Showing papers on "Histone demethylation published in 2010"

The histone demethylase UTX enables RB-dependent cell fate control

Abstract: Trimethylation of histone H3 on Lys 27 (H3K27me3) is key for cell fate regulation. The H3K27me3 demethylase UTX functions in development and tumor suppression with undefined mechanisms. Here, genome-wide chromatin occupancy analysis of UTX and associated histone modifications reveals distinct classes of UTX target genes, including genes encoding Retinoblastoma (RB)-binding proteins. UTX removes H3K27me3 and maintains expression of several RB-binding proteins, enabling cell cycle arrest. Genetic interactions in mammalian cells and Caenorhabditis elegans show that UTX regulates cell fates via RB-dependent pathways. Thus, UTX defines an evolutionarily conserved mechanism to enable coordinate transcription of a RB network in cell fate control.

Essential functions of the histone demethylase lid.

Abstract: Drosophila Little imaginal discs (Lid) is a recently described member of the JmjC domain class of histone demethylases that specifically targets trimethylated histone H3 lysine 4 (H3K4me3). To understand its biological function, we have utilized a series of Lid deletions and point mutations to assess the role that each domain plays in histone demethylation, in animal viability, and in cell growth mediated by the transcription factor dMyc. Strikingly, we find that lid mutants are rescued to adulthood by either wildtype or enzymatically inactive Lid expressed under the control of its endogenous promoter, demonstrating that Lid's demethylase activity is not essential for development. In contrast, ubiquitous expression of UAS-Lid transgenes lacking its JmjN, C-terminal PHD domain, and C5HC2 zinc finger were unable to rescue lid homozygous mutants, indicating that these domains carry out Lid's essential developmental functions. Although Lid-dependent demethylase activity is not essential, dynamic removal of H3K4me3 may still be an important component of development, as we have observed a genetic interaction between lid and another H3K4me3 demethylase, dKDM2. We also show that Lid's essential C-terminal PHD finger binds specifically to di- and trimethylated H3K4 and that this activity is required for Lid to function in dMyc-induced cell growth. Taken together, our findings highlight the importance of Lid function in the regulated removal and recognition of H3K4me3 during development.

Hypoxia induces trimethylated H3 lysine 4 by inhibition of JARID1A demethylase.

Abstract: Histone H3 Lysine 4 (H3K4) tri-methylation (H3K4me3) at the promoter region of genes has been linked to transcriptional activation In the present study, we found that hypoxia (1 % oxygen) increased H3K4me3 in both normal human bronchial epithelial Beas-2B cells and human lung carcinoma A549 cells The increase of H3K4me3 by hypoxia was likely caused by the inhibition of H3K4 demethylating activity, as hypoxia still increased H3K4me3 in methionine-deficient medium Furthermore, an in vitro histone demethylation assay demonstrated that 1% oxygen decreased the activity of H3K4 demethylases in Beas-2B nuclear extract since ambient oxygen tensions were required for the demethylation reaction to proceed Hypoxia only minimally increased H3K4me3 in the BEAS-2B cells with knockdown of JARID1A, which is the major histone H3K4 demethylase in this cell line However, the mRNA and protein levels of JARID1A were not affected by hypoxia GeneChip and pathway analysis in JARID1A knockdown Beas-2B cells revealed that JARID1A regulates the expression of hundreds of genes involved in different cellular functions, including tumorigenesis Knocking down of JARID1A increased H3K4me3 at the promoters of HMOX1 and DAF genes Thus, these results indicate that hypoxia may target JARID1A activity which in turn increases H3K4me3 at both the global and gene specific levels, leading to the altered programs of gene expression and tumor progression

A plant-specific histone H3 lysine 4 demethylase represses the floral transition in Arabidopsis.

Abstract: Histone demethylation regulates chromatin structure and gene expression, and is catalyzed by various histone demethylases. Trimethylation of histone H3 at lysine 4 (H3K4) is coupled to active gene expression; trimethyl H3K4 is demethylated by Jumonj C (JmjC) domain-containing demethylases in mammals. Here we report that a plant-specific JmjC domain-containing protein known as PKDM7B (At4g20400) demethylates trimethyl H3K4. PKDM7B mediates H3K4 demethylation in a key floral promoter, FLOWERING LOCUS T (FT), and an FT homolog, TWIN SISTER OF FT (TSF), and represses their expression to inhibit the floral transition in Arabidopsis. Our findings suggest that there are at least two distinct sub-families of JmjC domain-containing demethylases that demethylate the active trimethyl H3K4 mark in eukaryotic genes, and reveal a plant-specific JmjC domain enzyme capable of H3K4 demethylation.

Analysis of Jmjd6 Cellular Localization and Testing for Its Involvement in Histone Demethylation

Abstract: Methylation of residues in histone tails is part of a network that regulates gene expression. JmjC domain containing proteins catalyze the oxidative removal of methyl groups on histone lysine residues. Here, we report studies to test the involvement of Jumonji domain-containing protein 6 (Jmjd6) in histone lysine demethylation. Jmjd6 has recently been shown to hydroxylate RNA splicing factors and is known to be essential for the differentiation of multiple tissues and cells during embryogenesis. However, there have been conflicting reports as to whether Jmjd6 is a histone-modifying enzyme. Immunolocalization studies reveal that Jmjd6 is distributed throughout the nucleoplasm outside of regions containing heterochromatic DNA, with occasional localization in nucleoli. During mitosis, Jmjd6 is excluded from the nucleus and reappears in the telophase of the cell cycle. Western blot analyses confirmed that Jmjd6 forms homo-multimers of different molecular weights in the nucleus and cytoplasm. A comparison of mono-, di-, and tri-methylation states of H3K4, H3K9, H3K27, H3K36, and H4K20 histone residues in wildtype and Jmjd6-knockout cells indicate that Jmjd6 is not involved in the demethylation of these histone lysine residues. This is further supported by overexpression of enzymatically active and inactive forms of Jmjd6 and subsequent analysis of histone methylation patterns by immunocytochemistry and western blot analysis. Finally, treatment of cells with RNase A and DNase I indicate that Jmjd6 may preferentially associate with RNA/RNA complexes and less likely with chromatin. Taken together, our results provide further evidence that Jmjd6 is unlikely to be involved in histone lysine demethylation. We confirmed that Jmjd6 forms multimers and showed that nuclear localization of the protein involves association with a nucleic acid matrix.

Histone demethylase JHDM2A is involved in male infertility and obesity.

Abstract: Recent studies indicate that histone lysine methylation is subject to enzyme-catalyzed reversion, and jumonji C (JmjC) domain-containing proteins have been identified as one of the members of histone demethylases. Although an increasing number of histone demethylases have been identified and biochemically characterized, their biological functions are poorly characterized. To elucidate the physiological functions, we generated the knockout mouse model of dimethylated or monomethylated histone 3 lysine 9 (H3K9me2/1)-specific JmjC domain-containing histone demethylase 2A (JHDM2A; also known as JMJD1A and KDM3A) and showed that JHDM2A is essential for spermatogenesis. Jhdm2a-deficient mice exhibited impaired postmeiotic chromatin condensation, which caused infertility, even though the hormonal levels were maintained. Further molecular and biochemical analysis revealed that JHDM2A directly bound to the core promoter regions of transition nuclear protein 1 (Tnp1) and protamine 1 (Prm1) genes, and it induced the transcriptional activation of these genes by removing H3K9 methylation, which is known as a silencing marker of gene transcription. This work uncovered a role for JHDM2A in spermatogenesis and identified 2 downstream genes that are critical for sperm nuclear condensation. In addition, we also showed that JHDM2A plays a role in regulating fat metabolic gene expression in muscle and brown fat tissue, and the knockout mice exhibited obesity and hyperlipidemia. Thus, JHDM2A possesses organ/tissue-specific target genes, and impairment of this molecule cannot be compensated by other JmjC-containing histone demethylases, suggesting the importance of this molecule in vivo.

Small-Molecule-Based Inhibition of Histone Demethylation in Cells Assessed by Quantitative Mass Spectrometry

Abstract: Post-translational modifications on histones are an important mechanism for the regulation of gene expression and are involved in all aspects of cell growth and differentiation, as well as pathological processes including neurodegeneration, autoimmunity, and cancer. A major challenge within the chromatin field is to develop methods for the quantitative analysis of histone modifications. Here we report a mass spectrometry (MS) approach based on ultraperformance liquid chromatography high/low collision switching (UPLC-MSE) to monitor histone modifications in cells. This approach is exemplified by the analysis of trimethylated lysine-9 levels in histone H3, following a simple chemical derivatization procedure with d6-acetic anhydride. This method was used to study the inhibition of histone demethylases with pyridine-2,4-dicarboxylic acid (PDCA) derivatives in cells. Our results show that the PDCA-dimethyl ester inhibits JMJD2A catalyzed demethylation of lysine-9 on histone H3 in human HEK 293T cells. Demethy...

Role of H3K4 demethylases in complex neurodevelopmental diseases

Abstract: Significant neurological disorders can result from subtle perturbations of gene regulation that are often linked to epigenetic regulation. Proteins that regulate the methylation of lysine 4 of histone H3 (H3K4) and play a central role in epigenetic regulation, and mutations in genes encoding these enzymes have been identified in both autism and Rett syndrome. The H3K4 demethylases remove methyl groups from lysine 4 leading to loss of RNA polymerase binding and transcriptional repression. When these proteins are mutated, brain development is altered. Currently, little is known regarding how these gene regulators function at the genomic level. In this article, we will discuss findings that link H3K4 demethylases to neurodevelopment and neurological disease.

Regulation of adipogenesis by nuclear receptor PPARγ is modulated by the histone demethylase JMJD2C.

Abstract: A potential strategy to combat obesity and its associated complications involves modifying gene expression in adipose cells to reduce lipid accumulation. The nuclear receptor Peroxisome Proliferator-activated receptor gamma (PPARγ) is the master regulator of adipose cell differentiation and its functional activation is currently used as a therapeutic approach for Diabetes Mellitus type 2. However, total activation of PPARγ induces undesirable secondary effects that might be set with a partial activation. A group of proteins that produce histone demethylation has been shown to modify the transcriptional activity of nuclear receptors. Here we describe the repressive action of the jumonji domain containing 2C/lysine demethylase 4 C (JMJD2C/KDM4C) on PPARγ transcriptional activation. JMJD2C significantly reduced the rosiglitazone stimulated PPARγ activation. This effect was mainly observed in experiments performed using the Tudor domains that may interact with histone deacetylase class 1 (HDAC) and this interaction probably reduces the mediated activation of PPARγ. Trichostatin A, a HDAC inhibitor, reduces the repressive effect of JMJD2C. When JMJD2C was over-expressed in 3T3-L1 cells, a reduction of differentiation was observed with the Tudor domain. In summary, we herein describe JMJD2C-mediated reduction of PPARgamma transcriptional activation as well as preadipocyte differentiation. This novel action of JMJD2C might have an important role in new therapeutic approaches to treat obesity and its complications.

Regulation of alternative macrophage activation by chromatin remodeling

Abstract: The regulation of gene expression through changes in chromatin structure is increasingly recognized as a chief component of activation of cells of the immune response. It now seems that histone demethylation of the promoter of the gene encoding the transcription factor IRF4 contributes to alternative macrophage activation.

Analysis of hairless corepressor mutants to characterize molecular cooperation with the vitamin D receptor in promoting the mammalian hair cycle.

Abstract: The mammalian hair cycle requires both the vitamin D receptor (VDR) and the hairless (Hr) corepressor, each of which is expressed in the hair follicle. Hr interacts directly with VDR to repress VDR-targeted transcription. Herein, we further map the VDR-interaction domain to regions in the C-terminal half of Hr that contain two LXXLL-like pairs of motifs known to mediate contact of Hr with the RAR-related orphan receptor alpha and with the thyroid hormone receptor, respectively. Site-directed mutagenesis indicates that all four hydrophobic motifs are required for VDR transrepression by Hr. Point mutation of rat Hr at conserved residues corresponding to natural mutants causing alopecia in mice (G985W and a C-terminal deletion ΔAK) and in humans (P95S, C422Y, E611G, R640Q, C642G, N988S, D1030N, A1040T, V1074M and V1154D), as well as alteration of residues in the C-terminal Jumonji C domain implicated in histone demethylation activity (C1025G/E1027G and H1143G) revealed that all Hr mutants retained VDR association, and that transrepressor activity was selectively abrogated in C642G, G985W, N988S, D1030N, V1074M, H1143G and V1154D. Four of these latter Hr mutants (C642G, N988S, D1030N and V1154D) were found to associate normally with histone deacetylase-3. Finally, we identified three regions of human VDR necessary for association with Hr, namely residues 109–111, 134–201, and 202–303. It is concluded that Hr and VDR interact via multiple protein-protein interfaces, with Hr recruiting histone deacetylases and possibly itself catalyzing histone demethylation to effect chromatin remodeling and repress the transcription of VDR target genes that control the hair cycle.

The JmjC domain of Gis1 is dispensable for transcriptional activation

Abstract: Yeast Gis1 protein functions as a transcription factor after nutrient limitation and oxidative stress. In this report, we show that Gis1 also regulates the induction of several genes involved in spore wall synthesis during sporulation. Gis1 contains a JmjC domain near its N-terminus. In many proteins, JmjC domains provide histone demethylase activity. Whether the JmjC domain of Gis1 contributes to its transcriptional activation is still unknown. Here, we show that gis1 point mutations that abolish Fe (II) and α-ketoglutarate binding, known cofactors in other JmjC proteins, are still able to induce transcription normally during glucose starvation and sporulation. Even the deletion of the entire JmjC domain does not affect transcriptional activation by Gis1. Moreover, the JmjC domain is not required for the toxicity associated with Gis1 overexpression. The data demonstrate that the JmjC domain is dispensable for transcriptional activation by Gis1 during nutrient stress and sporulation.

Method of treatment

Abstract: The present invention relates to methods for treatment or prevention of autoimmune and inflammatory diseases and conditions by inhibiting or modifying histone demethylation In a further aspect the invention relates to a method for identifying agents useful in said methods of treatment The invention particularly describes the role of certain histone demethylase enzymes in these diseases and conditions and their use as therapeutic and screening targets