome pairs in the genus Astyanax, and specific chromosomal features may contribute to the genomic organization of the H3 histone and 5S rRNA genes.

  13. Novel and shared neoantigen derived from histone 3 variant H3.3K27M mutation for glioma T cell therapy.

    Science.gov (United States)

    Chheda, Zinal S; Kohanbash, Gary; Okada, Kaori; Jahan, Naznin; Sidney, John; Pecoraro, Matteo; Yang, Xinbo; Carrera, Diego A; Downey, Kira M; Shrivastav, Shruti; Liu, Shuming; Lin, Yi; Lagisetti, Chetana; Chuntova, Pavlina; Watchmaker, Payal B; Mueller, Sabine; Pollack, Ian F; Rajalingam, Raja; Carcaboso, Angel M; Mann, Matthias; Sette, Alessandro; Garcia, K Christopher; Hou, Yafei; Okada, Hideho

    2017-12-04

    The median overall survival for children with diffuse intrinsic pontine glioma (DIPG) is less than one year. The majority of diffuse midline gliomas, including more than 70% of DIPGs, harbor an amino acid substitution from lysine (K) to methionine (M) at position 27 of histone 3 variant 3 (H3.3). From a CD8+ T cell clone established by stimulation of HLA-A2+ CD8+ T cells with synthetic peptide encompassing the H3.3K27M mutation, complementary DNA for T cell receptor (TCR) α- and β-chains were cloned into a retroviral vector. TCR-transduced HLA-A2+ T cells efficiently killed HLA-A2+H3.3K27M+ glioma cells in an antigen- and HLA-specific manner. Adoptive transfer of TCR-transduced T cells significantly suppressed the progression of glioma xenografts in mice. Alanine-scanning assays suggested the absence of known human proteins sharing the key amino acid residues required for recognition by the TCR, suggesting that the TCR could be safely used in patients. These data provide us with a strong basis for developing T cell-based therapy targeting this shared neoepitope. © 2018 Chheda et al.

  14. Alcohol-induced histone H3K9 hyperacetylation and cardiac hypertrophy are reversed by a histone acetylases inhibitor anacardic acid in developing murine hearts.

    Science.gov (United States)

    Peng, Chang; Zhang, Weihua; Zhao, Weian; Zhu, Jing; Huang, Xupei; Tian, Jie

    2015-06-01

    The expression of cardiac genes is precisely regulated, and any perturbation may cause developmental defects. In a previous study, we demonstrated that alcohol consumption during pregnancy could lead to uncontrolled expressions of cardiac genes and eventually result in cardiac dysplasia. However, the underlying mechanisms remain unclear. In the present study, we have investigated the alcohol-induced cardiac hypertrophy and its potential mechanisms. Furthermore, the protective effect of anacardic acid against the alcohol-induced cardiac hypertrophy has been explored in experimental mice. C57BL/6 pregnant mice were gavaged with 56% ethanol or saline and the hearts of their fetus were collected for analysis. Binding of p300, CBP, PCAF, SRC1, except GCN5, were increased to the NKX2.5 promoter in fetal mouse hearts exposed to alcohol. Increased acetylation of H3K9 and increased mRNA expression of NKX2.5, β-MHC and Cx43 were observed in the same samples. Treatment with a pan-acetylase inhibitor, anacardic acid, reduced the binding affinity of p300 and PCAF to the NKX2.5, β-MHC, Cx43 promoters and attenuated H3K9 hyperacetylation. Interestingly, anacardic acid down-regulated over-expression of these cardiac genes induced by alcohol and ultimately attenuated ethanol-induced cardiac hypertrophy in fetal mice. Our results indicate that alcohol exposure during pregnancy could lead to fetal cardiac hypertrophy. The over-expression of NKX2.5, β-MHC, Cx43 mediated by p300 and PCAF may be critical mechanisms of alcohol-induced cardiac hypertrophy. Anacardic acid can down-regulate the over-expression of cardiac genes and reverse cardiac hypertrophy caused by alcohol treatment in pregnant mice, suggesting it could be a potential therapeutic agent for the treatment of cardiac hypertrophy. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  15. Mitotic instability in wheat x Thinopyrum ponticum derivatives revealed by chromosome counting, nuclear DNA content and histone H3 phosphorylation pattern.

    Science.gov (United States)

    Brasileiro-Vidal, A C; Brammer, S; Puertas, M J; Zanatta, A C; Prestes, A; Moraes-Fernandes, M I B; Guerra, M

    2005-06-01

    To evaluate the mitotic stability of Triticum aestivum x Thinopyrum ponticum derivatives (BC(2)F(7) and BC(2)F(5) doubled haploids), chromosome counting by both conventional and immunostaining techniques, and measurement of DNA content were performed. The wheat progenitor line, PF 839197, the wheat recurrent parent CEP 19 and the control Chinese Spring were also investigated. In the hybrid derivatives, chromosome number ranged from 2n=36 to 60, with a predominance of chromosome numbers higher than 2n=42, that was confirmed by determination of nuclear DNA content. Chinese Spring' and PF 839197 were stable, but CEP 19 showed chromosome number variation (20%). Analyses of non-pretreated cells revealed the presence of anaphase bridges, lagging chromatids, chromosome fragments and micronuclei. Immunostaining with an antibody recognizing histone H3 phosphorylated showed dicentric chromatids forming anaphase bridges and pericentromeric phosphorylation at centric chromosome fragments but not at lagging chromatids. The possible causes of the observed mitotic instability are discussed.

  16. Nuclear localization of CPI-17, a protein phosphatase-1 inhibitor protein, affects histone H3 phosphorylation and corresponds to proliferation of cancer and smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Eto, Masumi, E-mail: masumi.eto@jefferson.edu [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kirkbride, Jason A.; Chugh, Rishika; Karikari, Nana Kofi [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kim, Jee In [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Cardiovascular Research Institute, Kyungpook National University School of Medicine, Daegu 700-422 (Korea, Republic of)

    2013-04-26

    Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation.

  17. Genome-wide quantitative analysis of histone H3 lysine 4 trimethylation in wild house mouse liver: environmental change causes epigenetic plasticity.

    Directory of Open Access Journals (Sweden)

    Angelika G Börsch-Haubold

    Full Text Available In mammals, exposure to toxic or disease-causing environments can change epigenetic marks that are inherited independently of the intrauterine environment. Such inheritance of molecular phenotypes may be adaptive. However, studies demonstrating molecular evidence for epigenetic inheritance have so far relied on extreme treatments, and are confined to inbred animals. We therefore investigated whether epigenomic changes could be detected after a non-drastic change in the environment of an outbred organism. We kept two populations of wild-caught house mice (Mus musculus domesticus for several generations in semi-natural enclosures on either standard diet and light cycle, or on an energy-enriched diet with longer daylight to simulate summer. As epigenetic marker for active chromatin we quantified genome-wide histone-3 lysine-4 trimethylation (H3K4me3 from liver samples by chromatin immunoprecipitation and high-throughput sequencing as well as by quantitative polymerase chain reaction. The treatment caused a significant increase of H3K4me3 at metabolic genes such as lipid and cholesterol regulators, monooxygenases, and a bile acid transporter. In addition, genes involved in immune processes, cell cycle, and transcription and translation processes were also differently marked. When we transferred young mice of both populations to cages and bred them under standard conditions, most of the H3K4me3 differences were lost. The few loci with stable H3K4me3 changes did not cluster in metabolic functional categories. This is, to our knowledge, the first quantitative study of an epigenetic marker in an outbred mammalian organism. We demonstrate genome-wide epigenetic plasticity in response to a realistic environmental stimulus. In contrast to disease models, the bulk of the epigenomic changes we observed were not heritable.

  18. Genome-wide quantitative analysis of histone H3 lysine 4 trimethylation in wild house mouse liver: environmental change causes epigenetic plasticity.

    Science.gov (United States)

    Börsch-Haubold, Angelika G; Montero, Inka; Konrad, Kathryn; Haubold, Bernhard

    2014-01-01

    In mammals, exposure to toxic or disease-causing environments can change epigenetic marks that are inherited independently of the intrauterine environment. Such inheritance of molecular phenotypes may be adaptive. However, studies demonstrating molecular evidence for epigenetic inheritance have so far relied on extreme treatments, and are confined to inbred animals. We therefore investigated whether epigenomic changes could be detected after a non-drastic change in the environment of an outbred organism. We kept two populations of wild-caught house mice (Mus musculus domesticus) for several generations in semi-natural enclosures on either standard diet and light cycle, or on an energy-enriched diet with longer daylight to simulate summer. As epigenetic marker for active chromatin we quantified genome-wide histone-3 lysine-4 trimethylation (H3K4me3) from liver samples by chromatin immunoprecipitation and high-throughput sequencing as well as by quantitative polymerase chain reaction. The treatment caused a significant increase of H3K4me3 at metabolic genes such as lipid and cholesterol regulators, monooxygenases, and a bile acid transporter. In addition, genes involved in immune processes, cell cycle, and transcription and translation processes were also differently marked. When we transferred young mice of both populations to cages and bred them under standard conditions, most of the H3K4me3 differences were lost. The few loci with stable H3K4me3 changes did not cluster in metabolic functional categories. This is, to our knowledge, the first quantitative study of an epigenetic marker in an outbred mammalian organism. We demonstrate genome-wide epigenetic plasticity in response to a realistic environmental stimulus. In contrast to disease models, the bulk of the epigenomic changes we observed were not heritable.

  19. Genome-wide profiling of histone modifications (H3K9me2 and H4K12ac) and gene expression in rust (uromyces appendiculatus) inoculated common bean (Phaseolus vulgaris L)

    OpenAIRE

    Ayyappan, Vasudevan; Kalavacharla, Venu (Kal); Thimmapuram, Jyothi; Bhide, Ketaki P; Sripathi, Venkateswara R.

    2015-01-01

    Histone modifications such as methylation and acetylation play a significant role in controlling gene expression in unstressed and stressed plants. Genome-wide analysis of such stress-responsive modifications and genes in non-model crops is limited. We report the genome-wide profiling of histone methylation (H3K9me2) and acetylation (H4K12ac) in common bean (Phaseolus vulgaris L.) under rust (Uromyces appendiculatus) stress using two high-throughput approaches, chromatin immunoprecipitation s...

  20. Structure/Function Analysis of Recurrent Mutations in SETD2 Protein Reveals a Critical and Conserved Role for a SET Domain Residue in Maintaining Protein Stability and Histone H3 Lys-36 Trimethylation.

    Science.gov (United States)

    Hacker, Kathryn E; Fahey, Catherine C; Shinsky, Stephen A; Chiang, Yun-Chen J; DiFiore, Julia V; Jha, Deepak Kumar; Vo, Andy H; Shavit, Jordan A; Davis, Ian J; Strahl, Brian D; Rathmell, W Kimryn

    2016-09-30

    The yeast Set2 histone methyltransferase is a critical enzyme that plays a number of key roles in gene transcription and DNA repair. Recently, the human homologue, SETD2, was found to be recurrently mutated in a significant percentage of renal cell carcinomas, raising the possibility that the activity of SETD2 is tumor-suppressive. Using budding yeast and human cell line model systems, we examined the functional significance of two evolutionarily conserved residues in SETD2 that are recurrently mutated in human cancers. Whereas one of these mutations (R2510H), located in the Set2 Rpb1 interaction domain, did not result in an observable defect in SETD2 enzymatic function, a second mutation in the catalytic domain of this enzyme (R1625C) resulted in a complete loss of histone H3 Lys-36 trimethylation (H3K36me3). This mutant showed unchanged thermal stability as compared with the wild type protein but diminished binding to the histone H3 tail. Surprisingly, mutation of the conserved residue in Set2 (R195C) similarly resulted in a complete loss of H3K36me3 but did not affect dimethylated histone H3 Lys-36 (H3K36me2) or functions associated with H3K36me2 in yeast. Collectively, these data imply a critical role for Arg-1625 in maintaining the protein interaction with H3 and specific H3K36me3 function of this enzyme, which is conserved from yeast to humans. They also may provide a refined biochemical explanation for how H3K36me3 loss leads to genomic instability and cancer. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Repeated forced swim stress affects the expression of pCREB and ΔFosB and the acetylation of histone H3 in the rostral ventromedial medulla and locus coeruleus.

    Science.gov (United States)

    Imbe, Hiroki; Kimura, Akihisa

    2016-10-01

    The rostral ventromedial medulla (RVM) and locus coeruleus (LC) play crucial roles in descending pain modulation system. In the present study we examined the expression of phospho-cAMP response element-binding protein (pCREB) and ΔFosB and the acetylation of histone H3 in the RVM and LC after forced swim stress (FS) and complete Freund's adjuvant (CFA) injection to clarify changes in descending pain modulatory system in a rat model of stress-induced hyperalgesia. FS (day 1, 10min; days 2-3, 20min) induced a significant increase in the expression of pCREB and ΔFosB and the acetylation of histone H3 in the RVM, whereas the FS induced a significant increase only in the acetylation of histone H3 in the LC. CFA injection into the hindpaw did not induce a significant change in those expression and acetylation. Quantitative image analysis demonstrated that the numbers of pCREB-, acetylated histone H3- and ΔFosB-IR cells in the RVM were significantly higher in the FS group than those in the naive group. The CFA injection after the FS did not affect the FS-induced increases in the expression of pCREB and ΔFosB and the acetylation of histone H3 in the RVM even though nullified the increase in the acetylation of histone H3 in the LC. These findings suggest different neuroplasticities between the RVM and LC after the FS, which may be involved in activity change of descending pain modulatory system after the CFA injection. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Loss of the DNA methyltransferase MET1 Induces H3K9 hypermethylation at PcG target genes and redistribution of H3K27 trimethylation to transposons in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Angelique Deleris

    Full Text Available Dimethylation of histone H3 lysine 9 (H3K9m2 and trimethylation of histone H3 lysine 27 (H3K27m3 are two hallmarks of transcriptional repression in many organisms. In Arabidopsis thaliana, H3K27m3 is targeted by Polycomb Group (PcG proteins and is associated with silent protein-coding genes, while H3K9m2 is correlated with DNA methylation and is associated with transposons and repetitive sequences. Recently, ectopic genic DNA methylation in the CHG context (where H is any base except G has been observed in globally DNA hypomethylated mutants such as met1, but neither the nature of the hypermethylated loci nor the biological significance of this epigenetic phenomenon have been investigated. Here, we generated high-resolution, genome-wide maps of both H3K9m2 and H3K27m3 in wild-type and met1 plants, which we integrated with transcriptional data, to explore the relationships between these two marks. We found that ectopic H3K9m2 observed in met1 can be due to defects in IBM1-mediated H3K9m2 demethylation at some sites, but most importantly targets H3K27m3-marked genes, suggesting an interplay between these two silencing marks. Furthermore, H3K9m2/DNA-hypermethylation at these PcG targets in met1 is coupled with a decrease in H3K27m3 marks, whereas CG/H3K9m2 hypomethylated transposons become ectopically H3K27m3 hypermethylated. Our results bear interesting similarities with cancer cells, which show global losses of DNA methylation but ectopic hypermethylation of genes previously marked by H3K27m3.

  3. Preferential recruitment of the maternal centromere-specific histone H3 (CENH3) in oat (Avena sativa L.) × pearl millet (Pennisetum glaucum L.) hybrid embryos.

    Science.gov (United States)

    Ishii, Takayoshi; Sunamura, Naohiro; Matsumoto, Ayaka; Eltayeb, Amin Elsadig; Tsujimoto, Hisashi

    2015-12-01

    Chromosome elimination occurs frequently in interspecific hybrids between distantly related species in Poaceae. However, chromosomes from both parents behave stably in a hybrid of female oat (Avena sativa L.) pollinated by pearl millet (Pennisetum glaucum L.). To analyze the chromosome behavior in this hybrid, we cloned the centromere-specific histone H3 (CENH3) genes of oat and pearl millet and produced a pearl millet-specific anti-CENH3 antibody. Application of this antibody together with a grass species common anti-CENH3 antibody revealed the dynamic CENH3 composition of the hybrid cells before and after fertilization. Despite co-expression of CENH3 genes encoded by oat and pearl millet, only an oat-type CENH3 was incorporated into the centromeres of both species in the hybrid embryo. Oat CENH3 enables a functional centromere in pearl millet chromosomes in an oat genetic background. Comparison of CENH3 genes among Poaceae species that show chromosome elimination in interspecific hybrids revealed that the loop 1 regions of oat and pearl millet CENH3 exhibit exceptionally high similarity.

  4. Smad4 mediated BMP2 signal is essential for the regulation of GATA4 and Nkx2.5 by affecting the histone H3 acetylation in H9c2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Si, Lina; Shi, Jin; Gao, Wenqun [Heart Centre, Children’s Hospital of Chongqing Medical University, 136 Zhongshan 2nd Road, Yu Zhong District, Chongqing 400014 (China); Ministry of Education Key Laboratory of Child Development and Disorders, Key Laboratory of Pediatrics in Chongqing, Chongqing International Science and Technology Cooperation Center for Child Development and Disorders, 136 Zhongshan 2nd Road, Yu Zhong District, Chongqing 400014 (China); Zheng, Min [Heart Centre, Children’s Hospital of Chongqing Medical University, 136 Zhongshan 2nd Road, Yu Zhong District, Chongqing 400014 (China); Liu, Lingjuan; Zhu, Jing [Ministry of Education Key Laboratory of Child Development and Disorders, Key Laboratory of Pediatrics in Chongqing, Chongqing International Science and Technology Cooperation Center for Child Development and Disorders, 136 Zhongshan 2nd Road, Yu Zhong District, Chongqing 400014 (China); Tian, Jie, E-mail: jietian@cqmu.edu.cn [Heart Centre, Children’s Hospital of Chongqing Medical University, 136 Zhongshan 2nd Road, Yu Zhong District, Chongqing 400014 (China)

    2014-07-18

    Highlights: • BMP2 can upregulated cardiac related gene GATA4, Nkx2.5, MEF2c and Tbx5. • Inhibition of Smad4 decreased BMP2-induced hyperacetylation of histone H3. • Inhibition of Smad4 diminished BMP2-induced overexpression of GATA4 and Nkx2.5. • Inhibition of Smad4 decreased hyperacetylated H3 in the promoter of GATA4 and Nkx2.5. • Smad4 is essential for BMP2 induced hyperacetylated histone H3. - Abstract: BMP2 signaling pathway plays critical roles during heart development, Smad4 encodes the only common Smad protein in mammals, which is a pivotal nuclear mediator. Our previous studies showed that BMP2 enhanced the expression of cardiac transcription factors in part by increasing histone H3 acetylation. In the present study, we tested the hypothesis that Smad4 mediated BMP2 signaling pathway is essential for the expression of cardiac core transcription factors by affecting the histone H3 acetylation. We successfully constructed a lentivirus-mediated short hairpin RNA interference vector targeting Smad4 (Lv-Smad4) in rat H9c2 embryonic cardiac myocytes (H9c2 cells) and demonstrated that it suppressed the expression of the Smad4 gene. Cultured H9c2 cells were transfected with recombinant adenoviruses expressing human BMP2 (AdBMP2) with or without Lv-Smad4. Quantitative real-time RT-PCR analysis showed that knocking down of Smad4 substantially inhibited both AdBMP2-induced and basal expression levels of cardiac transcription factors GATA4 and Nkx2.5, but not MEF2c and Tbx5. Similarly, chromatin immunoprecipitation (ChIP) analysis showed that knocking down of Smad4 inhibited both AdBMP2-induced and basal histone H3 acetylation levels in the promoter regions of GATA4 and Nkx2.5, but not of Tbx5 and MEF2c. In addition, Lv-Smad4 selectively suppressed AdBMP2-induced expression of HAT p300, but not of HAT GCN5 in H9c2 cells. The data indicated that inhibition of Smad4 diminished both AdBMP2 induced and basal histone acetylation levels in the promoter regions of

  5. HP1 recruits activity-dependent neuroprotective protein to H3K9me3 marked pericentromeric heterochromatin for silencing of major satellite repeats.

    Directory of Open Access Journals (Sweden)

    Kerstin Mosch

    Full Text Available H3 lysine 9 trimethylation (H3K9me3 is a histone posttranslational modification (PTM that has emerged as hallmark of pericentromeric heterochromatin. This constitutive chromatin domain is composed of repetitive DNA elements, whose transcription is differentially regulated. Mammalian cells contain three HP1 proteins, HP1α, HP1β and HP1γ These have been shown to bind to H3K9me3 and are thought to mediate the effects of this histone PTM. However, the mechanisms of HP1 chromatin regulation and the exact functional role at pericentromeric heterochromatin are still unclear. Here, we identify activity-dependent neuroprotective protein (ADNP as an H3K9me3 associated factor. We show that ADNP does not bind H3K9me3 directly, but that interaction is mediated by all three HP1 isoforms in vitro. However, in cells ADNP localization to areas of pericentromeric heterochromatin is only dependent on HP1α and HP1β. Besides a PGVLL sequence patch we uncovered an ARKS motif within the ADNP homeodomain involved in HP1 dependent H3K9me3 association and localization to pericentromeric heterochromatin. While knockdown of ADNP had no effect on HP1 distribution and heterochromatic histone and DNA modifications, we found ADNP silencing major satellite repeats. Our results identify a novel factor in the translation of H3K9me3 at pericentromeric heterochromatin that regulates transcription.

  6. A fluorescence resonance energy transfer-based method for histone methyltransferases

    DEFF Research Database (Denmark)

    Devkota, Kanchan; Lohse, Brian; Nyby Jakobsen, Camilla

    2015-01-01

    A simple dye–quencher fluorescence resonance energy transfer (FRET)-based assay for methyltransferases was developed and used to determine kinetic parameters and inhibitory activity at EHMT1 and EHMT2. Peptides mimicking the truncated histone H3 tail were functionalized in each end with a dye...... and a quencher, respectively. When lysine-9 residues in the peptides were methylated, they were protected from cleavage by endoproteinase–EndoLysC, whereas unmethylated peptides were cleaved, resulting in an increase in fluorescent intensity....

  7. Elucidation of the Two H3K36me3 Histone Methyltransferases Set2 and Ash1 in Fusarium fujikuroi Unravels Their Different Chromosomal Targets and a Major Impact of Ash1 on Genome Stability.

    Science.gov (United States)

    Janevska, Slavica; Baumann, Leonie; Sieber, Christian M K; Münsterkötter, Martin; Ulrich, Jonas; Kämper, Jörg; Güldener, Ulrich; Tudzynski, Bettina

    2017-11-16

    In this work, we present a comprehensive analysis of the H3K36 histone methyltransferases Set2 and Ash1 in the filamentous ascomycete Fusarium fujikuroi In Saccharomyces cerevisiae, one single methyltransferase, Set2, confers all H3K36 methylation, while there are two members of the Set2 family in filamentous fungi, and even more H3K36 methyltransferases in higher eukaryotes. Whereas the yeast Set2 homolog has been analyzed in fungi previously, the second member of the Set2 family, designated Ash1, has not been described for any filamentous fungus. Western blot and ChIP-Seq analyses confirmed that F. fujikuroi Set2 and Ash1 are H3K36-specific histone methyltransferases that deposit H3K36me3 at specific loci: Set2 is most likely responsible for H3K36 methylation of euchromatic regions of the genome, while Ash1 methylates H3K36 at the subtelomeric regions (facultative heterochromatin) of all chromosomes including the accessory chromosome XII. Our data indicate that H3K36me3 cannot be considered a hallmark of euchromatin in F. fujikuroi, and likely also other filamentous fungi, making them different to what is known about nuclear characteristics in yeast and higher eukaryotes. We suggest that the H3K36 methylation mark exerts specific functions when deposited at euchromatic or subtelomeric regions by Set2 or Ash1, respectively. We found an enhanced level of H3K27me3, an increased instability of subtelomeric regions and losses of the accessory chromosome XII over time in Δash1 mutants, indicating an involvement of Ash1 in DNA repair processes. Further phenotypic analyses revealed a role of H3K36 methylation in vegetative growth, sporulation, secondary metabolite biosynthesis and virulence in F. fujikuroi. Copyright © 2017, Genetics.

  8. Synaptic activity induces dramatic changes in the geometry of the cell nucleus: interplay between nuclear structure, histone H3 phosphorylation, and nuclear calcium signaling.

    Science.gov (United States)

    Wittmann, Malte; Queisser, Gillian; Eder, Anja; Wiegert, J Simon; Bengtson, C Peter; Hellwig, Andrea; Wittum, Gabriel; Bading, Hilmar

    2009-11-25

    Synaptic activity initiates many adaptive responses in neurons. Here we report a novel form of structural plasticity in dissociated hippocampal cultures and slice preparations. Using a recently developed algorithm for three-dimensional image reconstruction and quantitative measurements of cell organelles, we found that many nuclei from hippocampal neurons are highly infolded and form unequally sized nuclear compartments. Nuclear infoldings are dynamic structures, which can radically transform the geometry of the nucleus in response to neuronal activity. Action potential bursting causing synaptic NMDA receptor activation dramatically increases the number of infolded nuclei via a process that requires the ERK-MAP kinase pathway and new protein synthesis. In contrast, death-signaling pathways triggered by extrasynaptic NMDA receptors cause a rapid loss of nuclear infoldings. Compared with near-spherical nuclei, infolded nuclei have a larger surface and increased nuclear pore complex immunoreactivity. Nuclear calcium signals evoked by cytosolic calcium transients are larger in small nuclear compartments than in the large compartments of the same nucleus; moreover, small compartments are more efficient in temporally resolving calcium signals induced by trains of action potentials in the theta frequency range (5 Hz). Synaptic activity-induced phosphorylation of histone H3 on serine 10 was more robust in neurons with infolded nuclei compared with neurons with near-spherical nuclei, suggesting a functional link between nuclear geometry and transcriptional regulation. The translation of synaptic activity-induced signaling events into changes in nuclear geometry facilitates the relay of calcium signals to the nucleus, may lead to the formation of nuclear signaling microdomains, and could enhance signal-regulated transcription.

  9. Evolutionary dynamics of 5S rDNA location in acridid grasshoppers and its relationship with H3 histone gene and 45S rDNA location.

    Science.gov (United States)

    Cabral-de-Mello, Diogo C; Cabrero, Josefa; López-León, María Dolores; Camacho, Juan Pedro M

    2011-07-01

    We analyze the chromosomal location of 5S rDNA clusters in 29 species of grasshoppers belonging to the family Acrididae. There was extensive variation among species for the number and location of 5S rDNA sites. Out of 148 sites detected, 75% were proximally located, 21.6% were interstitial, and only 3.4% were distal. The number of 5S rDNA sites per species varied from a single chromosome pair (in six species) to all chromosome pairs (in five species), with a range of intermediate situations. Thirteen chromosomes from eight species carried two 5S rDNA clusters. At intraspecific level, differences among populations were detected in Eyprepocnemis plorans, and some heteromorphisms have also been observed in some species. Double FISH for 5S rDNA and H3 histone gene DNA, performed on 17 of these 29 species, revealed that both markers are sometimes placed in a same chromosome but at different location, whereas they appeared to co-localize in five species (Calliptamus barbarus, Heteracris adpersa, Aiolopus strepens, Oedipoda charpentieri and O. coerulescens). Double fiber-FISH in A. strepens and O. coerulescens showed that the two DNAs are closely interspersed with variable relative amounts of both classes of DNA. Finally, no correlation was observed between the number of 5S and 45S rDNA clusters in 23 species where this information was available. These results are discussed in the light of possible mechanisms of spread that led to the extensive variation in the number of clusters observed for both rDNA types in acridid grasshoppers.

  10. PR-Set7 establishes a repressive trans-tail histone code that regulates differentiation.

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    Sims, Jennifer K; Rice, Judd C

    2008-07-01

    Posttranslational modifications of the DNA-associated histone proteins play fundamental roles in eukaryotic transcriptional regulation. We previously discovered a novel trans-tail histone code involving monomethylated histone H4 lysine 20 (H4K20) and H3 lysine 9 (H3K9); however, the mechanisms that establish this code and its function in transcription were unknown. In this report, we demonstrate that H3K9 monomethylation is dependent upon the PR-Set7 H4K20 monomethyltransferase but independent of its catalytic function, indicating that PR-Set7 recruits an H3K9 monomethyltransferase to establish the trans-tail histone code. We determined that this histone code is involved in a transcriptional regulatory pathway in vivo whereby monomethylated H4K20 binds the L3MBTL1 repressor protein to repress specific genes, including RUNX1, a critical regulator of hematopoietic differentiation. The selective loss of monomethylated H4K20 at the RUNX1 promoter resulted in the displacement of L3MBTL1 and a concomitant increase in RUNX1 transcription. Importantly, the lack of monomethylated H4K20 in the human K562 multipotent cell line was specifically associated with spontaneous megakaryocytic differentiation, in part, by activating RUNX1. Our findings demonstrate that this newly described repression pathway is required for regulating proper megakaryopoiesis and suggests that it is likely to function similarly in other multipotent cell types to regulate specific differentiation pathways.

  11. Solar Simulated Ultraviolet Radiation Induces Global Histone Hypoacetylation in Human Keratinocytes.

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    Xiaoru Zhang

    Full Text Available Ultraviolet radiation (UVR from sunlight is the primary effector of skin DNA damage. Chromatin remodeling and histone post-translational modification (PTM are critical factors in repairing DNA damage and maintaining genomic integrity, however, the dynamic changes of histone marks in response to solar UVR are not well characterized. Here we report global changes in histone PTMs induced by solar simulated UVR (ssUVR. A decrease in lysine acetylation of histones H3 and H4, particularly at positions of H3 lysine 9, lysine 56, H4 lysine 5, and lysine 16, was found in human keratinocytes exposed to ssUVR. These acetylation changes were highly associated with ssUVR in a dose-dependent and time-specific manner. Interestingly, H4K16ac, a mark that is crucial for higher order chromatin structure, exhibited a persistent reduction by ssUVR that was transmitted through multiple cell divisions. In addition, the enzymatic activities of histone acetyltransferases were significantly reduced in irradiated cells, which may account for decreased global acetylation. Moreover, depletion of histone deacetylase SIRT1 in keratinocytes rescued ssUVR-induced H4K16 hypoacetylation. These results indicate that ssUVR affects both HDAC and HAT activities, leading to reduced histone acetylation.

  12. Solar Simulated Ultraviolet Radiation Induces Global Histone Hypoacetylation in Human Keratinocytes.

    Science.gov (United States)

    Zhang, Xiaoru; Kluz, Thomas; Gesumaria, Lisa; Matsui, Mary S; Costa, Max; Sun, Hong

    2016-01-01

    Ultraviolet radiation (UVR) from sunlight is the primary effector of skin DNA damage. Chromatin remodeling and histone post-translational modification (PTM) are critical factors in repairing DNA damage and maintaining genomic integrity, however, the dynamic changes of histone marks in response to solar UVR are not well characterized. Here we report global changes in histone PTMs induced by solar simulated UVR (ssUVR). A decrease in lysine acetylation of histones H3 and H4, particularly at positions of H3 lysine 9, lysine 56, H4 lysine 5, and lysine 16, was found in human keratinocytes exposed to ssUVR. These acetylation changes were highly associated with ssUVR in a dose-dependent and time-specific manner. Interestingly, H4K16ac, a mark that is crucial for higher order chromatin structure, exhibited a persistent reduction by ssUVR that was transmitted through multiple cell divisions. In addition, the enzymatic activities of histone acetyltransferases were significantly reduced in irradiated cells, which may account for decreased global acetylation. Moreover, depletion of histone deacetylase SIRT1 in keratinocytes rescued ssUVR-induced H4K16 hypoacetylation. These results indicate that ssUVR affects both HDAC and HAT activities, leading to reduced histone acetylation.

  13. Adding a Lysine Mimic in the Design of Potent Inhibitors of Histone Lysine Methyltransferases

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Yanqi; Ganesh, Thota; Horton, John R.; Spannhoff, Astrid; Liu, Jin; Sun, Aiming; Zhang, Xing; Bedford, Mark T.; Shinkai, Yoichi; Snyder, James P.; Cheng, Xiaodong (Emory); (Kyoto); (Texas)

    2010-07-19

    Dynamic histone lysine methylation involves the activities of modifying enzymes (writers), enzymes removing modifications (erasers), and readers of the histone code. One common feature of these activities is the recognition of lysines in methylated and unmethylated states, whether they are substrates, reaction products, or binding partners. We applied the concept of adding a lysine mimic to an established inhibitor (BIX-01294) of histone H3 lysine 9 methyltransferases G9a and G9a-like protein by including a 5-aminopentyloxy moiety, which is inserted into the target lysine-binding channel and becomes methylated by G9a-like protein, albeit slowly. The compound enhances its potency in vitro and reduces cell toxicity in vivo. We suggest that adding a lysine or methyl-lysine mimic should be considered in the design of small-molecule inhibitors for other methyl-lysine writers, erasers, and readers.

  14. Combinatorial H3K9acS10ph Histone Modification in IgH Locus S Regions Targets 14-3-3 Adaptors and AID to Specify Antibody Class-Switch DNA Recombination

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    Guideng Li

    2013-11-01

    Full Text Available Class-switch DNA recombination (CSR is central to the antibody response, in that it changes the immunoglobulin heavy chain (IgH constant region, thereby diversifying biological effector functions of antibodies. The activation-induced cytidine deaminase (AID-centered CSR machinery excises and rejoins DNA between an upstream (donor and a downstream (acceptor S region, which precede the respective constant region DNA. AID is stabilized on S regions by 14-3-3 adaptors. These adaptors display a high affinity for 5′-AGCT-3′ repeats, which recur in all S regions. However, how 14-3-3, AID, and the CSR machinery target exclusively the donor and acceptor S regions is poorly understood. Here, we show that histone methyltransferases and acetyltransferases are induced by CD40 or Toll-like receptor signaling and catalyze H3K4me3 and H3K9ac/K14ac histone modifications, which are enriched in S regions but do not specify the S region targets of CSR. By contrast, the combinatorial H3K9acS10ph modification specifically marks the S regions set to recombine and directly recruits 14-3-3 adaptors for AID stabilization there. Inhibition of the enzymatic activity of GCN5 and PCAF histone acetyltransferases reduces H3K9acS10ph in S regions, 14-3-3 and AID stabilization, and CSR. Thus, H3K9acS10ph is a histone code that is “written” specifically in S regions and is “read” by 14-3-3 adaptors to target AID for CSR as an important biological outcome.

  15. Inhibition of MDA-MB-231 breast cancer cell proliferation and tumor growth by apigenin through induction of G2/M arrest and histone H3 acetylation-mediated p21(WAF1/CIP1) expression.

    Science.gov (United States)

    Tseng, Tsui-Hwa; Chien, Ming-Hsien; Lin, Wea-Lung; Wen, Yu-Ching; Chow, Jyh-Ming; Chen, Chi-Kuan; Kuo, Tsang-Chih; Lee, Wei-Jiunn

    2017-02-01

    Apigenin (4',5,7-trihydroxyflavone), a flavonoid commonly found in fruits and vegetables, has anticancer properties in various malignant cancer cells. However, the molecular basis of the anticancer effect remains to be elucidated. In this study, we investigated the cellular mechanisms underlying the induction of cell cycle arrest by apigenin. Our results showed that apigenin at the nonapoptotic induction concentration inhibited cell proliferation and induced cell cycle arrest at the G2/M phase in the MDA-MB-231 breast cancer cell line. Immunoblot analysis indicated that apigenin suppressed the expression of cyclin A, cyclin B, and cyclin-dependent kinase-1 (CDK1), which control the G2-to-M phase transition in the cell cycle. In addition, apigenin upregulated p21(WAF1/CIP1) and increased the interaction of p21(WAF1/CIP1) with proliferating cell nuclear antigen (PCNA), which inhibits cell cycle progression. Furthermore, apigenin significantly inhibited histone deacetylase (HDAC) activity and induced histone H3 acetylation. The subsequent chromatin immunoprecipitation (ChIP) assay indicated that apigenin increased acetylation of histone H3 in the p21(WAF1/CIP1) promoter region, resulting in the increase of p21(WAF1/CIP1) transcription. In a tumor xenograft model, apigenin effectively delayed tumor growth. In these apigenin-treated tumors, we also observed reductions in the levels of cyclin A and cyclin B and increases in the levels of p21(WAF1/CIP1) and acetylated histone H3. These findings demonstrate for the first time that apigenin can be used in breast cancer prevention and treatment through epigenetic regulation. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 434-444, 2017. © 2016 Wiley Periodicals, Inc.

  16. Epigenetics and sex differences in the brain: A genome-wide comparison of histone-3 lysine-4 trimethylation (H3K4me3) in male and female mice.

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    Shen, Erica Y; Ahern, Todd H; Cheung, Iris; Straubhaar, Juerg; Dincer, Aslihan; Houston, Isaac; de Vries, Geert J; Akbarian, Schahram; Forger, Nancy G

    2015-06-01

    Many neurological and psychiatric disorders exhibit gender disparities, and sex differences in the brain likely explain some of these effects. Recent work in rodents points to a role for epigenetics in the development or maintenance of neural sex differences, although genome-wide studies have so far been lacking. Here we review the existing literature on epigenetics and brain sexual differentiation and present preliminary analyses on the genome-wide distribution of histone-3 lysine-4 trimethylation in a sexually dimorphic brain region in male and female mice. H3K4me3 is a histone mark primarily organized as 'peaks' surrounding the transcription start site of active genes. We microdissected the bed nucleus of the stria terminalis and preoptic area (BNST/POA) in adult male and female mice and used ChIP-Seq to compare the distribution of H3K4me3 throughout the genome. We found 248 genes and loci with a significant sex difference in H3K4me3. Of these, the majority (71%) had larger H3K4me3 peaks in females. Comparisons with existing databases indicate that genes and loci with increased H3K4me3 in females are associated with synaptic function and with expression atlases from related brain areas. Based on RT-PCR, only a minority of genes with a sex difference in H3K4me3 has detectable sex differences in expression at baseline conditions. Together with previous findings, our data suggest that there may be sex biases in the use of epigenetic marks. Such biases could underlie sex differences in vulnerabilities to drugs or diseases that disrupt specific epigenetic processes. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. CHK1-driven histone H3.3 serine 31 phosphorylation is important for chromatin maintenance and cell survival in human ALT cancer cells.

    Science.gov (United States)

    Chang, Fiona T M; Chan, F Lyn; R McGhie, James D; Udugama, Maheshi; Mayne, Lynne; Collas, Philippe; Mann, Jeffrey R; Wong, Lee H

    2015-03-11

    Human ALT cancers show high mutation rates in ATRX and DAXX. Although it is well known that the absence of ATRX/DAXX disrupts H3.3 deposition at heterochromatin, its impact on H3.3 deposition and post-translational modification in the global genome remains unclear. Here, we explore the dynamics of phosphorylated H3.3 serine 31 (H3.3S31ph) in human ALT cancer cells. While H3.3S31ph is found only at pericentric satellite DNA repeats during mitosis in most somatic human cells, a high level of H3.3S31ph is detected on the entire chromosome in ALT cells, attributable to an elevated CHK1 activity in these cells. Drug inhibition of CHK1 activity during mitosis and expression of mutant H3.3S31A in these ALT cells result in a decrease in H3.3S31ph levels accompanied with increased levels of phosphorylated H2AX serine 139 on chromosome arms and at the telomeres. Furthermore, the inhibition of CHK1 activity in these cells also reduces cell viability. Our findings suggest a novel role of CHK1 as an H3.3S31 kinase, and that CHK1-mediated H3.3S31ph plays an important role in the maintenance of chromatin integrity and cell survival in ALT cancer cells. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  18. Molecular landscape of modified histones in Drosophila heterochromatic genes and euchromatin-heterochromatin transition zones.

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    Jiro C Yasuhara

    2008-01-01

    Full Text Available Constitutive heterochromatin is enriched in repetitive sequences and histone H3-methylated-at-lysine 9. Both components contribute to heterochromatin's ability to silence euchromatic genes. However, heterochromatin also harbors hundreds of expressed genes in organisms such as Drosophila. Recent studies have provided a detailed picture of sequence organization of D. melanogaster heterochromatin, but how histone modifications are associated with heterochromatic sequences at high resolution has not been described. Here, distributions of modified histones in the vicinity of heterochromatic genes of normal embryos and embryos homozygous for a chromosome rearrangement were characterized using chromatin immunoprecipitation and genome tiling arrays. We found that H3-di-methylated-at-lysine 9 (H3K9me2 was depleted at the 5' ends but enriched throughout transcribed regions of heterochromatic genes. The profile was distinct from that of euchromatic genes and suggests that heterochromatic genes are integrated into, rather than insulated from, the H3K9me2-enriched domain. Moreover, the profile was only subtly affected by a Su(var3-9 null mutation, implicating a histone methyltransferase other than SU(VAR3-9 as responsible for most H3K9me2 associated with heterochromatic genes in embryos. On a chromosomal scale, we observed a sharp transition to the H3K9me2 domain, which coincided with increased retrotransposon density in the euchromatin-heterochromatin (eu-het transition zones on the long chromosome arms. Thus, a certain density of retrotransposons, rather than specific boundary elements, may demarcate Drosophila pericentric heterochromatin. We also demonstrate that a chromosome rearrangement that created a new eu-het junction altered H3K9me2 distribution and induced new euchromatic sites of enrichment as far as several megabases away from the breakpoint. Taken together, the findings argue against simple classification of H3K9me as the definitive signature

  19. The SET-domain protein SUVR5 mediates H3K9me2 deposition and silencing at stimulus response genes in a DNA methylation-independent manner.

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    Elena Caro

    Full Text Available In eukaryotic cells, environmental and developmental signals alter chromatin structure and modulate gene expression. Heterochromatin constitutes the transcriptionally inactive state of the genome and in plants and mammals is generally characterized by DNA methylation and histone modifications such as histone H3 lysine 9 (H3K9 methylation. In Arabidopsis thaliana, DNA methylation and H3K9 methylation are usually colocated and set up a mutually self-reinforcing and stable state. Here, in contrast, we found that SUVR5, a plant Su(var3-9 homolog with a SET histone methyltransferase domain, mediates H3K9me2 deposition and regulates gene expression in a DNA methylation-independent manner. SUVR5 binds DNA through its zinc fingers and represses the expression of a subset of stimulus response genes. This represents a novel mechanism for plants to regulate their chromatin and transcriptional state, which may allow for the adaptability and modulation necessary to rapidly respond to extracellular cues.

  20. ATRX ADD domain links an atypical histone methylation recognition mechanism to human mental-retardation syndrome

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    Iwase, Shigeki; Xiang, Bin; Ghosh, Sharmistha; Ren, Ting; Lewis, Peter W.; Cochrane, Jesse C.; Allis, C. David; Picketts, David J.; Patel, Dinshaw J.; Li, Haitao; Shi, Yang (Harvard-Med); (Ottawa Hosp.); (MSKCC); (Rockefeller); (CH-Boston); (Tsinghua); (Mass. Gen. Hosp.)

    2011-07-19

    ATR-X (alpha-thalassemia/mental retardation, X-linked) syndrome is a human congenital disorder that causes severe intellectual disabilities. Mutations in the ATRX gene, which encodes an ATP-dependent chromatin-remodeler, are responsible for the syndrome. Approximately 50% of the missense mutations in affected persons are clustered in a cysteine-rich domain termed ADD (ATRX-DNMT3-DNMT3L, ADD{sub ATRX}), whose function has remained elusive. Here we identify ADD{sub ATRX} as a previously unknown histone H3-binding module, whose binding is promoted by lysine 9 trimethylation (H3K9me3) but inhibited by lysine 4 trimethylation (H3K4me3). The cocrystal structure of ADD{sub ATRX} bound to H3{sub 1-15}K9me3 peptide reveals an atypical composite H3K9me3-binding pocket, which is distinct from the conventional trimethyllysine-binding aromatic cage. Notably, H3K9me3-pocket mutants and ATR-X syndrome mutants are defective in both H3K9me3 binding and localization at pericentromeric heterochromatin; thus, we have discovered a unique histone-recognition mechanism underlying the ATR-X etiology.

  1. ATRX ADD Domain Links an Atypical Histone Methylation Recognition Mechanism to Human Mental-Retardation Syndrome

    Energy Technology Data Exchange (ETDEWEB)

    S Iwase; B Xiang; S Ghosh; T Ren; P Lewis; J Cochrane; C Allis; D Picketts; D Patel; et al.

    2011-12-31

    ATR-X (alpha-thalassemia/mental retardation, X-linked) syndrome is a human congenital disorder that causes severe intellectual disabilities. Mutations in the ATRX gene, which encodes an ATP-dependent chromatin-remodeler, are responsible for the syndrome. Approximately 50% of the missense mutations in affected persons are clustered in a cysteine-rich domain termed ADD (ATRX-DNMT3-DNMT3L, ADD{sub ATRX}), whose function has remained elusive. Here we identify ADD{sub ATRX} as a previously unknown histone H3-binding module, whose binding is promoted by lysine 9 trimethylation (H3K9me3) but inhibited by lysine 4 trimethylation (H3K4me3). The cocrystal structure of ADD{sub ATRX} bound to H3{sub 1-15}K9me3 peptide reveals an atypical composite H3K9me3-binding pocket, which is distinct from the conventional trimethyllysine-binding aromatic cage. Notably, H3K9me3-pocket mutants and ATR-X syndrome mutants are defective in both H3K9me3 binding and localization at pericentromeric heterochromatin; thus, we have discovered a unique histone-recognition mechanism underlying the ATR-X etiology.

  2. A wide reprogramming of histone H3 modifications during male meiosis I in rice is dependent on the Argonaute protein MEL1.

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    Liu, Hua; Nonomura, Ken-Ichi

    2016-10-01

    The roles of epigenetic mechanisms, including small-RNA-mediated silencing, in plant meiosis largely remain unclear, despite their importance in plant reproduction. This study unveiled that rice chromosomes are reprogrammed during the premeiosis-to-meiosis transition in pollen mother cells (PMCs). This large-scale meiotic chromosome reprogramming (LMR) continued throughout meiosis I, during which time H3K9 dimethylation (H3K9me2) was increased, and H3K9 acetylation and H3S10 phosphorylation were broadly decreased, with an accompanying immunostaining pattern shift of RNA polymerase II. LMR was dependent on the rice Argonaute protein, MEIOSIS ARRESTED AT LEPTOTENE1 (MEL1), which is specifically expressed in germ cells prior to meiosis, because LMR was severely diminished in mel1 mutant anthers. Pivotal meiotic events, such as pre-synaptic centromere association, DNA double-strand break initiation and synapsis of homologous chromosomes, were also disrupted in this mutant. Interestingly, and as opposed to the LMR loss in most chromosomal regions, aberrant meiotic protein loading and hypermethylation of H3K9 emerged on the nucleolar organizing region in the mel1 PMCs. These results suggest that MEL1 plays important roles in epigenetic LMR to promote faithful homologous recombination and synapsis during rice meiosis. © 2016. Published by The Company of Biologists Ltd.

  3. Quantifying histone and RNA polymerase II post-translational modification dynamics in mother and daughter cells.

    Science.gov (United States)

    Stasevich, Timothy J; Sato, Yuko; Nozaki, Naohito; Kimura, Hiroshi

    2014-12-01

    Post-translational histone modifications are highly correlated with transcriptional activity, but the relative timing of these marks and their dynamic interplay during gene regulation remains controversial. To shed light on this problem and clarify the connections between histone modifications and transcription, we demonstrate how FabLEM (Fab-based Live Endogenous Modification labeling) can be used to simultaneously track histone H3 Lysine 9 acetylation (H3K9ac) together with RNA polymerase II Serine 2 and Serine 5 phosphorylation (RNAP2 Ser2ph/Ser5ph) in single living cells and their progeny. We provide a detailed description of the FabLEM methodology, including helpful tips for preparing and loading fluorescently conjugated antigen binding fragments (Fab) into cells for optimal results. We also introduce simple procedures for analyzing and visualizing FabLEM data, including color-coded scatterplots to track correlations between modifications through the cell cycle and temporal cross-correlation analysis to dissect modification dynamics. Using these methods, we find significant correlations that span cell generations, with a relatively strong correlation between H3K9ac and Ser5ph that appears to peak a few hours before mitosis and may reflect the bookmarking of genes for efficient re-initiation following mitosis. The techniques we have developed are broadly applicable and should help clarify how histone modifications dynamically contribute to gene regulation. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Dose- and time- effect responses of DNA methylation and histone H3K9 acetylation changes induced by traffic-related air pollution.

    Science.gov (United States)

    Ding, Rui; Jin, Yongtang; Liu, Xinneng; Ye, Huaizhuang; Zhu, Ziyi; Zhang, Yuan; Wang, Ting; Xu, Yinchun

    2017-03-03

    As an important risk factor of respiratory disorders, traffic-related air pollution (TRAP) has caused extensive concerns. Epigenetic change has been considered a link between TRAP and respiratory diseases. However, the exact effects of TRAP on epigenetic changes are still unclear. Here we investigated the dose- and time- effect responses of TRAP on DNA methylations and H3K9 acetylation (H3K9ac) in both blood and lung tissues of rats. The findings showed that every 1 μg/m3 increase of TRAP components were associated with changes in %5 mC (95% CI) in LINE-1, iNOS, p16CDKN2A, and APC ranging from -0.088% (-0.150, -0.026) to 0.102 (0.049, 0.154), as well as 0.276 (0.053, 0.498) to 0.475 (0.103, 0.848) ng/mg increase of H3K9ac. In addition, every 1 more day exposure at high level of TRAP (in tunnel) also significantly changed the levels of DNA methylation (ranging from -0.842% to 0.248%) and H3K9ac (16.033 and 15.718 ng/mg pro in PBMC and lung tissue, respectively) changes. Season and/or sex could interact with air pollutants in affecting DNA methylation and H3K9ac. The findings showed that TRAP exposure is dose- and time- dependently associated with the changes of DNA methylation and H3K9ac.

  5. H3K9me3-binding proteins are dispensable for SETDB1/H3K9me3-dependent retroviral silencing

    Directory of Open Access Journals (Sweden)

    Maksakova Irina A

    2011-07-01

    Full Text Available Abstract Background Endogenous retroviruses (ERVs are parasitic sequences whose derepression is associated with cancer and genomic instability. Many ERV families are silenced in mouse embryonic stem cells (mESCs via SETDB1-deposited trimethylated lysine 9 of histone 3 (H3K9me3, but the mechanism of H3K9me3-dependent repression remains unknown. Multiple proteins, including members of the heterochromatin protein 1 (HP1 family, bind H3K9me2/3 and are involved in transcriptional silencing in model organisms. In this work, we address the role of such H3K9me2/3 "readers" in the silencing of ERVs in mESCs. Results We demonstrate that despite the reported function of HP1 proteins in H3K9me-dependent gene repression and the critical role of H3K9me3 in transcriptional silencing of class I and class II ERVs, the depletion of HP1α, HP1β and HP1γ, alone or in combination, is not sufficient for derepression of these elements in mESCs. While loss of HP1α or HP1β leads to modest defects in DNA methylation of ERVs or spreading of H4K20me3 into flanking genomic sequence, respectively, neither protein affects H3K9me3 or H4K20me3 in ERV bodies. Furthermore, using novel ERV reporter constructs targeted to a specific genomic site, we demonstrate that, relative to Setdb1, knockdown of the remaining known H3K9me3 readers expressed in mESCs, including Cdyl, Cdyl2, Cbx2, Cbx7, Mpp8, Uhrf1 and Jarid1a-c, leads to only modest proviral reactivation. Conclusion Taken together, these results reveal that each of the known H3K9me3-binding proteins is dispensable for SETDB1-mediated ERV silencing. We speculate that H3K9me3 might maintain ERVs in a silent state in mESCs by directly inhibiting deposition of active covalent histone marks.

  6. H3K9me3 demethylase Kdm4d facilitates the formation of pre-initiative complex and regulates DNA replication.

    Science.gov (United States)

    Wu, Rentian; Wang, Zhiquan; Zhang, Honglian; Gan, Haiyun; Zhang, Zhiguo

    2017-01-09

    DNA replication is tightly regulated to occur once and only once per cell cycle. How chromatin, the physiological substrate of DNA replication machinery, regulates DNA replication remains largely unknown. Here we show that histone H3 lysine 9 demethylase Kdm4d regulates DNA replication in eukaryotic cells. Depletion of Kdm4d results in defects in DNA replication, which can be rescued by the expression of H3K9M, a histone H3 mutant transgene that reverses the effect of Kdm4d on H3K9 methylation. Kdm4d interacts with replication proteins, and its recruitment to DNA replication origins depends on the two pre-replicative complex components (origin recognition complex [ORC] and minichromosome maintenance [MCM] complex). Depletion of Kdm4d impairs the recruitment of Cdc45, proliferating cell nuclear antigen (PCNA), and polymerase δ, but not ORC and MCM proteins. These results demonstrate a novel mechanism by which Kdm4d regulates DNA replication by reducing the H3K9me3 level to facilitate formation of pre-initiative complex. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  7. Acetate supplementation modulates brain histone acetylation and decreases interleukin-1β expression in a rat model of neuroinflammation

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    Soliman Mahmoud L

    2012-03-01

    Full Text Available Abstract Background Long-term acetate supplementation reduces neuroglial activation and cholinergic cell loss in a rat model of lipopolysaccharide-induced neuroinflammation. Additionally, a single dose of glyceryl triacetate, used to induce acetate supplementation, increases histone H3 and H4 acetylation and inhibits histone deacetylase activity and histone deacetylase-2 expression in normal rat brain. Here, we propose that the therapeutic effect of acetate in reducing neuroglial activation is due to a reversal of lipopolysaccharide-induced changes in histone acetylation and pro-inflammatory cytokine expression. Methods In this study, we examined the effect of a 28-day-dosing regimen of glyceryl triacetate, to induce acetate supplementation, on brain histone acetylation and interleukin-1β expression in a rat model of lipopolysaccharide-induced neuroinflammation. The effect was analyzed using Western blot analysis, quantitative real-time polymerase chain reaction and enzymic histone deacetylase and histone acetyltransferase assays. Statistical analysis was performed using one-way analysis of variance, parametric or nonparametric when appropriate, followed by Tukey's or Dunn's post-hoc test, respectively. Results We found that long-term acetate supplementation increased the proportion of brain histone H3 acetylated at lysine 9 (H3K9, histone H4 acetylated at lysine 8 and histone H4 acetylated at lysine 16. However, unlike a single dose of glyceryl triacetate, long-term treatment increased histone acetyltransferase activity and had no effect on histone deacetylase activity, with variable effects on brain histone deacetylase class I and II expression. In agreement with this hypothesis, neuroinflammation reduced the proportion of brain H3K9 acetylation by 50%, which was effectively reversed with acetate supplementation. Further, in rats subjected to lipopolysaccharide-induced neuroinflammation, the pro-inflammatory cytokine interleukin-1β protein

  8. Inhibition of histone H3K9 acetylation by anacardic acid can correct the over-expression of Gata4 in the hearts of fetal mice exposed to alcohol during pregnancy.

    Science.gov (United States)

    Peng, Chang; Zhu, Jing; Sun, Hui-Chao; Huang, Xu-Pei; Zhao, Wei-An; Zheng, Min; Liu, Ling-Juan; Tian, Jie

    2014-01-01

    Cardiovascular malformations can be caused by abnormalities in Gata4 expression during fetal development. In a previous study, we demonstrated that ethanol exposure could lead to histone hyperacetylation and Gata4 over-expression in fetal mouse hearts. However, the potential mechanisms of histone hyperacetylation and Gata4 over-expression induced by ethanol remain unclear. Pregnant mice were gavaged with ethanol or saline. Fetal mouse hearts were collected for analysis. The results of ethanol fed groups showed that global HAT activity was unusually high in the hearts of fetal mice while global HDAC activity remained unchanged. Binding of P300, CBP, PCAF, SRC1, but not GCN5, were increased on the Gata4 promoter relative to the saline treated group. Increased acetylation of H3K9 and increased mRNA expression of Gata4, α-MHC, cTnT were observed in these hearts. Treatment with the pan-histone acetylase inhibitor, anacardic acid, reduced the binding of P300, PCAF to the Gata4 promoter and reversed H3K9 hyperacetylation in the presence of ethanol. Interestingly, anacardic acid attenuated over-expression of Gata4, α-MHC and cTnT in fetal mouse hearts exposed to ethanol. Our results suggest that P300 and PCAF may be critical regulatory factors that mediate Gata4 over-expression induced by ethanol exposure. Alternatively, P300, PCAF and Gata4 may coordinate over-expression of cardiac downstream genes in mouse hearts exposed to ethanol. Anacardic acid may thus protect against ethanol-induced Gata4, α-MHC, cTnT over-expression by inhibiting the binding of P300 and PCAF to the promoter region of these genes.

  9. Genome-Wide Profiling of Histone Modifications (H3K9me2 and H4K12ac and Gene Expression in Rust (Uromyces appendiculatus Inoculated Common Bean (Phaseolus vulgaris L..

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    Vasudevan Ayyappan

    Full Text Available Histone modifications such as methylation and acetylation play a significant role in controlling gene expression in unstressed and stressed plants. Genome-wide analysis of such stress-responsive modifications and genes in non-model crops is limited. We report the genome-wide profiling of histone methylation (H3K9me2 and acetylation (H4K12ac in common bean (Phaseolus vulgaris L. under rust (Uromyces appendiculatus stress using two high-throughput approaches, chromatin immunoprecipitation sequencing (ChIP-Seq and RNA sequencing (RNA-Seq. ChIP-Seq analysis revealed 1,235 and 556 histone methylation and acetylation responsive genes from common bean leaves treated with the rust pathogen at 0, 12 and 84 hour-after-inoculation (hai, while RNA-Seq analysis identified 145 and 1,763 genes differentially expressed between mock-inoculated and inoculated plants. The combined ChIP-Seq and RNA-Seq analyses identified some key defense responsive genes (calmodulin, cytochrome p450, chitinase, DNA Pol II, and LRR and transcription factors (WRKY, bZIP, MYB, HSFB3, GRAS, NAC, and NMRA in bean-rust interaction. Differential methylation and acetylation affected a large proportion of stress-responsive genes including resistant (R proteins, detoxifying enzymes, and genes involved in ion flux and cell death. The genes identified were functionally classified using Gene Ontology (GO and EuKaryotic Orthologous Groups (KOGs. The Kyoto Encyclopedia of Genes and Genomes (KEGG pathway analysis identified a putative pathway with ten key genes involved in plant-pathogen interactions. This first report of an integrated analysis of histone modifications and gene expression involved in the bean-rust interaction as reported here provides a comprehensive resource for other epigenomic regulation studies in non-model species under stress.

  10. Genome-Wide Profiling of Histone Modifications (H3K9me2 and H4K12ac) and Gene Expression in Rust (Uromyces appendiculatus) Inoculated Common Bean (Phaseolus vulgaris L.).

    Science.gov (United States)

    Ayyappan, Vasudevan; Kalavacharla, Venu; Thimmapuram, Jyothi; Bhide, Ketaki P; Sripathi, Venkateswara R; Smolinski, Tomasz G; Manoharan, Muthusamy; Thurston, Yaqoob; Todd, Antonette; Kingham, Bruce

    2015-01-01

    Histone modifications such as methylation and acetylation play a significant role in controlling gene expression in unstressed and stressed plants. Genome-wide analysis of such stress-responsive modifications and genes in non-model crops is limited. We report the genome-wide profiling of histone methylation (H3K9me2) and acetylation (H4K12ac) in common bean (Phaseolus vulgaris L.) under rust (Uromyces appendiculatus) stress using two high-throughput approaches, chromatin immunoprecipitation sequencing (ChIP-Seq) and RNA sequencing (RNA-Seq). ChIP-Seq analysis revealed 1,235 and 556 histone methylation and acetylation responsive genes from common bean leaves treated with the rust pathogen at 0, 12 and 84 hour-after-inoculation (hai), while RNA-Seq analysis identified 145 and 1,763 genes differentially expressed between mock-inoculated and inoculated plants. The combined ChIP-Seq and RNA-Seq analyses identified some key defense responsive genes (calmodulin, cytochrome p450, chitinase, DNA Pol II, and LRR) and transcription factors (WRKY, bZIP, MYB, HSFB3, GRAS, NAC, and NMRA) in bean-rust interaction. Differential methylation and acetylation affected a large proportion of stress-responsive genes including resistant (R) proteins, detoxifying enzymes, and genes involved in ion flux and cell death. The genes identified were functionally classified using Gene Ontology (GO) and EuKaryotic Orthologous Groups (KOGs). The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis identified a putative pathway with ten key genes involved in plant-pathogen interactions. This first report of an integrated analysis of histone modifications and gene expression involved in the bean-rust interaction as reported here provides a comprehensive resource for other epigenomic regulation studies in non-model species under stress.

  11. Nucleosomal histone proteins of L. donovani: a combination of recombinant H2A, H2B, H3 and H4 proteins were highly immunogenic and offered optimum prophylactic efficacy against Leishmania challenge in hamsters.

    Science.gov (United States)

    Baharia, Rajendra K; Tandon, Rati; Sahasrabuddhe, Amogh A; Sundar, Shyam; Dube, Anuradha

    2014-01-01

    The present study includes cloning and expression of recombinant Leishmania donovani histone proteins (rLdH2B, rLdH3, rLdH2A and rLdH4), assessment of their immunogenicity in Leishmania infected cured patients/endemic contacts as well as in cured hamsters and finally evaluation of their prophylactic efficacy in hamsters against L. donovani challenge. All recombinant proteins were expressed and purified from the heterologous bacterial host system. Leishmania infected cured patients/endemic contacts as well as cured hamsters exhibited significantly higher proliferative responses to individual recombinant histones and their pooled combination (rLdH2B+rLdH3+rLdH2A+rLdH4) than those of L.donovani infected hosts. The L.donovani soluble antigens (SLD) stimulated PBMCs of cured/exposed and Leishmania patients to produce a mixed Thl/Th2-type cytokine profile, whereas rLdH2B, rLdH3, rLdH2A, rLdH4 and pooled combination (rLdH2-4) stimulated the production of Th1 cytokines IFN-γ, IL-12 and TNF-α but not Th2 cytokines IL-4 or IL-10. The immunogenicity of these histone proteins along with their combination was also checked in cured hamsters where they stimulated higher lymphoproliferation and Nitric oxide production in lymphocytes of cured hamsters than that of infected controls. Moreover, significantly increased IgG2 response, an indicative of cell mediated immunity, was observed in cured hamsters against these individual proteins and their combination as compared to infected hamsters. Further, it was demonstrated that rLdH2B, rLdH3, rLdH2A and rLdH4 and pooled combination were able to provide considerable protection for hamsters against L. donovani challenge. The efficacy was supported by the increased inducible Nitric Oxide Synthase (iNOS) mRNA transcripts and Th1-type cytokines--IFN-γ, IL-12 and TNF-α and down-regulation of IL-4, IL-10 and TGF-β. Hence, it is inferred that pooled rLdH2-4 elicits Thl-type of immune responses exclusively and confer considerable protection

  12. H3K9 methylation regulates growth and development in Aspergillus fumigatus.

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    Palmer, Jonathan M; Perrin, Robyn M; Dagenais, Taylor R T; Keller, Nancy P

    2008-12-01

    In most species, chromatin remodeling mediates critical biological processes ranging from development to disease states. In fungi within the genus Aspergillus, chromatin remodeling may regulate expression of metabolic gene clusters, but other processes regulated by chromatin structure remain to be elucidated. In many eukaryotic species, methylation of lysine 9 of histone 3 (H3K9) is a hallmark of heterochromatin formation and subsequent gene silencing. The sole H3K9 methyltransferase in Schizosaccharomyces pombe is Clr4. We report that disruption of the Clr4 homolog in the pathogenic mold Aspergillus fumigatus (ClrD), which is involved in both mono- and trimethylation of H3K9, results in several growth abnormalities. Developmental defects in DeltaAfclrD include reduction in radial growth, reduction in conidial production, and delayed conidiation after developmental competence mediated by delayed expression of brlA, the master regulator of conidiophore development. Sensitivity of DeltaAfclrD to 6-azauracil suggests that ClrD influences transcriptional processing in A. fumigatus. Despite growth abnormalities, macrophage assays suggest ClrD may be dispensable for host interactions.

  13. The epigenetic effects of aspirin: the modification of histone H3 lysine 27 acetylation in the prevention of colon carcinogenesis in azoxymethane- and dextran sulfate sodium-treated CF-1 mice.

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    Guo, Yue; Liu, Yue; Zhang, Chengyue; Su, Zheng-Yuan; Li, Wenji; Huang, Mou-Tuan; Kong, Ah-Ng

    2016-06-01

    Colorectal cancer (CRC) is the third most common cancer worldwide. Chronic inflammation appears to enhance the risk of CRC. Emerging evidence has suggested that epigenetic mechanisms play an important role in CRC. Aspirin [acetylsalicylic acid (ASA)] has been shown to prevent CRC; however, the epigenetic mechanisms of its action remain unknown. This study investigated the protective role of ASA in azoxymethane (AOM)-initiated and dextran sulfate sodium (DSS)-promoted colitis-associated colon cancer (CAC) and examined the epigenetic effects, particularly on histone 3 lysine 27 acetylation (H3K27ac), underlying the preventive effect of ASA. CF-1 mice were fed with AIN-93M diet with or without 0.02% ASA from 1 week prior to AOM initiation until the mice were killed 20 weeks after AOM injection. Our results showed that AOM/DSS + ASA significantly suppressed inflammatory colitis symptoms and tumor multiplicity. AOM/DSS + ASA reduced AOM/DSS-induced protein expression and the activity of histone deacetylases (HDACs) and globally restored H3K27ac. Furthermore, AOM/DSS + ASA inhibited AOM/DSS-induced enrichment of H3K27ac in the promoters of inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) that corresponded to the dramatic suppression of the messenger RNA (mRNA) and protein levels. Surprisingly, no significant changes in the H3K27ac abundance in the prostaglandin-endoperoxide synthase 2 (Cox-2) promoters or in the Cox-2 mRNA and protein expression were observed. Collectively, our results suggest that a potential novel epigenetic mechanism underlies the chemopreventive effects of ASA, and this mechanism attenuates CAC in AOM/DSS-induced CF-1 mice via the inhibition of HDACs and the modification of H3K27ac marks that suppress iNOS, TNF-α and IL-6. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  14. Repeated forced swim stress prior to complete Freund's adjuvant injection enhances mechanical hyperalgesia and attenuates the expression of pCREB and ΔFosB and the acetylation of histone H3 in the insular cortex of rat.

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    Imbe, H; Kimura, A

    2015-08-20

    Exposure to stressors causes substantial effects on the perception and response to pain. In several animal models, chronic stress produces hyperalgesia. The insular (IC) and anterior cingulate cortices (ACC) are the regions exhibiting most reliable pain-related activity. And the IC and ACC play an important role in pain modulation via descending pain modulatory system. In the present study we examined the expression of phospho-cAMP response element-binding protein (pCREB) and ΔFosB and the acetylation of histone H3 in the IC and ACC after forced swim stress (FS) and complete Freund's adjuvant (CFA) injection to clarify changes in the cerebral cortices that affect the activity of the descending pain modulatory system in rats with stress-induced hyperalgesia. CFA injection into the hindpaw or FS (day 1, 10min; days 2-3, 20min) induced a significant increase in the expression of pCREB and ΔFosB and the acetylation of histone H3 in the IC. Quantitative image analysis showed that the numbers of ΔFosB-immunoreactivity (IR) cells in the bilateral anterior and posterior IC (AIC and PIC) were significantly higher in the CFA group (AIC R, 548.0±98.6; AIC L, 433.5±89.4; PIC R, 546.1±72.8; PIC L, 415.5±53.5) than those in the naive group (AIC R, 86.6±14.8; AIC L, 85.5±24.7; PIC R, 124.5±29.9; PIC L, 107.0±19.8, p<0.01). However the FS prior to the CFA injection enhanced the mechanical hyperalgesia and attenuated the expression of pCREB and ΔFosB and the acetylation of histone H3 in the IC. There was no significant difference in the numbers of ΔFosB-IR cells in the bilateral PIC between the FS+CFA and naive groups. These findings suggest neuroplasticity in the IC after the FS, which may be involved in the enhancement of CFA-induced mechanical hyperalgesia through dysfunction of the descending pain modulatory system. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

  15. DNA methylation in an intron of the IBM1 histone demethylase gene stabilizes chromatin modification patterns.

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    Rigal, Mélanie; Kevei, Zoltán; Pélissier, Thierry; Mathieu, Olivier

    2012-06-29

    The stability of epigenetic patterns is critical for genome integrity and gene expression. This highly coordinated process involves interrelated positive and negative regulators that impact distinct epigenetic marks, including DNA methylation and dimethylation at histone H3 lysine 9 (H3K9me2). In Arabidopsis, mutations in the DNA methyltransferase MET1, which maintains CG methylation, result in aberrant patterns of other epigenetic marks, including ectopic non-CG methylation and the relocation of H3K9me2 from heterochromatin into gene-rich chromosome regions. Here, we show that the expression of the H3K9 demethylase IBM1 (increase in BONSAI methylation 1) requires DNA methylation. Surprisingly, the regulatory methylated region is contained in an unusually large intron that is conserved in IBM1 orthologues. The re-establishment of IBM1 expression in met1 mutants restored the wild-type H3K9me2 nuclear patterns, non-CG DNA methylation and transcriptional patterns at selected loci, which included DNA demethylase genes. These results provide a mechanistic explanation for long-standing puzzling observations in met1 mutants and reveal yet another layer of control in the interplay between DNA methylation and histone modification, which stabilizes DNA methylation patterns at genes.

  16. Abnormal Histone Methylation is Responsible for Increased VEGF165a Secretion from Airway Smooth Muscle Cells in Asthma

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    Clifford, Rachel L.; John, Alison E.; Brightling, Christopher E.; Knox, Alan J.

    2012-01-01

    Vascular Endothelial Growth Factor (VEGF), a key angiogenic molecule, is aberrantly expressed in several diseases including asthma where it contributes to bronchial vascular remodelling and chronic inflammation. Asthmatic human airway smooth muscle (HASM) cells hypersecrete VEGF but the mechanism is unclear. Here we defined the mechanism in HASM cells from non-asthmatic (NA) and asthmatic (A) patients. We found that asthmatic cells lacked a repression complex at the VEGF promoter which was present in non-asthmatic cells. Recruitment of G9A, trimethylation of histone H3 at lysine 9 (H3K9me3) and a resultant decrease in RNA polymerase II (RNA pol II) at the VEGF promoter was critical to repression of VEGF secretion in non-asthmatic cells. At the asthmatic promoter H3K9me3 was absent due to failed recruitment of G9a; RNA pol II binding, in association with TAF1, was increased, H3K4me3 was present and Sp1 binding was exaggerated and sustained. In contrast DNA methylation and histone acetylation were similar in A and NA cells. This is the first study to show that airway cells in asthma have altered epigenetic regulation of remodelling gene(s). Histone methylation at genes such as VEGF may be an important new therapeutic target. PMID:22689881

  17. G9a-mediated histone methylation regulates cadmium-induced male fertility damage in pubertal mice.

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    Li, Min; Liu, Chuan; Yang, Lingling; Zhang, Lei; Chen, Chunhai; He, Mindi; Lu, Yonghui; Feng, Wei; Pi, Huifeng; Zhang, Yanwen; Zhong, Min; Yu, Zhengping; Zhou, Zhou

    2016-06-11

    Increasing evidence suggests that cadmium (Cd) is associated with male fertility damage. However, the effects of histone modification on Cd-induced male fertility damage remain obscure. This study aims to evaluate the roles of histone methylation mediated by euchromatin histone methyltransferase (EHMT2/G9a) in regulating Cd-induced male fertility damage. We exposed 4-week-old male C57BL/6J mice to Cd by intraperitoneal injection at 2mg/kg for 1, 3 and 5days. Our data showed that Cd exposure decreased the numbers of impregnated females and litter sizes, which was concomitant with sperm count reduction, histological changes in the cauda epididymal ducts and seminiferous epithelium, and testicular cell apoptosis as evaluated by terminal dUTP nick-end labeling (TUNEL) assay and immunoblotting with increased levels of cleaved caspase 3, PARP and Bax and a decreased level of Bcl-2. Cd-induced male fertility damage was accompanied by enhanced G9a activity followed by increased histone H3 lysine 9 monomethylation (H3K9me1) and dimethylation (H3K9me2) in testes. Furthermore, inhibition of G9a by BIX-01294 normalized H3K9me1 and H3K9me2 to basal levels and prevented Cd-induced male fertility damage and testicular cell apoptosis. Our present study revealed that G9a-mediated histone methylation plays a critical role in Cd-induced male fertility damage and testicular cell apoptosis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  18. KDM4C, a H3K9me3 Histone Demethylase, is Involved in the Maintenance of Human ESCC-Initiating Cells by Epigenetically Enhancing SOX2 Expression

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    Xiang Yuan

    2016-10-01

    Full Text Available Our studies investigating the existence of tumor-initiating cell (TIC populations in human esophageal squamous cell carcinoma (ESCC had identified a subpopulation of cells isolated from ESCC patient-derived tumor specimens marked by an ALDHbri+ phenotype bear stem cell-like features. Importantly, KDM4C, a histone demethylase was enhanced in ALDHbri+ subpopulation, suggesting that strategies interfering with KDM4C may be able to target these putative TICs. In the present study, by genetic and chemical means, we demonstrated that, KDM4C blockade selectively decreased the ESCC ALDHbri+ TICs population in vitro and specifically targeted the TICs in ALDHbri+-derived xenograft, retarding engraftment. Subsequent studies of the KDM4C functional network identified a subset of pluripotency-associated genes (PAGs and aldehyde dehydrogenase family members to be preferentially down-regulated in KDM4C inhibited ALDHbri+ TICs. We further supported a model in which KDM4C maintains permissive histone modifications with a low level of H3K9 methylation at the promoters of several PAGs. Moreover, ectopic expression of SOX2 restored KDM4C inhibition-dependent ALDHbri+ TIC properties. We further confirmed these findings by showing that the cytoplasmic and nuclear KDM4C staining increased with adverse pathologic phenotypes and poor patient survival. Such staining pattern of intracellular KDM4C appeared to overlap with the expression of SOX2 and ALDH1. Collectively, our findings provided the insights into the development of novel therapeutic strategies based on the inhibition of KDM4C pathway for the eliminating of ESCC TIC compartment.

  19. Dynamic and flexible H3K9me3 bridging via HP1β dimerization establishes a plastic state of condensed chromatin

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    Hiragami-Hamada, Kyoko; Soeroes, Szabolcs; Nikolov, Miroslav; Wilkins, Bryan; Kreuz, Sarah; Chen, Carol; De La Rosa-Velázquez, Inti A.; Zenn, Hans Michael; Kost, Nils; Pohl, Wiebke; Chernev, Aleksandar; Schwarzer, Dirk; Jenuwein, Thomas; Lorincz, Matthew; Zimmermann, Bastian; Walla, Peter Jomo; Neumann, Heinz; Baubec, Tuncay; Urlaub, Henning; Fischle, Wolfgang

    2016-01-01

    Histone H3 trimethylation of lysine 9 (H3K9me3) and proteins of the heterochromatin protein 1 (HP1) family are hallmarks of heterochromatin, a state of compacted DNA essential for genome stability and long-term transcriptional silencing. The mechanisms by which H3K9me3 and HP1 contribute to chromatin condensation have been speculative and controversial. Here we demonstrate that human HP1β is a prototypic HP1 protein exemplifying most basal chromatin binding and effects. These are caused by dimeric and dynamic interaction with highly enriched H3K9me3 and are modulated by various electrostatic interfaces. HP1β bridges condensed chromatin, which we postulate stabilizes the compacted state. In agreement, HP1β genome-wide localization follows H3K9me3-enrichment and artificial bridging of chromatin fibres is sufficient for maintaining cellular heterochromatic conformation. Overall, our findings define a fundamental mechanism for chromatin higher order structural changes caused by HP1 proteins, which might contribute to the plastic nature of condensed chromatin. PMID:27090491

  20. Evaluating the role of epigenetic histone modifications in the metabolic memory of type 1 diabetes.

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    Miao, Feng; Chen, Zhuo; Genuth, Saul; Paterson, Andrew; Zhang, Lingxiao; Wu, Xiwei; Li, Sierra Min; Cleary, Patricia; Riggs, Arthur; Harlan, David M; Lorenzi, Gayle; Kolterman, Orville; Sun, Wanjie; Lachin, John M; Natarajan, Rama

    2014-05-01

    We assessed whether epigenetic histone posttranslational modifications are associated with the prolonged beneficial effects (metabolic memory) of intensive versus conventional therapy during the Diabetes Control and Complications Trial (DCCT) on the progression of microvascular outcomes in the long-term Epidemiology of Diabetes Interventions and Complications (EDIC) study. We performed chromatin immunoprecipitation linked to promoter tiling arrays to profile H3 lysine-9 acetylation (H3K9Ac), H3 lysine-4 trimethylation (H3K4Me3), and H3K9Me2 in blood monocytes and lymphocytes obtained from 30 DCCT conventional treatment group subjects (case subjects: mean DCCT HbA1c level >9.1% [76 mmol/mol] and progression of retinopathy or nephropathy by EDIC year 10 of follow-up) versus 30 DCCT intensive treatment subjects (control subjects: mean DCCT HbA1c level 15 genes related to the nuclear factor-κB inflammatory pathway and were enriched in genes related to diabetes complications. These results suggest an association between HbA1c level and H3K9Ac, and a possible epigenetic explanation for metabolic memory in humans.

  1. Molecular Targets of Chromatin Repressive Mark H3K9me3 in Primate Progenitor Cells within Adult Neurogenic Niches

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    Michael R Foret

    2014-07-01

    Full Text Available Histone 3 Lysine 9 (H3K9 methylation is known to be associated with pericentric heterochromatin and important in genomic stability. In this study, we show that trimethylation at H3K9 (H3K9me3 is enriched in an adult neural stem cell niche- the subventricular zone (SVZ on the walls of the lateral ventricle in both rodent and non-human primate baboon brain. Previous studies have shown that there is significant correlation between baboon and human regarding genomic similarity and brain structure, suggesting that findings in baboon are relevant to human. To understand the function of H3K9me3 in this adult neurogenic niche, we performed genome-wide analyses using ChIP-Seq (chromatin immunoprecipitation and deep-sequencing and RNA-Seq for in vivo SVZ cells purified from baboon brain. Through integrated analyses of ChIP-Seq and RNA-Seq, we found that H3K9me3-enriched genes associated with cellular maintenance, post-transcriptional and translational modifications, signaling pathways, and DNA replication are expressed, while genes involved in axon/neuron, hepatic stellate cell, or immune-response activation are not expressed. As neurogenesis progresses in the adult SVZ, cell fate restriction is essential to direct proper lineage commitment. Our findings highlight that H3K9me3 repression in undifferentiated SVZ cells is engaged in the maintenance of cell type integrity, implicating a role for H3K9me3 as an epigenetic mechanism to control cell fate transition within this adult germinal niche.

  2. H3K36 Methylation Antagonizes PRC2-mediated H3K27 Methylation*

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    Yuan, Wen; Xu, Mo; Huang, Chang; Liu, Nan; Chen, She; Zhu, Bing

    2011-01-01

    H3K27 methylation mediated by the histone methyltransferase complex PRC2 is critical for transcriptional regulation, Polycomb silencing, Drosophila segmentation, mammalian X chromosome inactivation, and cancer. PRC2-mediated H3K27 methylation can spread along the chromatin and propagate the repressive chromatin environment; thus, chromatin components that antagonize the activity of PRC2 are important for restraining Polycomb silencing. Here we report that in HeLa cells, H3 histones unmethylat...

  3. Data for the identification of proteins and post-translational modifications of proteins associated to histones H3 and H4 in S. cerevisiae, using tandem affinity purification coupled with mass spectrometry.

    Science.gov (United States)

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

    2016-03-01

    Tandem affinity purification method (TAP) allows the efficient purification of native protein complexes which incorporate a target protein fused with the TAP tag. Purified multiprotein complexes can then be subjected to diverse types of proteomic analyses. Here we describe the data acquired after applying the TAP strategy on histones H3 and H4 coupled with mass spectrometry to identify associated proteins and protein post-translational modifications in the budding yeast, Saccharomyces cerevisiae. The mass spectrometry dataset described here consists of 14 files generated from four different analyses in a 5600 Triple TOF (Sciex) by information-dependent acquisition (IDA) LC-MS/MS. The above files contain information about protein identification, protein relative abundance, and PTMs identification. The instrumental raw data from these files has been also uploaded to the ProteomeXchange Consortium via the PRIDE partner repository, with the dataset identifier PRIDE: PXD002671 and http://dx.doi.org/10.6019/PXD002671. These data are discussed and interpreted in http://dx.doi.org/10.1016/j.jprot.2016.01.004. Valero et al. (2016) [1].

  4. Abnormal histone methylation is responsible for increased vascular endothelial growth factor 165a secretion from airway smooth muscle cells in asthma.

    Science.gov (United States)

    Clifford, Rachel L; John, Alison E; Brightling, Christopher E; Knox, Alan J

    2012-07-15

    Vascular endothelial growth factor (VEGF), a key angiogenic molecule, is aberrantly expressed in several diseases including asthma where it contributes to bronchial vascular remodeling and chronic inflammation. Asthmatic human airway smooth muscle cells hypersecrete VEGF, but the mechanism is unclear. In this study, we defined the mechanism in human airway smooth muscle cells from nonasthmatic and asthmatic patients. We found that asthmatic cells lacked a repression complex at the VEGF promoter, which was present in nonasthmatic cells. Recruitment of G9A, trimethylation of histone H3 at lysine 9 (H3K9me3), and a resultant decrease in RNA polymerase II at the VEGF promoter was critical to repression of VEGF secretion in nonasthmatic cells. At the asthmatic promoter, H3K9me3 was absent because of failed recruitment of G9a; RNA polymerase II binding, in association with TATA-binding protein-associated factor 1, was increased; H3K4me3 was present; and Sp1 binding was exaggerated and sustained. In contrast, DNA methylation and histone acetylation were similar in asthmatic and nonasthmatic cells. This is the first study, to our knowledge, to show that airway cells in asthma have altered epigenetic regulation of remodeling gene(s). Histone methylation at genes such as VEGF may be an important new therapeutic target.

  5. Involvement of Histone Lysine Methylation in p21 Gene Expression in Rat Kidney In Vivo and Rat Mesangial Cells In Vitro under Diabetic Conditions

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    Xiangjun Li

    2016-01-01

    Full Text Available Diabetic nephropathy (DN, a common complication associated with type 1 and type 2 diabetes mellitus (DM, characterized by glomerular mesangial expansion, inflammation, accumulation of extracellular matrix (ECM protein, and hypertrophy, is the major cause of end-stage renal disease (ESRD. Increasing evidence suggested that p21-dependent glomerular and mesangial cell (MC hypertrophy play key roles in the pathogenesis of DN. Recently, posttranscriptional modifications (PTMs have uncovered novel molecular mechanisms involved in DN. However, precise regulatory mechanism of histone lysine methylation (HKme mediating p21 related hypertrophy associated with DN is not clear. We evaluated the roles of HKme and histone methyltransferase (HMT SET7/9 in p21 gene expression in glomeruli of diabetic rats and in high glucose- (HG- treated rat mesangial cells (RMCs. p21 gene expression was upregulated in diabetic rats glomeruli; chromatin immunoprecipitation (ChIP assays showed decreased histone H3-lysine9-dimethylation (H3K9me2 accompanied with enhanced histone H3-lysine4-methylation (H3K4me1/3 and SET7/9 occupancies at the p21 promoter. HG-treated RMCs exhibited increased p21 mRNA, H3K4me level, SET7/9 recruitment, and inverse H3K9me, which were reversed by TGF-β1 antibody. These data uncovered key roles of H3Kme and SET7/9 responsible for p21 gene expression in vivo and in vitro under diabetic conditions and confirmed preventive effect of TGF-β1 antibody on DN.

  6. FGF2 is a target and a trigger of epigenetic mechanisms associated with differences in emotionality: partnership with H3K9me3.

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    Chaudhury, Sraboni; Aurbach, Elyse L; Sharma, Vikram; Blandino, Peter; Turner, Cortney A; Watson, Stanley J; Akil, Huda

    2014-08-12

    Posttranslational modifications of histone tails in chromatin template can result from environmental experiences such as stress and substance abuse. However, the role of epigenetic modifications as potential predisposing factors in affective behavior is less well established. To address this question, we used our selectively bred lines of high responder (bHR) and low responder (bLR) rats that show profound and stable differences in affective responses, with bLRs being prone to anxiety- and depression-like behavior and bHRs prone to addictive behavior. We first asked whether these phenotypes are associated with basal differences in epigenetic profiles. Our results reveal broad between-group differences in basal levels of trimethylated histone protein H3 at lysine 9 (H3K9me3) in hippocampus (HC), amygdala, and nucleus accumbens. Moreover, levels of association of H3K9me3 at Glucocorticoid Receptor (GR) and Fibroblast growth Factor 2 (FGF2) promoters differ reciprocally between bHRs and bLRs in these regions, consistent with these genes' opposing levels of expression and roles in modulating anxiety behavior. Importantly, this basal epigenetic pattern is modifiable by FGF2, a factor that modulates anxiety behavior. Thus, early-life FGF2, which decreases anxiety, altered the levels of H3K9me3 and its binding at FGF2 and GR promoters of bLRs rendering them more similar to bHRs. Conversely, knockdown of HC FGF2 altered both anxiety behavior and levels of H3K9me3 in bHRs, rendering them more bLR-like. These findings implicate FGF2 as a modifier of epigenetic mechanisms associated with emotional responsiveness, and point to H3K9me3 as a key player in the regulation of affective vulnerability.

  7. Epigenetic regulation of oogenesis and germ stem cell maintenance by the Drosophila histone methyltransferase Eggless/dSetDB1.

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    Clough, Emily; Tedeschi, Thomas; Hazelrigg, Tulle

    2014-04-15

    The Drosophila melanogaster histone lysine methyltransferase (HKMT) Eggless (Egg/dSETDB1) catalyzes methylation of Histone H3 lysine 9 (H3K9), a signature of repressive heterochromatin. Our previous studies showed that H3K9 methylation by Egg is required for oogenesis. Here we analyze a set of EMS-induced mutations in the egg gene, identify the molecular lesions of these mutations, and compare the effects on oogenesis of both strong loss-of-function and weak hypomorphic alleles. These studies show that H3K9 methylation by Egg is required for multiple stages of oogenesis. Mosaic expression experiments show that the egg gene is not required intrinsically in the germ cells for their early differentiation, but is required in the germ cells for their survival past stage 5 of oogenesis. egg is also required in germ stem cells for their maintenance, since egg- germ stem cells initially survive but are not maintained as females age. Mosaic analysis also reveals that the early egg chamber budding defects in egg- ovaries are due to an intrinsic requirement for egg in follicle stem cells and their descendents, and that egg plays a non-autonomous role in somatic cells in the germarium to influence the differentiation of early germ cells. Copyright © 2014. Published by Elsevier Inc.

  8. Effect of histone acetylation modification with MGCD0103, a histone deacetylase inhibitor, on nuclear reprogramming and the developmental competence of porcine somatic cell nuclear transfer embryos.

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    Jin, Long; Zhu, Hai-Ying; Guo, Qing; Li, Xiao-Chen; Zhang, Yu-Chen; Cui, Cheng-Du; Li, Wen-Xue; Cui, Zheng-Yun; Yin, Xi-Jun; Kang, Jin-Dan

    2017-01-01

    Cloning remains as an important technique to enhance the reconstitution and distribution of animal population with high-genetic merit. One of the major detrimental factors of this technique is the abnormal epigenetic modifications. MGCD0103 is known as a histone deacetylase inhibitor. In this study, we investigated the effect of MGCD0103 on the in vitro blastocyst formation rate in porcine somatic cell nuclear transferred (SCNT) embryos and expression in acetylation of the histone H3 lysine 9 and histone H4 lysine 12. We compared the in vitro embryonic development of SCNT embryos treated with different concentrations of MGCD0103 for 24 hours. Our results reported that treating with 0.2-μM MGCD0103 for 24 hours effectively improved the development of SCNT embryos, in comparison to the control group (blastocyst formation rate, 25.5 vs. 10.7%, P MGCD0103 for various intervals after activation. Treatment for 6 hours significantly improved the development of pig SCNT embryos, compared with the control group (blastocyst formation rate, 21.2 vs. 10.5%, P MGCD0103 supplementation significantly (P MGCD0103-treated SCNT embryos were transferred into two surrogate sows, one of whom became pregnant and three fetuses developed. These results suggest that MGCD0103 can enhance the nuclear reprogramming and improve in vitro developmental potential of porcine SCNT embryos. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Hypoxia-induced down-regulation of neprilysin by histone modification in mouse primary cortical and hippocampal neurons.

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    Zheng Wang

    Full Text Available Amyloid β-peptide (Aβ accumulation leads to neurodegeneration and Alzheimer's disease (AD. Aβ metabolism is a dynamic process in the Aβ production and clearance that requires neprilysin (NEP and other enzymes to degrade Aβ. It has been reported that NEP expression is significantly decreased in the brain of AD patients. Previously we have documented hypoxia is a risk factor for Aβ generation in vivo and in vitro through increasing Aβ generation by altering β-cleavage and γ-cleavage of APP and down-regulating NEP, and causing tau hyperphosphorylation. Here, we investigated the molecular mechanisms of hypoxia-induced down-regulation of NEP. We found a significant decrease in NEP expression at the mRNA and protein levels after hypoxic treatment in mouse primary cortical and hippocampal neurons. Chromatin immunoprecipitation (ChIP assays and relative quantitative PCR (q-PCR revealed an increase of histone H3-lysine9 demethylation (H3K9me2 and a decrease of H3 acetylation (H3-Ace in the NEP promoter regions following hypoxia. In addition, we found that hypoxia caused up-regulation of histone methyl transferase (HMT G9a and histone deacetylases (HDACs HDAC-1. Decreased expression of NEP during hypoxia can be prevented by application with the epigenetic regulators 5-Aza-2'-deoxycytidine (5-Aza, HDACs inhibitor sodium valproate (VA, and siRNA-mediated knockdown of G9a or HDAC1. DNA methylation PCR data do not support that hypoxia affects the methylation of NEP promoters. This study suggests that hypoxia may down-regulate NEP by increasing H3K9me2 and decreasing H3-Ace modulation.

  10. DNA methylation and histone modifications regulate de novo shoot regeneration in Arabidopsis by modulating WUSCHEL expression and auxin signaling.

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    Wei Li

    2011-08-01

    Full Text Available Plants have a profound capacity to regenerate organs from differentiated somatic tissues, based on which propagating plants in vitro was made possible. Beside its use in biotechnology, in vitro shoot regeneration is also an important system to study de novo organogenesis. Phytohormones and transcription factor WUSCHEL (WUS play critical roles in this process but whether and how epigenetic modifications are involved is unknown. Here, we report that epigenetic marks of DNA methylation and histone modifications regulate de novo shoot regeneration of Arabidopsis through modulating WUS expression and auxin signaling. First, functional loss of key epigenetic genes-including METHYLTRANSFERASE1 (MET1 encoding for DNA methyltransferase, KRYPTONITE (KYP for the histone 3 lysine 9 (H3K9 methyltransferase, JMJ14 for the histone 3 lysine 4 (H3K4 demethylase, and HAC1 for the histone acetyltransferase-resulted in altered WUS expression and developmental rates of regenerated shoots in vitro. Second, we showed that regulatory regions of WUS were developmentally regulated by both DNA methylation and histone modifications through bisulfite sequencing and chromatin immunoprecipitation. Third, DNA methylation in the regulatory regions of WUS was lost in the met1 mutant, thus leading to increased WUS expression and its localization. Fourth, we did a genome-wide transcriptional analysis and found out that some of differentially expressed genes between wild type and met1 were involved in signal transduction of the phytohormone auxin. We verified that the increased expression of AUXIN RESPONSE FACTOR3 (ARF3 in met1 indeed was due to DNA demethylation, suggesting DNA methylation regulates de novo shoot regeneration by modulating auxin signaling. We propose that DNA methylation and histone modifications regulate de novo shoot regeneration by modulating WUS expression and auxin signaling. The study demonstrates that, although molecular components involved in organogenesis

  11. Pharmacological reversal of histone methylation presensitizes pancreatic cancer cells to nucleoside drugs: in vitro optimization and novel nanoparticle delivery studies.

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    Sau Wai Hung

    Full Text Available We evaluated the potential of an investigational histone methylation reversal agent, 3-deazaneplanocin A (DZNep, in improving the chemosensitivity of pancreatic cancer to nucleoside analogs (i.e., gemcitabine. DZNep brought delayed but selective cytotoxicity to pancreatic cancer cells without affecting normal human pancreatic ductal epithelial (HPDE cells. Co-exposure of DZNep and gemcitabine induced cytotoxic additivity or synergism in both well- and poorly-differentiated pancreatic cell lines by increased apoptosis. In contrast, DZNep exerted antagonism with gemcitabine against HPDE cells with significant reduction in cytotoxicity compared with the gemcitabine-alone regimen. DZNep marginally depended on purine nucleoside transporters for its cytotoxicity, but the transport dependence was circumvented by acyl derivatization. Drug exposure studies revealed that a short priming with DZNep followed by gemcitabine treatment rather than co-treatment of both agents to produce a maximal chemosensitization response in both gemcitabine-sensitive and gemcitabine-resistant pancreatic cancer cells. DZNep rapidly and reversibly decreased trimethylation of histone H3 lysine 27 but increased trimethylation of lysine 9 in an EZH2- and JMJD1A/2C-dependent manner, respectively. However, DZNep potentiation of nucleoside analog chemosensitization was found to be temporally coupled to trimethylation changes in lysine 27 and not lysine 9. Polymeric nanoparticles engineered to chronologically release DZNep followed by gemcitabine produced pronounced chemosensitization and dose-lowering effects. Together, our results identify that an optimized DZNep exposure can presensitize pancreatic cancer cells to anticancer nucleoside analogs through the reversal of histone methylation, emphasizing the promising clinical utilities of epigenetic reversal agents in future pancreatic cancer combination therapies.

  12. H3 lysine 4 is acetylated at active gene promoters and is regulated by H3 lysine 4 methylation.

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    Benoit Guillemette

    2011-03-01

    Full Text Available Methylation of histone H3 lysine 4 (H3K4me is an evolutionarily conserved modification whose role in the regulation of gene expression has been extensively studied. In contrast, the function of H3K4 acetylation (H3K4ac has received little attention because of a lack of tools to separate its function from that of H3K4me. Here we show that, in addition to being methylated, H3K4 is also acetylated in budding yeast. Genetic studies reveal that the histone acetyltransferases (HATs Gcn5 and Rtt109 contribute to H3K4 acetylation in vivo. Whilst removal of H3K4ac from euchromatin mainly requires the histone deacetylase (HDAC Hst1, Sir2 is needed for H3K4 deacetylation in heterochomatin. Using genome-wide chromatin immunoprecipitation (ChIP, we show that H3K4ac is enriched at promoters of actively transcribed genes and located just upstream of H3K4 tri-methylation (H3K4me3, a pattern that has been conserved in human cells. We find that the Set1-containing complex (COMPASS, which promotes H3K4me2 and -me3, also serves to limit the abundance of H3K4ac at gene promoters. In addition, we identify a group of genes that have high levels of H3K4ac in their promoters and are inadequately expressed in H3-K4R, but not in set1Δ mutant strains, suggesting that H3K4ac plays a positive role in transcription. Our results reveal a novel regulatory feature of promoter-proximal chromatin, involving mutually exclusive histone modifications of the same histone residue (H3K4ac and H3K4me.

  13. Methamphetamine causes differential alterations in gene expression and patterns of histone acetylation/hypoacetylation in the rat nucleus accumbens.

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    Tracey A Martin

    Full Text Available Methamphetamine (METH addiction is associated with several neuropsychiatric symptoms. Little is known about the effects of METH on gene expression and epigenetic modifications in the rat nucleus accumbens (NAC. Our study investigated the effects of a non-toxic METH injection (20 mg/kg on gene expression, histone acetylation, and the expression of the histone acetyltransferase (HAT, ATF2, and of the histone deacetylases (HDACs, HDAC1 and HDAC2, in that structure. Microarray analyses done at 1, 8, 16 and 24 hrs after the METH injection identified METH-induced changes in the expression of genes previously implicated in the acute and longterm effects of psychostimulants, including immediate early genes and corticotropin-releasing factor (Crf. In contrast, the METH injection caused time-dependent decreases in the expression of other genes including Npas4 and cholecystokinin (Cck. Pathway analyses showed that genes with altered expression participated in behavioral performance, cell-to-cell signaling, and regulation of gene expression. PCR analyses confirmed the changes in the expression of c-fos, fosB, Crf, Cck, and Npas4 transcripts. To determine if the METH injection caused post-translational changes in histone markers, we used western blot analyses and identified METH-mediated decreases in histone H3 acetylated at lysine 9 (H3K9ac and lysine 18 (H3K18ac in nuclear sub-fractions. In contrast, the METH injection caused time-dependent increases in acetylated H4K5 and H4K8. The changes in histone acetylation were accompanied by decreased expression of HDAC1 but increased expression of HDAC2 protein levels. The histone acetyltransferase, ATF2, showed significant METH-induced increased in protein expression. These results suggest that METH-induced alterations in global gene expression seen in rat NAC might be related, in part, to METH-induced changes in histone acetylation secondary to changes in HAT and HDAC expression. The causal role that HATs and

  14. Genome-Wide Profiling of Histone Modifications (H3K9me2 and H4K12ac) and Gene Expression in Rust (Uromyces appendiculatus) Inoculated Common Bean (Phaseolus vulgaris L.)

    National Research Council Canada - National Science Library

    Ayyappan, Vasudevan; Kalavacharla, Venu; Thimmapuram, Jyothi; Bhide, Ketaki P; Sripathi, Venkateswara R; Smolinski, Tomasz G; Manoharan, Muthusamy; Thurston, Yaqoob; Todd, Antonette; Kingham, Bruce

    2015-01-01

    ...) and RNA sequencing (RNA-Seq). ChIP-Seq analysis revealed 1,235 and 556 histone methylation and acetylation responsive genes from common bean leaves treated with the rust pathogen at 0, 12 and 84 hour-after-inoculation (hai...

  15. Genome-Wide Profiling of Histone Modifications (H3K9me2 and H4K12ac) and Gene Expression in Rust (Uromyces appendiculatus) Inoculated Common Bean (Phaseolus vulgaris L.): e0132176

    National Research Council Canada - National Science Library

    Vasudevan Ayyappan; Venu Kalavacharla; Jyothi Thimmapuram; Ketaki P Bhide; Venkateswara R Sripathi; Tomasz G Smolinski; Muthusamy Manoharan; Yaqoob Thurston; Antonette Todd; Bruce Kingham

    2015-01-01

    ...) and RNA sequencing (RNA-Seq). ChIP-Seq analysis revealed 1,235 and 556 histone methylation and acetylation responsive genes from common bean leaves treated with the rust pathogen at 0, 12 and 84 hour-after-inoculation (hai...

  16. Prenatal Exposure to a Maternal High-Fat Diet Affects Histone Modification of Cardiometabolic Genes in Newborn Rats

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    Bijaya Upadhyaya

    2017-04-01

    Full Text Available Infants born to women with diabetes or obesity are exposed to excess circulating fuels during fetal heart development and are at higher risk of cardiac diseases. We have previously shown that late-gestation diabetes, especially in conjunction with a maternal high-fat (HF diet, impairs cardiac functions in rat-offspring. This study investigated changes in genome-wide histone modifications in newborn hearts from rat-pups exposed to maternal diabetes and HF-diet. Chromatin-immunoprecipitation-sequencing revealed a differential peak distribution on gene promoters in exposed pups with respect to acetylation of lysines 9 and 14 and to trimethylation of lysines 4 and 27 in histone H3 (all, false discovery rate, FDR < 0.1. In the HF-diet exposed offspring, 54% of the annotated genes showed the gene-activating mark trimethylated lysine 4. Many of these genes (1 are associated with the “metabolic process” in general and particularly with “positive regulation of cholesterol biosynthesis” (FDR = 0.03; (2 overlap with 455 quantitative trait loci for blood pressure, body weight, serum cholesterol (all, FDR < 0.1; and (3 are linked to cardiac disease susceptibility/progression, based on disease ontology analyses and scientific literature. These results indicate that maternal HF-diet changes the cardiac histone signature in offspring suggesting a fuel-mediated epigenetic reprogramming of cardiac tissue in utero.

  17. Chromatin Modulation of Herpesvirus Lytic Gene Expression: Managing Nucleosome Density and Heterochromatic Histone Modifications

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    Thomas M. Kristie

    2016-03-01

    Full Text Available Like their cellular hosts, herpesviruses are subject to the regulatory impacts of chromatin assembled on their genomes. Upon infection, these viruses are assembled into domains of chromatin with heterochromatic signatures that suppress viral gene expression or euchromatic characteristics that promote gene expression. The organization and modulation of these chromatin domains appear to be intimately linked to the coordinated expression of the different classes of viral genes and thus ultimately play an important role in the progression of productive infection or the establishment and maintenance of viral latency. A recent report from the Knipe laboratory (J. S. Lee, P. Raja, and D. M. Knipe, mBio 7:e02007-15, 2016 contributes to the understanding of the dynamic modulation of chromatin assembled on the herpes simplex virus genome by monitoring the levels of characteristic heterochromatic histone modifications (histone H3 lysine 9 and 27 methylation associated with a model viral early gene during the progression of lytic infection. Additionally, this study builds upon previous observations that the viral immediate-early protein ICP0 plays a role in reducing the levels of heterochromatin associated with the early genes.

  18. Analysis of the subcellular localization of the human histone methyltransferase SETDB1

    Energy Technology Data Exchange (ETDEWEB)

    Tachibana, Keisuke, E-mail: nya@phs.osaka-u.ac.jp [Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Gotoh, Eiko; Kawamata, Natsuko [Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Ishimoto, Kenji [Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Laboratory for System Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Uchihara, Yoshie [Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871 (Japan); Iwanari, Hiroko [Department of Quantitative Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Sugiyama, Akira; Kawamura, Takeshi [Radioisotope Center, The University of Tokyo, 2-11-16 Yayoi, Bunkyo, Tokyo 113-0032 (Japan); Mochizuki, Yasuhiro [Department of Quantitative Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Tanaka, Toshiya [Laboratory for System Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Sakai, Juro [Division of Metabolic Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Hamakubo, Takao [Department of Quantitative Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); Kodama, Tatsuhiko [Laboratory for System Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8904 (Japan); and others

    2015-10-02

    SET domain, bifurcated 1 (SETDB1) is a histone methyltransferase that methylates lysine 9 on histone H3. Although it is important to know the localization of proteins to elucidate their physiological function, little is known of the subcellular localization of human SETDB1. In the present study, to investigate the subcellular localization of hSETDB1, we established a human cell line constitutively expressing enhanced green fluorescent protein fused to hSETDB1. We then generated a monoclonal antibody against the hSETDB1 protein. Expression of both exogenous and endogenous hSETDB1 was observed mainly in the cytoplasm of various human cell lines. Combined treatment with the nuclear export inhibitor leptomycin B and the proteasome inhibitor MG132 led to the accumulation of hSETDB1 in the nucleus. These findings suggest that hSETDB1, localized in the nucleus, might undergo degradation by the proteasome and be exported to the cytosol, resulting in its detection mainly in the cytosol. - Highlights: • Endogenous human SETDB1 was localized mainly in the cytoplasm. • Combined treatment with LMB and MG132 led to accumulation of human SETDB1 in the nucleus. • HeLa cells expressing EFGP-hSETDB1 are useful for subcellular localization analyses.

  19. Genome-wide ChIP-seq mapping and analysis of butyrate-induced H3K9 and H3K27 acetylation and epigenomic landscape alteration in bovine cells

    Science.gov (United States)

    Utilizing next-generation sequencing technology, combined with ChIP (Chromatin Immunoprecipitation) technology, we analyzed histone modification (acetylation) induced by butyrate and the large-scale mapping of the epigenomic landscape of normal histone H3 and acetylated histone H3K9 and H3K27. To d...

  20. Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood.

    Science.gov (United States)

    Kyzar, Evan J; Zhang, Huaibo; Sakharkar, Amul J; Pandey, Subhash C

    2017-09-01

    Alcohol exposure in adolescence is an important risk factor for the development of alcoholism in adulthood. Epigenetic processes are implicated in the persistence of adolescent alcohol exposure-related changes, specifically in the amygdala. We investigated the role of histone methylation mechanisms in the persistent effects of adolescent intermittent ethanol (AIE) exposure in adulthood. Adolescent rats were exposed to 2 g/kg ethanol (2 days on/off) or intermittent n-saline (AIS) during postnatal days (PND) 28-41 and used for behavioral and epigenetic studies. We found that AIE exposure caused a long-lasting decrease in mRNA and protein levels of lysine demethylase 1(Lsd1) and mRNA levels of Lsd1 + 8a (a neuron-specific splice variant) in specific amygdaloid structures compared with AIS-exposed rats when measured at adulthood. Interestingly, AIE increased histone H3 lysine 9 dimethylation (H3K9me2) levels in the central nucleus of the amygdala (CeA) and medial nucleus of the amygdala (MeA) in adulthood without producing any change in H3K4me2 protein levels. Acute ethanol challenge (2 g/kg) in adulthood attenuated anxiety-like behaviors and the decrease in Lsd1 + 8a mRNA levels in the amygdala induced by AIE. AIE caused an increase in H3K9me2 occupancy at the brain-derived neurotrophic factor exon IV promoter in the amygdala that returned to baseline after acute ethanol challenge in adulthood. These results indicate that AIE specifically modulates epizymes involved in H3K9 dimethylation in the amygdala in adulthood, which are possibly responsible for AIE-induced chromatin remodeling and adult psychopathology such as anxiety. © Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  1. Hypoxia and nickel inhibit histone demethylase JMJD1A and repress Spry2 expression in human bronchial epithelial BEAS-2B cells.

    Science.gov (United States)

    Chen, Haobin; Kluz, Thomas; Zhang, Ronghe; Costa, Max

    2010-12-01

    Epigenetic silencing of tumor suppressor genes commonly occurs in human cancers via increasing DNA methylation and repressive histone modifications at gene promoters. However, little is known about how pathogenic environmental factors contribute to cancer development by affecting epigenetic regulatory mechanisms. Previously, we reported that both hypoxia and nickel (an environmental carcinogen) increased global histone H3 lysine 9 methylation in cells through inhibiting a novel class of iron- and α-ketoglutarate-dependent histone demethylases. Here, we investigated whether inhibition of histone demethylase JMJD1A by hypoxia and nickel could lead to repression/silencing of JMJD1A-targeted gene(s). By using Affymetrix GeneChip and ChIP-on-chip technologies, we identified Spry2 gene, a key regulator of receptor tyrosine kinase/extracellular signal-regulated kinase (ERK) signaling, as one of the JMJD1A-targeted genes in human bronchial epithelial BEAS-2B cells. Both hypoxia and nickel exposure increased the level of H3K9me2 at the Spry2 promoter by inhibiting JMJD1A, which probably led to a decreased expression of Spry2 in BEAS-2B cells. Repression of Spry2 potentiated the nickel-induced ERK phosphorylation, and forced expression of Spry2 in BEAS-2B cells decreased the nickel-induced ERK phosphorylation and significantly suppressed nickel-induced anchorage-independent growth. Taken together, our results suggest that histone demethylases could be targets of environmental carcinogens and their inhibition may lead to altered gene expression and eventually carcinogenesis.

  2. Histone methyltransferase Setdb1 is indispensable for Meckel's cartilage development.

    Science.gov (United States)

    Yahiro, Kohei; Higashihori, Norihisa; Moriyama, Keiji

    2017-01-22

    The histone methyltransferase Setdb1 represses gene expression by catalyzing lysine 9 of histone H3 trimethylation. Given that the conventional knockout of Setdb1 is embryo-lethal at the implantation stage, its role in craniofacial development is poorly understood. Here, we investigated the role of Setdb1, using conditional knockout mice-in which Setdb1 was deleted in the Meckel's cartilage (Setdb1 CKO)-and the mouse chondrogenic cell line ATDC5-in which Setdb1 was inhibited by siRNA. Deletion of Setdb1 in Meckel's cartilage, the supportive tissue in the embryonic mandible, led to its enlargement, instead of the degeneration that normally occurs. Chondrocytes from the Meckel's cartilage of Setdb1 CKO mice showed increased size. Furthermore, at embryonic days 16.5 and 18.5, part of the perichondrium was disrupted and mineralization was observed in the Meckel's cartilage. Proliferation analysis showed that inhibition of Setdb1 caused increased proliferation in chondrocytes in the Meckel's cartilage as well as in ATDC5 cells. Quantitative RT-PCR showed decreased expression of chondrogenic genes, such as Sox9, Mmp13, Collagen II, and Aggrecan, as a result of Setdb1 inhibition in ATDC5 cells. Along with these phenomenons, SMAD-dependent BMP signaling was significantly increased by the loss of Setdb1 in both the Meckel's cartilage of Setdb1 CKO mice and ATDC5 cells. Therefore, the abnormal development of Meckel's cartilage in Setdb1 CKO mice is partly due to the enhanced SMAD-dependent BMP signaling. Overall, to our knowledge, the present study is the first to show that epigenetic regulation by Setdb1 is indispensable for the embryonic development of Meckel's cartilage. Copyright © 2016. Published by Elsevier Inc.

  3. Flexibility in crosstalk between H2B ubiquitination and H3 methylation in vivo (Corrigendum)

    NARCIS (Netherlands)

    Vlaming, Hanneke; van Welsem, Tibor; de Graaf, Erik L; Ontoso, David; Altelaar, A F Maarten; San-Segundo, Pedro A; Heck, Albert J R; van Leeuwen, Fred

    2014-01-01

    Histone H2B ubiquitination is a dynamic modification that promotes methylation of histone H3K79 and H3K4. This crosstalk is important for the DNA damage response and has been implicated in cancer. Here, we show that in engineered yeast strains, ubiquitins tethered to every nucleosome promote H3K79

  4. Flexibility in crosstalk between H2B ubiquitination and H3 methylation in vivo

    NARCIS (Netherlands)

    Vlaming, Hanneke; van Welsem, Tibor; de Graaf, Erik L; Ontoso, David; Altelaar, A F Maarten; San-Segundo, Pedro A; Heck, Albert J R; van Leeuwen, Fred

    2014-01-01

    Histone H2B ubiquitination is a dynamic modification that promotes methylation of histone H3K79 and H3K4. This crosstalk is important for the DNA damage response and has been implicated in cancer. Here, we show that in engineered yeast strains, ubiquitins tethered to every nucleosome promote H3K79

  5. Trimethylation of Histone 3 lysine 27 (H3K27me3) ChIP-PCR and transcriptional expression data of Ef1-alpha, cyp26A, HoxC10, HoxD10 and HoxD11 in the Xenopus XTC cell line.

    Science.gov (United States)

    Vieira, Warren; Sahin, Hande; Wells, Kaylee; McCusker, Catherine

    2017-12-01

    Trimethylation of Histone 3 lysine 27 (H3K27me3) is a chromatin modification that is associated with transcriptional repression (Cao et al., 2002; Sarma et al., 2008; Pengelly et al., 2013) [1], [2], [3]. In this article we performed anti-H3K27me3 Chromosomal Immunoprecipitation (ChIP-PCR), to detect the abundance of H3K27me3 marks on Ef1-alpha, cyp26A, HoxC10, HoxD10 and HoxD11 in the Xenopus XTC cell line. We also performed RT-PCR for these genes to determine whether their expression is detectable in the XTC cell culture. The data we present here are the fold enrichment of Ef1-alpha, cyp26A, HoxC10, HoxD10 and HoxD11 on anti-H3K27me3 ChIP compared to no antibody controls. We also present RT-PCR data on the above listed genes.

  6. Downregulation of histone methyltransferase genes SUV39H1 and SUV39H2 increases telomere length in embryonic stem-like cells and embryonic fibroblasts in pigs.

    Science.gov (United States)

    Dang-Nguyen, Thanh Quang; Haraguchi, Seiki; Furusawa, Tadashi; Somfai, Tamas; Kaneda, Masahiro; Watanabe, Shinya; Akagi, Satoshi; Kikuchi, Kazuhiro; Tajima, Atsushi; Nagai, Takashi

    2013-01-01

    Telomere is a nucleoprotein structure at the ends of chromosomes that helps to protect the ends of chromosomes from being fused with other chromosomes. Knockout of histone methyltransferases Suv39h1 and Suv39h2 increases the telomere length in murine cells, whereas downregulation of SUV39H1 and SUV39H2 genes decreases the telomere length in human cells, suggesting that telomere biology is different among mammalian species. However, epigenetic regulation of the telomere has not been studied in mammals other than the human and mouse. In the present study, the effect of knockdown of SUV39H1 and SUV39H2 genes on telomere length was examined in porcine embryonic stem-like cells (pESLCs) and porcine embryonic fibroblasts (PEFs). The telomeres in SUV39H1 and SUV39H2 knockdown (SUV39KD) pESLCs (37.1 ± 0.9 kb) were longer (Ptelomeres (22.1 ± 0.4 kb; Ptelomere elongation in SUV39KD pESLCs and SUV39KD PEFs. Relative levels of trimethylation of histone H3 lysine 9 and expressions of DNMT1, DNMT3A and DNMT3B were decreased in SUV39KD cells, suggesting that telomere lengthening in SUV39KD pESLCs and SUV39KD PEFs might be not only related to the loss of histone modification marks but also linked to the decrease in DNA methyltransferase in pigs.

  7. mRNA levels of related Abcb genes change opposite to each other upon histone deacetylase inhibition in drug-resistant rat hepatoma cells.

    Directory of Open Access Journals (Sweden)

    Adám Sike

    Full Text Available The multidrug-resistant phenotype of tumor cells is acquired via an increased capability of drug efflux by ABC transporters and causes serious problems in cancer treatment. With the aim to uncover whether changes induced by epigenetic mechanisms in the expression level of drug transporter genes correlates with changes in the drug resistance phenotypes of resistant cells, we studied the expression of drug transporters in rat hepatoma cell lines. We found that of the three major rat ABC transporter genes Abcb1a, Abcb1b and Abcc1 the activity of only Abcb1b increased significantly in colchicine-selected, drug-resistant cells. Increased transporter expression in drug-resistant cells results primarily from transcriptional activation. A change in histone modification at the regulatory regions of the chromosomally adjacent Abcb1a and Abcb1b genes differentially affects the levels of corresponding mRNAs. Transcriptional up- and down-regulation accompany an increase in acetylation levels of histone H3 lysine 9 at the promoter regions of Abcb1b and Abcb1a, respectively. Drug efflux activity, however, does not follow tightly the transcriptional activity of drug transporter genes in hepatoma cells. Our results point out the need for careful analysis of cause-and-effect relationships between changes in histone modification, drug transporter expression and drug resistance phenotypes.

  8. Male germ cell apoptosis and epigenetic histone modification induced by Tripterygium wilfordii Hook F.

    Directory of Open Access Journals (Sweden)

    Ji Xiong

    Full Text Available Multiglycosides of Tripterygium wilfordii Hook f (GTW, a Chinese herb-derived medicine used as a remedy for rheumatoid arthritis, are considered to be a reversible anti-fertility drug affecting the mammalian spermatids. However, the mechanism behind this effect is still unknown. To study the possible mechanism behind the impact of GTW on spermatogenesis, we administered 4 groups of 4-week-old male mice with different doses of GTW. We found a dose-dependent decrease in the number of germ cells after 40 days of GTW treatment, and an increase in apoptotic cells from the low-dose to the high-dose group. During this same period the dimethylated level of histone H3 lysine 9 (H3K9me2 in GTW-treated testes germ cells declined. Additionally, spermatogonial stem cells (SSCs from 6-day-old mice were isolated to evaluate the possible effect of GTW or triptolide on development of SSCs. We found a significantly higher incidence of apoptosis and lower dimethylation level of H3K9me2 in the SSCs of GTW or triptolide treatment than in controls. Thus, these data suggest that the GTW-induced apoptosis might be responsible for the fertility impairment in mice. This damage could be traced back to the early stages of spermatogenesis. GTW also affected the epigenetic modification of H3K9 in spermatogenesis. Molecular dynamics simulation suggested that triptolide and dimethylated or trimethylated H3K9 might have similar interaction mechanisms with EED (embryonic ectoderm development. These candidate activation mechanisms provide the first glimpse into the pathway of GTW-induced gonad toxicity, which is crucial for further research and clinical application.

  9. H3K9 and H3K14 acetylation co-occur at many gene regulatory elements, while H3K14ac marks a subset of inactive inducible promoters in mouse embryonic stem cells.

    OpenAIRE

    Karmodiya Krishanpal; Krebs Arnaud R; Oulad-Abdelghani Mustapha; Kimura Hiroshi; Tora Laszlo

    2012-01-01

    Abstract Background Transcription regulation in pluripotent embryonic stem (ES) cells is a complex process that involves multitude of regulatory layers, one of which is post-translational modification of histones. Acetylation of specific lysine residues of histones plays a key role in regulating gene expression. Results Here we have investigated the genome-wide occurrence of two histone marks, acetylation of histone H3K9 and K14 (H3K9ac and H3K14ac), in mouse embryonic stem (mES) cells. Genom...

  10. Effect of cryopreservation and in vitro culture of bovine fibroblasts on histone acetylation levels and in vitro development of hand-made cloned embryos

    Science.gov (United States)

    Chacon, L.; Gomez, M.C.; Jenkins, J.A.; Leibo, S.P.; Wirtu, G.; Dresser, B.L.; Pope, C.E.

    2011-01-01

    In this study, the relative acetylation levels of histone 3 in lysine 9 (H3K9ac) in cultured and cryopreserved bovine fibroblasts was measured and we determined the influence of the epigenetic status of three cultured (C1, C2 and C3) donor cell lines on the in vitro development of reconstructed bovine embryos. Results showed that cryopreservation did not alter the overall acetylation levels of H3K9 in bovine fibroblasts analysed immediately after thawing (frozen/thawed) compared with fibroblasts cultured for a period of time after thawing. However, reduced cleavage rates were noted in embryos reconstructed with fibroblasts used immediately after thawing. Cell passage affects the levels of H3K9ac in bovine fibroblasts, decreasing after P1 and donor cells with lower H3K9ac produced a greater frequency of embryo development to the blastocyst stage. Cryopreservation did not influence the total cell and ICM numbers, or the ICM/TPD ratios of reconstructed embryos. However, the genetic source of donor cells did influence the total number of cells and the trophectoderm cell numbers, and the cell passage influenced the total ICM cell numbers. ?? Copyright Cambridge University Press 2010.

  11. [Histone variants and histone exchange].

    Science.gov (United States)

    Wu, Nan; Gui, Jian-Fang

    2006-04-01

    Histones, as the basic components of nucleosome, are essential to chromatin structure and function. To adapt to various states of chromatin, corresponding histone variants are incorporated in nucleosome, and certain modifications also occur on the variants' tails. These variants change the conformation and stability of nucleosome to facilitate transcriptional activation or deactivation, DNA repairing, heterochromatin formation, and others. During histone exchange, chromatin remodeling complex facilitates histone variant deposition into nucleosome, and different variants have diverse deposition pathways. Recently, research on histone variants is not only a new hotspot in epigenetics, but also a new annotation of "histone code". In addition, histone exchange reveals new changing mechanism of DNA-histone interaction.

  12. 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. PMID:25365782

  13. Interstellar H3+

    Science.gov (United States)

    Oka, Takeshi

    2006-01-01

    Protonated molecular hydrogen, H3+, is the simplest polyatomic molecule. It is the most abundantly produced interstellar molecule, next only to H2, although its steady state concentration is low because of its extremely high chemical reactivity. H3+ is a strong acid (proton donor) and initiates chains of ion-molecule reactions in interstellar space thus leading to formation of complex molecules. Here, I summarize the understandings on this fundamental species in interstellar space obtained from our infrared observations since its discovery in 1996 and discuss the recent observations and analyses of H3+ in the Central Molecular Zone near the Galatic center that led to a revelation of a vast amount of warm and diffuse gas existing in the region. PMID:16894171

  14. Histone demethylase JMJD2B is required for tumor cell proliferation and survival and is overexpressed in gastric cancer

    Energy Technology Data Exchange (ETDEWEB)

    Li, Wenjuan; Zhao, Li; Zang, Wen; Liu, Zhifang; Chen, Long; Liu, Tiantian [Department of Microbiology/Key Laboratory for Experimental Teratology of Chinese Ministry of Education, School of Medicine, Shandong University, 44 Wenhua Xi Road, Jinan 250012 (China); Xu, Dawei, E-mail: Dawei.Xu@ki.se [Department of Microbiology/Key Laboratory for Experimental Teratology of Chinese Ministry of Education, School of Medicine, Shandong University, 44 Wenhua Xi Road, Jinan 250012 (China); Department of Medicine, Division of Hematology, Karolinska University Hospital, Solna and Karolinska Institutet, Stockholm (Sweden); Jia, Jihui, E-mail: jiajihui@sdu.edu.cn [Department of Microbiology/Key Laboratory for Experimental Teratology of Chinese Ministry of Education, School of Medicine, Shandong University, 44 Wenhua Xi Road, Jinan 250012 (China)

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer JMJD2B is required for cell proliferation and in vivo tumorigenesis. Black-Right-Pointing-Pointer JMJD2B depletion induces apoptosis and/or cell cycle arrest. Black-Right-Pointing-Pointer JMJD2B depletion activates DNA damage response and enhances p53 stabilization. Black-Right-Pointing-Pointer JMJD2B is overexpressed in human primary gastric cancer. -- Abstract: Epigenetic alterations such as aberrant expression of histone-modifying enzymes have been implicated in tumorigenesis. Jumonji domain containing 2B (JMJD2B) is a newly identified histone demethylase that regulates chromatin structure or gene expression by removing methyl residues from trimethylated lysine 9 on histone H3. Recent observations have shown oncogenic activity of JMJD2B. We explored the functional role of JMJD2B in cancer cell proliferation, survival and tumorigenesis, and determined its expression profile in gastric cancer. Knocking down JMJD2B expression by small interfering RNA (siRNA) in gastric and other cancer cells inhibited cell proliferation and/or induced apoptosis and elevated the expression of p53 and p21{sup CIP1} proteins. The enhanced p53 expression resulted from activation of the DNA damage response pathway. JMJD2B knockdown markedly suppressed xenograft tumor growth in vivo in mice. Moreover, JMJD2B expression was increased in primary gastric-cancer tissues of humans. Thus, JMJD2B is required for sustained proliferation and survival of tumor cells in vitro and in vivo, and its aberrant expression may contribute to the pathogenesis of gastric cancer.

  15. Histone variants: emerging players in cancer biology

    Science.gov (United States)

    Vardabasso, Chiara; Hasson, Dan; Ratnakumar, Kajan; Chung, Chi-Yeh; Duarte, Luis F.

    2014-01-01

    Histone variants are key players in shaping chromatin structure, and, thus, in regulating fundamental cellular processes such as chromosome segregation and gene expression. Emerging evidence points towards a role for histone variants in contributing to tumor progression, and, recently, the first cancer-associated mutation in a histone variant-encoding gene was reported. In addition, genetic alterations of the histone chaperones that specifically regulate chromatin incorporation of histone variants are rapidly being uncovered in numerous cancers. Collectively, these findings implicate histone variants as potential drivers of cancer initiation and/or progression, and, therefore, targeting histone deposition or the chromatin remodeling machinery may be of therapeutic value. Here, we review the mammalian histone variants of the H2A and H3 families in their respective cellular functions, and their involvement in tumor biology. PMID:23652611

  16. Writers and readers of histone H3 methylation

    NARCIS (Netherlands)

    Warrier, R.A.

    2010-01-01

    The synthesis of RNA from a DNA template in eukaryotic cells is a complex process which is regulated at several stages. This is accomplished by RNA polymerase II and a horde of other transcription factors which together comprise the transcription machinery. Accessing information from DNA in the

  17. Distribution pattern of histone H3 phosphorylation at serine 10 ...

    Indian Academy of Sciences (India)

    Genetics and Plant Breeding, Universidade Federal de Lavras (UFLA), Campus Universitário, Caixa Postal 3037, CEP: 372000-000 Lavras-MG, Brazil; Universidade Federal de Lavras (UFLA), Campus Universitário, Caixa Postal 3037, CEP: 372000-000 Lavras-MG, Brazil; Empresa Brasileira de Pesquisa Agropecuária ...

  18. Arabidopsis ATRX Modulates H3.3 Occupancy and Fine-Tunes Gene Expression

    KAUST Repository

    Duc, Céline

    2017-07-07

    Histones are essential components of the nucleosome, the major chromatin subunit that structures linear DNA molecules and regulates access of other proteins to DNA. Specific histone chaperone complexes control the correct deposition of canonical histones and their variants to modulate nucleosome structure and stability. In this study, we characterize the Arabidopsis Alpha Thalassemia-mental Retardation X-linked (ATRX) ortholog and show that ATRX is involved in histone H3 deposition. Arabidopsis ATRX mutant alleles are viable, but show developmental defects and reduced fertility. Their combination with mutants of the histone H3.3 chaperone HIRA (Histone Regulator A) results in impaired plant survival, suggesting that HIRA and ATRX function in complementary histone deposition pathways. Indeed, ATRX loss of function alters cellular histone H3.3 pools and in consequence modulates the H3.1/H3.3 balance in the cell. H3.3 levels are affected especially at genes characterized by elevated H3.3 occupancy, including the 45S ribosomal DNA (45S rDNA) loci, where loss of ATRX results in altered expression of specific 45S rDNA sequence variants. At the genome-wide scale, our data indicate that ATRX modifies gene expression concomitantly to H3.3 deposition at a set of genes characterized both by elevated H3.3 occupancy and high expression. Altogether, our results show that ATRX is involved in H3.3 deposition and emphasize the role of histone chaperones in adjusting genome expression.

  19. A histone-mimicking interdomain linker in a multidomain protein modulates multivalent histone binding.

    Science.gov (United States)

    Kostrhon, Sebastian; Kontaxis, Georg; Kaufmann, Tanja; Schirghuber, Erika; Kubicek, Stefan; Konrat, Robert; Slade, Dea

    2017-10-27

    N-terminal histone tails are subject to many posttranslational modifications that are recognized by and interact with designated reader domains in histone-binding proteins. BROMO domain adjacent to zinc finger 2B (BAZ2B) is a multidomain histone-binding protein that contains two histone reader modules, a plant homeodomain (PHD) and a bromodomain (BRD), linked by a largely disordered linker. Although previous studies have reported specificity of the PHD domain for the unmodified N terminus of histone H3 and of the BRD domain for H3 acetylated at Lys14 (H3K14ac), the exact mode of H3 binding by BAZ2B and its regulation are underexplored. Here, using isothermal titration calorimetry and NMR spectroscopy, we report that acidic residues in the BAZ2B PHD domain are essential for H3 binding and that BAZ2B PHD-BRD establishes a polyvalent interaction with H3K14ac. Furthermore, we provide evidence that the disordered interdomain linker modulates the histone-binding affinity by interacting with the PHD domain. In particular, lysine-rich stretches in the linker, which resemble the positively charged N terminus of histone H3, reduce the binding affinity of the PHD finger toward the histone substrate. Phosphorylation, acetylation, or poly(ADP-ribosyl)ation of the linker residues may therefore act as a cellular mechanism to transiently tune BAZ2B histone-binding affinity. Our findings further support the concept of interdomain linkers serving a dual role in substrate binding by appropriately positioning the adjacent domains and by electrostatically modulating substrate binding. Moreover, inhibition of histone binding by a histone-mimicking interdomain linker represents another example of regulation of protein-protein interactions by intramolecular mimicry. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. File list: His.Liv.05.H3.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Liv.05.H3.AllCell mm9 Histone H3 Liver SRX027341,SRX100343,SRX100340,SRX100341,...SRX100342 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Liv.05.H3.AllCell.bed ...

  1. H3K23me2 is a new heterochromatic mark in Caenorhabditis elegans

    DEFF Research Database (Denmark)

    Vandamme, Julien; Sidoli, Simone; Mariani, Luca

    2015-01-01

    described in this organism. We used mass spectrometry based middle-down proteomics to analyze histone H3 N-terminal tails from C. elegans embryos for the presence, the relative abundance and the potential cross-talk of co-existing PTMs. This analysis highlighted that the lysine 23 of histone H3 (H3K23......) is extensively modified by methylation and that tri-methylated H3K9 (H3K9me3) is exclusively detected on histone tails with di-methylated H3K23 (H3K23me2). Chromatin immunoprecipitation approaches revealed a positive correlation between H3K23me2 and repressive marks. By immunofluorescence analyses, H3K23me2...... appears differentially regulated in germ and somatic cells, in part by the action of the histone demethylase JMJD-1.2. H3K23me2 is enriched in heterochromatic regions, localizing in H3K9me3 and heterochromatin protein like-1 (HPL-1)-positive foci. Biochemical analyses indicated that HPL-1 binds to H3K23me...

  2. Computational inference of H3K4me3 and H3K27ac domain length

    Directory of Open Access Journals (Sweden)

    Julian Zubek

    2016-03-01

    Full Text Available Background. Recent epigenomic studies have shown that the length of a DNA region covered by an epigenetic mark is not just a byproduct of the assaying technologies and has functional implications for that locus. For example, expanded regions of DNA sequences that are marked by enhancer-specific histone modifications, such as acetylation of histone H3 lysine 27 (H3K27ac domains coincide with cell-specific enhancers, known as super or stretch enhancers. Similarly, promoters of genes critical for cell-specific functions are marked by expanded H3K4me3 domains in the cognate cell type, and these can span DNA regions from 4–5kb up to 40–50kb in length. These expanded H3K4me3 domains are known as buffer domains or super promoters. Methods. To ask what correlates with—and potentially regulates—the length of loci marked with these two important histone marks, H3K4me3 and H3K27ac, we built Random Forest regression models. With these models, we computationally identified genomic and epigenomic patterns that are predictive for the length of these marks in seven ENCODE cell lines. Results. We found that certain epigenetic marks and transcription factors explain the variability of the length of H3K4me3 and H3K27ac marks across different cell types, which implies that the lengths of these two epigenetic marks are tightly regulated in a given cell type. Our source code for the regression models and data can be found at our GitHub page: https://github.com/zubekj/broad_peaks. Discussion. Our Random Forest based regression models enabled us to estimate the individual contribution of different epigenetic marks and protein binding patterns to the length of H3K4me3 and H3K27ac deposition patterns, therefore potentially revealing genomic signatures at cell specific regulatory elements.

  3. Distribution of introns in fungal histone genes.

    Directory of Open Access Journals (Sweden)

    Choong-Soo Yun

    Full Text Available Saccharomycotina and Taphrinomycotina lack intron in their histone genes, except for an intron in one of histone H4 genes of Yarrowia lipolytica. On the other hand, Basidiomycota and Perizomycotina have introns in their histone genes. We compared the distributions of 81, 47, 79, and 98 introns in the fungal histone H2A, H2B, H3, and H4 genes, respectively. Based on the multiple alignments of the amino acid sequences of histones, we identified 19, 13, 31, and 22 intron insertion sites in the histone H2A, H2B, H3, and H4 genes, respectively. Surprisingly only one hot spot of introns in the histone H2A gene is shared between Basidiomycota and Perizomycotina, suggesting that most of introns of Basidiomycota and Perizomycotina were acquired independently. Our findings suggest that the common ancestor of Ascomycota and Basidiomycota maybe had a few introns in the histone genes. In the course of fungal evolution, Saccharomycotina and Taphrinomycotina lost the histone introns; Basidiomycota and Perizomycotina acquired other introns independently. In addition, most of the introns have sequence similarity among introns of phylogenetically close species, strongly suggesting that horizontal intron transfer events between phylogenetically distant species have not occurred recently in the fungal histone genes.

  4. Distribution of introns in fungal histone genes.

    Science.gov (United States)

    Yun, Choong-Soo; Nishida, Hiromi

    2011-01-27

    Saccharomycotina and Taphrinomycotina lack intron in their histone genes, except for an intron in one of histone H4 genes of Yarrowia lipolytica. On the other hand, Basidiomycota and Perizomycotina have introns in their histone genes. We compared the distributions of 81, 47, 79, and 98 introns in the fungal histone H2A, H2B, H3, and H4 genes, respectively. Based on the multiple alignments of the amino acid sequences of histones, we identified 19, 13, 31, and 22 intron insertion sites in the histone H2A, H2B, H3, and H4 genes, respectively. Surprisingly only one hot spot of introns in the histone H2A gene is shared between Basidiomycota and Perizomycotina, suggesting that most of introns of Basidiomycota and Perizomycotina were acquired independently. Our findings suggest that the common ancestor of Ascomycota and Basidiomycota maybe had a few introns in the histone genes. In the course of fungal evolution, Saccharomycotina and Taphrinomycotina lost the histone introns; Basidiomycota and Perizomycotina acquired other introns independently. In addition, most of the introns have sequence similarity among introns of phylogenetically close species, strongly suggesting that horizontal intron transfer events between phylogenetically distant species have not occurred recently in the fungal histone genes.

  5. 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......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...... 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...

  6. Molecular Basis for the Regulation of the H3K4 Methyltransferase Activity of PRDM9

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2013-10-01

    Full Text Available PRDM9, a histone lysine methyltransferase, is a key determinant of the localization of meiotic recombination hot spots in humans and mice and the only vertebrate protein known to be involved in hybrid sterility. Here, we report the crystal structure of the PRDM9 methyltransferase domain in complex with a histone H3 peptide dimethylated on lysine 4 (H3K4me2 and S-adenosylhomocysteine (AdoHcy, which provides insights into the methyltransferase activity of PRDM proteins. We show that the genuine substrate of PRDM9 is histone H3 lysine 4 (H3K4 and that the enzyme possesses mono-, di-, and trimethylation activities. We also determined the crystal structure of PRDM9 in its autoinhibited state, which revealed a rearrangement of the substrate and cofactor binding sites by a concerted action of the pre-SET and post-SET domains, providing important insights into the regulatory mechanisms of histone lysine methyltransferase activity.

  7. Endometriosis is characterized by a distinct pattern of histone 3 and histone 4 lysine modifications.

    Science.gov (United States)

    Monteiro, Janice B; Colón-Díaz, Maricarmen; García, Miosotis; Gutierrez, Sylvia; Colón, Mariano; Seto, Edward; Laboy, Joaquín; Flores, Idhaliz

    2014-03-01

    The histone modification patterns in endometriosis have not been fully characterized. This gap in knowledge results in a poor understanding of the epigenetic mechanisms (and potential therapeutic targets) at play. We aimed to (1) assess global acetylation status of histone 3 (H3) and histone 4 (H4), (2) measure levels of H3 and H4 lysine (K) acetylation and methylation, and (3) to identify histone acetylation patterns in promoter regions of candidate genes in tissues from patients and controls. Global and K-specific acetylation/methylation levels of histones were measured in 24 lesions, 15 endometrium from patients, and 26 endometrium from controls. Chromatin immunoprecipitation (ChIP)-polymerase chain reaction was used to determine the histone acetylation status of the promoter regions of candidate genes in tissues. The lesions were globally hypoacetylated at H3 (but not H4) compared to eutopic endometrium from controls. Lesions had significantly lower levels of H3K9ac and H4K16ac compared to eutopic endometrium from patients and controls. Tissues from patients were hypermethylated at H3K4, H3K9, and H3K27 compared to endometrium from controls. The ChIP analysis showed hypoacetylation of H3/H4 within promoter regions of candidate genes known to be downregulated in endometriosis (e.g., HOXA10, ESR1, CDH1, and p21 (WAF1/Cip1) ) in lesions versus control endometrium. The stereoidogenic factor 1 (SF1) promoter region was enriched for acetylated H3 and H4 in lesions versus control tissues, correlating with its reported high expression in lesions. This study describes the histone code of lesions and endometrium from patients with endometriosis and provides support for a possible role of histone modification in modulation of gene expression in endometriosis.

  8. Inhibition of histone-mediated gene transfer in eucaryotic cells by anti-histone IgG.

    Science.gov (United States)

    Hasselmayer, O; Demirhan, I; Chandra, A; Bayer, M; Müller, R; Chandra, P

    2001-01-01

    In our laboratory, the gene transfer efficiency of some lipofection reagents (lipofectine, lipofectamine, DOTAP and Dosper) and histones H3 and H4 was compared to that of DEAE-Dextran (64). The histones H3 and H4 were found to have the highest transfection efficiency of all the agents tested. In the present study we have analyzed other parameters important for gene delivery by the histones H3 and H4. We transferred the HIV-1 tat gene to Jurkat cells and measured the transactivation of HIV-1-LTR by the transactivator protein, expressed in Jurkat cells. The expression of CAT as a reporter gene hybridized to LTR was a direct measure of transactivation potential. In order to investigate whether the transfection was only due to the positive ionic character of the histones H3 and H4 we tested other histones (H1 and H2A) and polylysine in our system. Under our experimental conditions, neither polylysine, nor the histones H1 and H2A were able to promote gene transfer in Jurkat cells. The inability of these reagents to promote gene transfer was not dependent on DNA condensation; in EMSA (Electrophoretic Mobility Shift Assay) all these reagents exhibited a strong retardation of DNA. In the presence of anti-histone-IgG the transfection potential of histones H3 and H4 was diminished in a concentration - dependent manner. To investigate whether the histone antibodies inhibited the condensation of DNA by histones we carried out gel retardation assays (EMSA) in the absence and in the presence of histone antibodies. Anti-histone-IgG had no effect on the retardation of histone-DNA complexes; on the contrary, retardation was increased. This observation has led us to postulate two models for the possible mechanism by which the histones H3 and H4 catalyze gene transfer in eucaryotic cells.

  9. Optoepigenetics

    DEFF Research Database (Denmark)

    Madsen, Andreas S; Olsen, Christian A

    2016-01-01

    Histone 3 lysine 9 acetylation (H3K9ac) levels were modulated by photoresponsive histone deacetylase (HDAC) inhibitors to perturb the transcription of genes involved in cell cycle regulation and mitochondrial function.......Histone 3 lysine 9 acetylation (H3K9ac) levels were modulated by photoresponsive histone deacetylase (HDAC) inhibitors to perturb the transcription of genes involved in cell cycle regulation and mitochondrial function....

  10. 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

  11. Integrated analysis and transcript abundance modelling of H3K4me3 and H3K27me3 in developing secondary xylem.

    Science.gov (United States)

    Hussey, Steven G; Loots, Mattheus T; van der Merwe, Karen; Mizrachi, Eshchar; Myburg, Alexander A

    2017-06-13

    Despite the considerable contribution of xylem development (xylogenesis) to plant biomass accumulation, its epigenetic regulation is poorly understood. Furthermore, the relative contributions of histone modifications to transcriptional regulation is not well studied in plants. We investigated the biological relevance of H3K4me3 and H3K27me3 in secondary xylem development using ChIP-seq and their association with transcript levels among other histone modifications in woody and herbaceous models. In developing secondary xylem of the woody model Eucalyptus grandis, H3K4me3 and H3K27me3 genomic spans were distinctly associated with xylogenesis-related processes, with (late) lignification pathways enriched for putative bivalent domains, but not early secondary cell wall polysaccharide deposition. H3K27me3-occupied genes, of which 753 (~31%) are novel targets, were enriched for transcriptional regulation and flower development and had significant preferential expression in roots. Linear regression models of the ChIP-seq profiles predicted ~50% of transcript abundance measured with strand-specific RNA-seq, confirmed in a parallel analysis in Arabidopsis where integration of seven additional histone modifications each contributed smaller proportions of unique information to the predictive models. This study uncovers the biological importance of histone modification antagonism and genomic span in xylogenesis and quantifies for the first time the relative correlations of histone modifications with transcript abundance in plants.

  12. 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...... 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...

  13. Top-down and Middle-down Protein Analysis Reveals that Intact and Clipped Human Histones Differ in Post-translational Modification Patterns

    DEFF Research Database (Denmark)

    Tvardovskiy, Andrey; Wrzesinski, Krzysztof; Sidoli, Simone

    2015-01-01

    . Using top-down and middle-down protein analysis by mass spectrometry, we report histone H2B and H3 N-terminal tail clipping in human hepatocytes and demonstrate a relationship between clipping and coexisting PTMs of histone H3. Histones H2B and H3 undergo proteolytic processing in primary human...

  14. EZH2 is a potential therapeutic target for H3K27M-mutant pediatric gliomas

    DEFF Research Database (Denmark)

    Mohammad, Faizaan; Weissmann, Simon; Leblanc, Benjamin Olivier

    2017-01-01

    Diffuse intrinsic pontine glioma (DIPG) is an aggressive brain tumor that is located in the pons and primarily affects children. Nearly 80% of DIPGs harbor mutations in histone H3 genes, wherein lysine 27 is substituted with methionine (H3K27M). H3K27M has been shown to inhibit polycomb repressiv...

  15. Hotspot Mutations in H3F3A and IDH1 Define Distinct Epigenetic and Biological Subgroups of Glioblastoma

    NARCIS (Netherlands)

    Sturm, Dominik; Witt, Hendrik; Hovestadt, Volker; Khuong-Quang, Dong-Anh; Jones, David T. W.; Konermann, Carolin; Pfaff, Elke; Tönjes, Martje; Sill, Martin; Bender, Sebastian; Kool, Marcel; Zapatka, Marc; Becker, Natalia; Zucknick, Manuela; Hielscher, Thomas; Liu, Xiao-Yang; Fontebasso, Adam M.; Ryzhova, Marina; Albrecht, Steffen; Jacob, Karine; Wolter, Marietta; Ebinger, Martin; Schuhmann, Martin U.; van Meter, Timothy; Frühwald, Michael C.; Hauch, Holger; Pekrun, Arnulf; Radlwimmer, Bernhard; Niehues, Tim; von Komorowski, Gregor; Dürken, Matthias; Kulozik, Andreas E.; Madden, Jenny; Donson, Andrew; Foreman, Nicholas K.; Drissi, Rachid; Fouladi, Maryam; Scheurlen, Wolfram; von Deimling, Andreas; Monoranu, Camelia; Roggendorf, Wolfgang; Herold-Mende, Christel; Unterberg, Andreas; Kramm, Christof M.; Felsberg, Jörg; Hartmann, Christian; Wiestler, Benedikt; Wick, Wolfgang; Milde, Till; Witt, Olaf; Lindroth, Anders M.; Schwartzentruber, Jeremy; Faury, Damien; Fleming, Adam; Zakrzewska, Magdalena; Liberski, Pawel P.; Zakrzewski, Krzysztof; Hauser, Peter; Garami, Miklos; Klekner, Almos; Bognar, Laszlo; Morrissy, Sorana; Cavalli, Florence; Taylor, Michael D.; van Sluis, Peter; Koster, Jan; Versteeg, Rogier; Volckmann, Richard; Mikkelsen, Tom; Aldape, Kenneth; Reifenberger, Guido; Collins, V. Peter; Majewski, Jacek; Korshunov, Andrey; Lichter, Peter; Plass, Christoph; Jabado, Nada; Pfister, Stefan M.

    2012-01-01

    Glioblastoma (GBM) is a brain tumor that carries a dismal prognosis and displays considerable heterogeneity. We have recently identified recurrent H3F3A mutations affecting two critical amino acids (K27 and G34) of histone H3.3 in one-third of pediatric GBM. Here, we show that each H3F3A mutation

  16. Inhibition of histone binding by supramolecular hosts

    Science.gov (United States)

    Allen, Hillary F.; Daze, Kevin D.; Shimbo, Takashi; Lai, Anne; Musselman, Catherine A.; Sims, Jennifer K.; Wade, Paul A.; Hof†, Fraser; Kutateladze, Tatiana G.

    2015-01-01

    The tandem PHD (plant homeodomain) fingers of the CHD4 (chromodomain helicase DNA-binding protein 4) ATPase are epigenetic readers that bind either unmodified histone H3 tails or H3K9me3 (histone H3 trimethylated at Lys9). This dual function is necessary for the transcriptional and chromatin remodelling activities of the NuRD (nucleosome remodelling and deacetylase) complex. In the present paper, we show that calixarene-based supramolecular hosts disrupt binding of the CHD4 PHD2 finger to H3K9me3, but do not affect the interaction of this protein with the H3K9me0 (unmodified histone H3) tail. A similar inhibitory effect, observed for the association of chromodomain of HP1γ (heterochromatin protein 1γ) with H3K9me3, points to a general mechanism of methyl-lysine caging by calixarenes and suggests a high potential for these compounds in biochemical applications. Immunofluorescence analysis reveals that the supramolecular agents induce changes in chromatin organization that are consistent with their binding to and disruption of H3K9me3 sites in living cells. The results of the present study suggest that the aromatic macrocyclic hosts can be used as a powerful new tool for characterizing methylation-driven epigenetic mechanisms. PMID:24576085

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

    OpenAIRE

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

    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 ...

  18. 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...

  19. 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

    to track histone modifications and histone variants during DNA replication and across the cell cycle. We show that post-translational modifications (PTMs) are transmitted with parental histones to newly replicated DNA. Di- and trimethylation marks are diluted twofold upon DNA replication, as a consequence...... 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...... of these marks extends over several cell generations. Together, our results reveal how histone marks propagate and demonstrate that chromatin states oscillate within the cell cycle....

  20. Molecular functions of the histone acetyltransferase chaperone complex Rtt109-Vps75

    Energy Technology Data Exchange (ETDEWEB)

    Berndsen, Christopher E; Tsubota, Toshiaki; Lindner, Scott E; Lee, Susan; Holton, James M; Kaufman, Paul D; Keck, James L; Denu, John M [UMASS, MED; (UCB); (UW-MED)

    2010-01-12

    Histone acetylation and nucleosome remodeling regulate DNA damage repair, replication and transcription. Rtt109, a recently discovered histone acetyltransferase (HAT) from Saccharomyces cerevisiae, functions with the histone chaperone Asf1 to acetylate lysine K56 on histone H3 (H3K56), a modification associated with newly synthesized histones. In vitro analysis of Rtt109 revealed that Vps75, a Nap1 family histone chaperone, could also stimulate Rtt109-dependent acetylation of H3K56. However, the molecular function of the Rtt109-Vps75 complex remains elusive. Here we have probed the molecular functions of Vps75 and the Rtt109-Vps75 complex through biochemical, structural and genetic means. We find that Vps75 stimulates the kcat of histone acetylation by {approx}100-fold relative to Rtt109 alone and enhances acetylation of K9 in the H3 histone tail. Consistent with the in vitro evidence, cells lacking Vps75 showed a substantial reduction (60%) in H3K9 acetylation during S phase. X-ray structural, biochemical and genetic analyses of Vps75 indicate a unique, structurally dynamic Nap1-like fold that suggests a potential mechanism of Vps75-dependent activation of Rtt109. Together, these data provide evidence for a multifunctional HAT-chaperone complex that acetylates histone H3 and deposits H3-H4 onto DNA, linking histone modification and nucleosome assembly.

  1. Histone Methylation and Epigenetic Silencing in Breast Cancer

    Science.gov (United States)

    2011-02-01

    methylation of histone H1 or nucleosomal histone H3, Mol. Cell 14 (2004) 183–193. [64] C. Martin , R. Cao, Y. Zhang, Substrate preferences of the EZH2...101] N.A. Lobo , Y. Shimono, D. Qian, M.F. Clarke, The biology of cancer stem cells, Annu. Rev. Cell Dev. Biol. 23 (2007) 675–699. [102] J. Stingl, C

  2. The Fork in the Road: Histone Partitioning During DNA Replication

    Directory of Open Access Journals (Sweden)

    Anthony T. Annunziato

    2015-06-01

    Full Text Available In the following discussion the distribution of histones at the replication fork is examined, with specific attention paid to the question of H3/H4 tetramer "splitting." After a presentation of early experiments surrounding this topic, more recent contributions are detailed. The implications of these findings with respect to the transmission of histone modifications and epigenetic models are also addressed.

  3. Histone propionylation is a mark of active chromatin.

    Science.gov (United States)

    Kebede, Adam F; Nieborak, Anna; Shahidian, Lara Zorro; Le Gras, Stephanie; Richter, Florian; Gómez, Diana Aguilar; Baltissen, Marijke P; Meszaros, Gergo; Magliarelli, Helena de Fatima; Taudt, Aaron; Margueron, Raphael; Colomé-Tatché, Maria; Ricci, Romeo; Daujat, Sylvain; Vermeulen, Michiel; Mittler, Gerhard; Schneider, Robert

    2017-12-01

    Histones are highly covalently modified, but the functions of many of these modifications remain unknown. In particular, it is unclear how histone marks are coupled to cellular metabolism and how this coupling affects chromatin architecture. We identified histone H3 Lys14 (H3K14) as a site of propionylation and butyrylation in vivo and carried out the first systematic characterization of histone propionylation. We found that H3K14pr and H3K14bu are deposited by histone acetyltransferases, are preferentially enriched at promoters of active genes and are recognized by acylation-state-specific reader proteins. In agreement with these findings, propionyl-CoA was able to stimulate transcription in an in vitro transcription system. Notably, genome-wide H3 acylation profiles were redefined following changes to the metabolic state, and deletion of the metabolic enzyme propionyl-CoA carboxylase altered global histone propionylation levels. We propose that histone propionylation, acetylation and butyrylation may act in combination to promote high transcriptional output and to couple cellular metabolism with chromatin structure and function.

  4. Roles of H3K27me2 and H3K27me3 Examined during Fate Specification of Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Aster H. Juan

    2016-10-01

    Full Text Available The polycomb repressive complex 2 (PRC2 methylates lysine 27 of histone H3 (H3K27 through its catalytic subunit Ezh2. PRC2-mediated di- and tri-methylation (H3K27me2/H3K27me3 have been interchangeably associated with gene repression. However, it remains unclear whether these two degrees of H3K27 methylation have different functions. In this study, we have generated isogenic mouse embryonic stem cells (ESCs with a modified H3K27me2/H3K27me3 ratio. Our findings document dynamic developmental control in the genomic distribution of H3K27me2 and H3K27me3 at regulatory regions in ESCs. They also reveal that modifying the ratio of H3K27me2 and H3K27me3 is sufficient for the acquisition and repression of defined cell lineage transcriptional programs and phenotypes and influences induction of the ESC ground state.

  5. DNA replication origin function is promoted by H3K4 di-methylation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Rizzardi, Lindsay F; Dorn, Elizabeth S; Strahl, Brian D; Cook, Jeanette Gowen

    2012-10-01

    DNA replication is a highly regulated process that is initiated from replication origins, but the elements of chromatin structure that contribute to origin activity have not been fully elucidated. To identify histone post-translational modifications important for DNA replication, we initiated a genetic screen to identify interactions between genes encoding chromatin-modifying enzymes and those encoding proteins required for origin function in the budding yeast Saccharomyces cerevisiae. We found that enzymes required for histone H3K4 methylation, both the histone methyltransferase Set1 and the E3 ubiquitin ligase Bre1, are required for robust growth of several hypomorphic replication mutants, including cdc6-1. Consistent with a role for these enzymes in DNA replication, we found that both Set1 and Bre1 are required for efficient minichromosome maintenance. These phenotypes are recapitulated in yeast strains bearing mutations in the histone substrates (H3K4 and H2BK123). Set1 functions as part of the COMPASS complex to mono-, di-, and tri-methylate H3K4. By analyzing strains lacking specific COMPASS complex members or containing H2B mutations that differentially affect H3K4 methylation states, we determined that these replication defects were due to loss of H3K4 di-methylation. Furthermore, histone H3K4 di-methylation is enriched at chromosomal origins. These data suggest that H3K4 di-methylation is necessary and sufficient for normal origin function. We propose that histone H3K4 di-methylation functions in concert with other histone post-translational modifications to support robust genome duplication.

  6. Structural insight into the recognition of the H3K4me3 mark by the TFIID subunit TAF3

    NARCIS (Netherlands)

    van Ingen, H.|info:eu-repo/dai/nl/297054651; van Schaik, F.M.A.; Wienk, H.|info:eu-repo/dai/nl/203884884; Ballering, J.|info:eu-repo/dai/nl/325784876; Rehmann, H.; Dechesne, A.C.|info:eu-repo/dai/nl/304841684; Kruijzer, J.A.W.; Liskamp, R.M.J.|info:eu-repo/dai/nl/069091315; Timmers, H.T.M.; Boelens, R.|info:eu-repo/dai/nl/070151407

    2008-01-01

    Trimethylation of lysine residue K4 of histone H3 (H3K4me3) strongly correlates with active promoters for RNA polymerase II-transcribed genes. Several reader proteins, including the basal transcription factor TFIID, for this nucleosomal mark have been identified. Its TAF3 subunit specifically binds

  7. Drosophila KDM2 is a H3K4me3 demethylase regulating nucleolar organization

    Directory of Open Access Journals (Sweden)

    Birchler James A

    2009-10-01

    Full Text Available Abstract Background CG11033 (dKDM2 is the Drosophila homolog of the gene KDM2B. dKDM2 has been known to possess histone lysine demethylase activity towards H3K36me2 in cell lines and it regulates H2A ubiquitination. The human homolog of the gene has dual activity towards H3K36me2 as well as H3K4me3, and plays an important role in cellular senescence. Findings We have used transgenic flies bearing an RNAi construct for the dKDM2 gene. The knockdown of dKDM2 gene was performed by crossing UAS-RNAi-dKDM2 flies with actin-Gal4 flies. Western blots of acid extracted histones and immunofluoresence analysis of polytene chromosome showed the activity of the enzyme dKDM2 to be specific for H3K4me3 in adult flies. Immunofluoresence analysis of polytene chromosome also revealed the presence of multiple nucleoli in RNAi knockdown mutants of dKDM2 and decreased H3-acetylation marks associated with active transcription. Conclusion Our findings indicate that dKDM2 is a histone lysine demethylase with specificity for H3K4me3 and regulates nucleolar organization.

  8. Resolving Heart Regeneration by Replacement Histone Profiling.

    Science.gov (United States)

    Goldman, Joseph Aaron; Kuzu, Guray; Lee, Nutishia; Karasik, Jaclyn; Gemberling, Matthew; Foglia, Matthew J; Karra, Ravi; Dickson, Amy L; Sun, Fei; Tolstorukov, Michael Y; Poss, Kenneth D

    2017-02-27

    Chromatin regulation is a principal mechanism governing animal development, yet it is unclear to what extent structural changes in chromatin underlie tissue regeneration. Non-mammalian vertebrates such as zebrafish activate cardiomyocyte (CM) division after tissue damage to regenerate lost heart muscle. Here, we generated transgenic zebrafish expressing a biotinylatable H3.3 histone variant in CMs and derived cell-type-specific profiles of histone replacement. We identified an emerging program of putative enhancers that revise H3.3 occupancy during regeneration, overlaid upon a genome-wide reduction of H3.3 from promoters. In transgenic reporter lines, H3.3-enriched elements directed gene expression in subpopulations of CMs. Other elements increased H3.3 enrichment and displayed enhancer activity in settings of injury- and/or Neuregulin1-elicited CM proliferation. Dozens of consensus sequence motifs containing predicted transcription factor binding sites were enriched in genomic regions with regeneration-responsive H3.3 occupancy. Thus, cell-type-specific regulatory programs of tissue regeneration can be revealed by genome-wide H3.3 profiling. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Structural biology of human H3K9 methyltransferases.

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2010-01-01

    Full Text Available SET domain methyltransferases deposit methyl marks on specific histone tail lysine residues and play a major role in epigenetic regulation of gene transcription. We solved the structures of the catalytic domains of GLP, G9a, Suv39H2 and PRDM2, four of the eight known human H3K9 methyltransferases in their apo conformation or in complex with the methyl donating cofactor, and peptide substrates. We analyzed the structural determinants for methylation state specificity, and designed a G9a mutant able to tri-methylate H3K9. We show that the I-SET domain acts as a rigid docking platform, while induced-fit of the Post-SET domain is necessary to achieve a catalytically competent conformation. We also propose a model where long-range electrostatics bring enzyme and histone substrate together, while the presence of an arginine upstream of the target lysine is critical for binding and specificity.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 plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

  10. Characterization of Chromatin Structure-associated Histone Modifications in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Chang Pyo Hong

    2012-09-01

    Full Text Available Chromatin structure and dynamics that are influenced by epigenetic marks, such as histone modification and DNA methylation, play a crucial role in modulating gene transcription. To understand the relationship between histone modifications and regulatory elements in breast cancer cells, we compared our chromatin immunoprecipitation sequencing (ChIP-Seq histone modification patterns for histone H3K4me1, H3K4me3, H3K9/16ac, and H3K27me3 in MCF-7 cells with publicly available formaldehyde-assisted isolation of regulatory elements (FAIRE-chip signals in human chromosomes 8, 11, and 12, identified by a method called FAIRE. Active regulatory elements defined by FAIRE were highly associated with active histone modifications, like H3K4me3 and H3K9/16ac, especially near transcription start sites. The H3K9/16ac-enriched genes that overlapped with FAIRE signals (FAIRE-H3K9/14ac were moderately correlated with gene expression levels. We also identified functional sequence motifs at H3K4me1-enriched FAIRE sites upstream of putative promoters, suggesting that regulatory elements could be associated with H3K4me1 to be regarded as distal regulatory elements. Our results might provide an insight into epigenetic regulatory mechanisms explaining the association of histone modifications with open chromatin structure in breast cancer cells.

  11. Cigarette smoke induces distinct chromatin histone modifications in lung cells: implication in pathogenesis of COPD and lung cancer

    Science.gov (United States)

    Sundar, Isaac K.; Nevid, Michael Z.; Friedman, Alan E.; Rahman, Irfan

    2014-01-01

    Cigarette smoke (CS)-mediated oxidative stress induces several signaling cascades, including kinases, which results in chromatin modifications (histone acetylation/deacetylation and histone methylation/demethylation). We have previously reported that CS induces chromatin remodeling in pro-inflammatory gene promoters; however, the underlying site-specific histone marks formed in histones H3 and H4 during CS exposure in lungs in vivo and in lung cells in vitro, which can either drive gene expression or repression are not known. We hypothesize that CS exposure in mouse and human bronchial epithelial cells (H292) can cause site-specific posttranslational histone modifications (PTMs) that may play an important role in the pathogenesis of CS-induced chronic lung diseases. We used a bottom-up mass spectrometry approach to identify some potentially novel histone marks, including acetylation, mono-methylation and di-methylation in specific lysine and arginine residues of histones H3 and H4 in mouse lungs and H292 cells. We found that CS-induced distinct posttranslational histone modification patterns in histone H3 and histone H4 in lung cells, which may be considered as usable biomarkers for CS-induced chronic lung diseases. These identified histone marks (histone H3 and histone H4) may play an important role in epigenetic state during the pathogenesis of smoking-induced chronic lung diseases, such as chronic obstructive pulmonary disease and lung cancer. PMID:24283195

  12. 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.

  13. Detection of histone modifications in plant leaves.

    Science.gov (United States)

    Jaskiewicz, Michal; Peterhansel, Christoph; Conrath, Uwe

    2011-09-23

    Chromatin structure is important for the regulation of gene expression in eukaryotes. In this process, chromatin remodeling, DNA methylation, and covalent modifications on the amino-terminal tails of histones H3 and H4 play essential roles(1-2). H3 and H4 histone modifications include methylation of lysine and arginine, acetylation of lysine, and phosphorylation of serine residues(1-2). These modifications are associated either with gene activation, repression, or a primed state of gene that supports more rapid and robust activation of expression after perception of appropriate signals (microbe-associated molecular patterns, light, hormones, etc.)(3-7). Here, we present a method for the reliable and sensitive detection of specific chromatin modifications on selected plant genes. The technique is based on the crosslinking of (modified) histones and DNA with formaldehyde(8,9), extraction and sonication of chromatin, chromatin immunoprecipitation (ChIP) with modification-specific antibodies(9,10), de-crosslinking of histone-DNA complexes, and gene-specific real-time quantitative PCR. The approach has proven useful for detecting specific histone modifications associated with C(4;) photosynthesis in maize(5,11) and systemic immunity in Arabidopsis(3).

  14. File list: His.Brs.50.H3K27ac3.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Brs.50.H3K27ac3.AllCell hg19 Histone H3K27ac3 Breast SRX685192,SRX1115312,SRX68...5193,SRX685196,SRX685195,SRX685194,SRX685191 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Brs.50.H3K27ac3.AllCell.bed ...

  15. File list: His.ALL.05.H3K27ac3.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.ALL.05.H3K27ac3.AllCell hg19 Histone H3K27ac3 All cell types SRX1115312,SRX1424...875,SRX685195,SRX685192,SRX685196,SRX685194,SRX685193,SRX685191 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.ALL.05.H3K27ac3.AllCell.bed ...

  16. File list: His.Brs.20.H3K27ac3.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Brs.20.H3K27ac3.AllCell hg19 Histo