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Results for “Histone Demethylases”

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At least 19 recordsLinked to original sources

PubMed2006

Reversal of histone lysine trimethylation by the JMJD2 family of histone demethylases.

Histone methylation regulates chromatin structure, transcription, and epigenetic state of the cell. Histone methylation is dynamically regulated by histone methylases and demethylases such as LSD1 and JHDM1, which mediate demethylation of di- and monomethylated histones. It has been unclear whether demethylases exist that reverse lysine trimethylation. We show the JmjC domain-containing protein JMJD2A reversed trimethylated H3-K9/K36 to di- but not mono- or unmethylated products. Overexpression of JMJD2A but not a catalytically inactive mutant reduced H3-K9/K36 trimethylation levels in cultured cells. In contrast, RNAi depletion of the C. elegans JMJD2A homolog resulted in an increase in general H3-K9Me3 and localized H3-K36Me3 levels on meiotic chromosomes and triggered p53-dependent germline apoptosis. Additionally, other human JMJD2 subfamily members also functioned as trimethylation-specific demethylases, converting H3-K9Me3 to H3-K9Me2 and H3-K9Me1, respectively. Our finding that this family of demethylases generates different methylated states at the same lysine residue provides a mechanism for fine-tuning histone methylation.

Animals↗

PubMed2006

Structural basis for CoREST-dependent demethylation of nucleosomes by the human LSD1 histone demethylase.

Histone methylation regulates diverse chromatin-templated processes, including transcription. Many transcriptional corepressor complexes contain lysine-specific demethylase 1 (LSD1) and CoREST that collaborate to demethylate mono- and dimethylated H3-K4 of nucleosomes. Here, we report the crystal structure of the LSD1-CoREST complex. LSD1-CoREST forms an elongated structure with a long stalk connecting the catalytic domain of LSD1 and the CoREST SANT2 domain. LSD1 recognizes a large segment of the H3 tail through a deep, negatively charged pocket at the active site and possibly a shallow groove on its surface. CoREST SANT2 interacts with DNA. Disruption of the SANT2-DNA interaction diminishes CoREST-dependent demethylation of nucleosomes by LSD1. The shape and dimension of LSD1-CoREST suggest its bivalent binding to nucleosomes, allowing efficient H3-K4 demethylation. This spatially separated, multivalent nucleosome binding mode may apply to other chromatin-modifying enzymes that generally contain multiple nucleosome binding modules.

Binding Sites↗

PubMed2006

A mechanism-based inactivator for histone demethylase LSD1.

Histone demethylase LSD1 is a flavin-dependent amine oxidase that catalyzes the oxidative removal of one or two methyl groups from the methyl-lysine-4 side chain of histone H3. We have designed and synthesized two peptide-based inhibitor analogues that block LSD1. One of these inhibitors, compound 1, contains a propargylamine functionality and shows time-dependent inactivation of LSD1. Peptide substrate, diMeK4H3-21, protected LSD1 against inactivation by 1 in a concentration-dependent fashion. Mass spectrometric analysis showed that 1 forms a covalent interaction with FAD. Compound 1 did not detectably inhibit monoamine oxidase B in the concentration range studied. Compound 1 is thus a selective, mechanism-based inactivator of LSD1 and is likely to serve as a useful tool in the study of histone modifications and chromatin remodeling.

Amino Acid Sequence↗

PubMed2005

Human histone demethylase LSD1 reads the histone code.

Human histone demethylase LSD1 is a flavin-dependent amine oxidase that catalyzes the specific removal of methyl groups from mono- and dimethylated Lys4 of histone H3. The N-terminal tail of H3 is subject to various covalent modifications, and a fundamental question in LSD1 biology is how these epigenetic marks affect the demethylase activity. We show that LSD1 does not have a strong preference for mono- or dimethylated Lys4 of H3. Substrate recognition is not confined to the residues neighboring Lys4, but it requires a sufficiently long peptide segment consisting of the N-terminal 20 amino acids of H3. Electrostatic interactions are an important factor in protein-substrate recognition, as indicated by the high sensitivity of Km to ionic strength. We have probed LSD1 for its ability to demethylate Lys4 in presence of a second modification on the same peptide substrate. Methylation of Lys9 does not affect enzyme catalysis. Conversely, Lys9 acetylation causes an almost 6-fold increase in the Km value, whereas phosphorylation of Ser10 totally abolishes activity. LSD1 is inhibited by a demethylated peptide with an inhibition constant of 1.8 microM, suggesting that LSD1 can bind to H3 independently of Lys4 methylation. LSD1 is a chromatin-modifying enzyme, which is able to read different epigenetic marks on the histone N-terminal tail and can serve as a docking module for the stabilization of the associated corepressor complex(es) on chromatin.

Amino Acid Sequence↗

PubMed2006

Purification of histone demethylases from HeLa cells.

Posttranslational histone modifications play an important role in regulating chromatin dynamics and function. One of the modifications, methylation, occurs on both lysine and arginine residues and participates in diverse range of biological processes including heterochromatin formation, X-chromosome inactivation, and transcriptional regulation. While acetylation, phosphorylation, and ubiquitylation are dynamically regulated by enzymes that catalyze the addition and removal of a particular modification, enzymes that are capable of removing methyl groups were not known until recently. Thus far, two families of histone demethylases with distinct cofactor requirements and reaction mechanisms have been identified. One is the FAD (flavin adenine dinucleotide)-dependent amine oxidase family LSD1 (lysine-specific demethylase), the other is the Fe(II) and alpha-KG (alpha-ketoglutarate)-dependent dioxygenase family JHDM (JmjC domain-containing histone demethylase). Identification and characterization of these histone demethylases is an important step towards understanding both the function and regulation of histone methylation. Here, we describe assays currently used for measuring histone demethylase activity and chromatography strategies used in purifying histone demethylases from HeLa cells.

Chromatography↗

PubMed2006

Functional interplay between histone demethylase and deacetylase enzymes.

Histone deacetylase (HDAC) inhibitors are a promising class of anticancer agents for the treatment of solid and hematological malignancies. The precise mechanism by which HDAC inhibitors mediate their effects on tumor cell growth, differentiation, and/or apoptosis is the subject of intense research. Previously we described a family of multiprotein complexes that contain histone deacetylase 1/2 (HDAC1/2) and the histone demethylase BHC110 (LSD1). Here we show that HDAC inhibitors diminish histone H3 lysine 4 (H3K4) demethylation by BHC110 in vitro. In vivo analysis revealed an increased H3K4 methylation concomitant with inhibition of nucleosomal deacetylation by HDAC inhibitors. Reconstitution of recombinant complexes revealed a functional connection between HDAC1 and BHC110 only when nucleosomal substrates were used. Importantly, while the enzymatic activity of BHC110 is required to achieve optimal deacetylation in vitro, in vivo analysis following ectopic expression of an enzymatically dead mutant of BHC110 (K661A) confirmed the functional cross talk between the demethylase and deacetylase enzymes. Our studies not only reveal an intimate link between the histone demethylase and deacetylase enzymes but also identify histone demethylation as a secondary target of HDAC inhibitors.

Acetylation↗

PubMed2006

Diversity within the JMJD2 histone demethylase family.

JMJD2A-D belong to the JmjC domain-containing family of histone demethylases. JMJD2D is the most structurally divergent JMJD2 protein as it lacks the PHD and Tudor domains present in JMJD2A-C. Here, we systematically analyzed the histone demethylase specificity of JMJD2 proteins in vivo. We found that JMJD2A and C demethylate tri- and dimethylated H3K9 and H3K36, whereas JMJD2D demethylates tri-, di-, and monomethylated H3K9. Enzymatic activity requires the N-terminal JmjN domain. It also contributes to efficient nuclear localization together with the PHD and Tudor domains of JMJD2A and C. Furthermore, JMJD2 proteins form homomers, and JMJD2A and C, but not JMJD2D, can also heteromerize. Finally, we show that JMJD2 proteins promoter-specifically repress or activate gene transcription. Altogether, our results reveal novel properties of and functional differences between JMJD2 proteins that may therefore have different effects on chromatin structure.

Blotting, Western↗

PubMed2025

Mechanism of histone demethylase KDM5A in osteoporotic fracture healing through epigenetic regulation of the miR-495/SKP2/Runx2 axis.

BACKGROUND: Osteoporosis represents a salient metabolic bone disorder. Histone demethylase plays a vital role in bone development and homeostasis. This study explored the mechanism of histone demethylase KDM5A affecting osteoporotic fracture healing via the miR-495/SKP2/Runx2 axis. METHODS: The murine model of osteoporotic fracture was established. The bone mineral density, maximum elastic stress, and maximum load were tested. The relative trabecular bone volume, bone trabecular thickness, and trabecular number at the proximal end of tibia were detected. The histopathological changes of femur tissues and bone microstructure were observed. Expressions of KDM5A and osteogenic factors were detected. The cell proliferation, alkaline phosphatase activity, and calcified nodules were measured. The binding relationships between KDM5A and miR-495 promoter, and miR-495 and SKP2 were verified. The interaction between SKP2 and Runx2 was detected. The ubiquitination level of Runx2 and the stability of Runx2 protein were detected. RESULTS: KDM5A was highly expressed in the murine model of osteoporotic fracture. Interference of KDM5A expression facilitated fracture healing in osteoporotic mice. KDM5A downregulated miR-495 expression by promoting the H3K4me3 methylation of the miR-495 promoter. Inhibition of miR-495 reversed the effect of KDM5A silencing on osteoblast proliferation, differentiation, and mineralization. miR-495 facilitated osteoblast proliferation, differentiation, and mineralization by targeting SKP2. SKP2 suppressed Runx2 expression through ubiquitination degradation. Inhibition of Runx2 reversed the promoting effect of SKP2 silencing on osteogenic differentiation. CONCLUSION: KDM5A attenuated the inhibition of miR-495 on SKP2 and promoted the ubiquitination degradation of Runx2 protein by SKP2, thereby repressing osteoblast differentiation and retarding osteoporotic fracture healing.

Animals↗

PubMed2005

Regulation of LSD1 histone demethylase activity by its associated factors.

LSD1 is a recently identified human lysine (K)-specific histone demethylase. LSD1 is associated with HDAC1/2; CoREST, a SANT domain-containing corepressor; and BHC80, a PHD domain-containing protein, among others. We show that CoREST endows LSD1 with the ability to demethylate nucleosomal substrates and that it protects LSD1 from proteasomal degradation in vivo. We find hyperacetylated nucleosomes less susceptible to CoREST/LSD1-mediated demethylation, suggesting that hypoacetylated nucleosomes may be the preferred physiological substrates. This raises the possibility that histone deacetylases and LSD1 may collaborate to generate a repressive chromatin environment. Consistent with this model, TSA treatment results in derepression of LSD1 target genes. While CoREST positively regulates LSD1 function, BHC80 inhibits CoREST/LSD1-mediated demethylation in vitro and may therefore confer negative regulation. Taken together, these findings suggest that LSD1-mediated histone demethylation is regulated dynamically in vivo. This is expected to have profound effects on gene expression under both physiological and pathological conditions.

Enzyme Stability↗

PubMed2024

Targeting the transcription factor YY1 is synthetic lethal with loss of the histone demethylase KDM5C.

An understanding of the enzymatic and scaffolding functions of epigenetic modifiers is important for the development of epigenetic therapies for cancer. The H3K4me2/3 histone demethylase KDM5C has been shown to regulate transcription. The diverse roles of KDM5C are likely determined by its interacting partners, which are still largely unknown. In this study, we screen for KDM5C-binding proteins and show that YY1 interacts with KDM5C. A synergistic antitumor effect is exerted when both KDM5C and YY1 are depleted, and targeting YY1 appears to be a vulnerability in KDM5C-deficient cancer cells. Mechanistically, KDM5C promotes global YY1 chromatin recruitment, especially at promoters. Moreover, an intact KDM5C JmjC domain but not KDM5C histone demethylase activity is required for KDM5C-mediated YY1 chromatin binding. Transcriptional profiling reveals that dual inhibition of KDM5C and YY1 increases transcriptional repression of cell cycle- and apoptosis-related genes. In summary, our work demonstrates a synthetic lethal interaction between YY1 and KDM5C and suggests combination therapies for cancer treatments.

YY1 Transcription Factor↗

PubMed2006

Solution structure of the SWIRM domain of human histone demethylase LSD1.

SWIRM is an evolutionarily conserved domain involved in several chromatin-modifying complexes. Recently, the LSD1 protein, which bears a SWIRM domain, was found to be a demethylase for Lys4-methylated histone H3. Here, we report a solution structure of the SWIRM domain of human LSD1. It forms a compact fold composed of 6 alpha helices, in which a 20 amino acid long helix (alpha4) is surrounded by 5 other short helices. The SWIRM domain structure could be divided into the N-terminal part (alpha1-alpha3) and the C-terminal part (alpha4-alpha6), which are connected to each other by a salt bridge. While the N-terminal part forms a SWIRM-specific structure, the C-terminal part adopts a helix-turn-helix (HTH)-related fold. We discuss a model in which the SWIRM domain acts as an anchor site for a histone tail.

Amino Acid Motifs↗

PubMed2002

A second catalytic domain in the Elp3 histone acetyltransferases: a candidate for histone demethylase activity?

A new subfamily of two-domain histone acetyltransferases (HATs) related to Elp3 has been identified. In addition to a HAT domain in the C terminus, these proteins have an N-terminal domain similar to the catalytic domain of S-adenosylmethionine radical enzymes. Two-domain organization is preserved in evolution, suggesting that both enzymatic activities are functionally or mechanistically coupled and directed towards highly conserved substrates. The functional implications of this similarity and a possible role for Elp3-related proteins as histone demethylases are discussed.

Acetyltransferases↗

PubMed2026

Histone demethylase PHF2 drives olanzapine-induced dyslipidemia via epigenomic rewiring of hepatic lipogenic genes.

Olanzapine, an atypical antipsychotic agent, is widely used in treating psychotic disorders, yet its metabolic side effects remain a clinical concern. Emerging evidence suggests that dynamic alterations in histone methylation are implicated in olanzapine-induced hepatic lipid metabolic disorders. PHF2, a JmjC family histone demethylase mediating H3K9me2 demethylation, functions as a transcriptional repressor by regulating downstream targets. To elucidate PHF2's role in this process, we utilized an olanzapine-induced dyslipidemia rat model. ChIP-qPCR analysis demonstrated a significant reduction in dimethylated histone H3 lysine 9 (H3K9me2) on the promoters of lipogenic genes (Fasn, Acc1, Scd1) in the liver, accompanied by elevated nuclear expression of PHF2 in olanzapine-treated rats. Co-immunoprecipitation (Co-IP) assays revealed a physical interaction between PHF2 and ChREBP, a glucose-responsive lipogenic transcription factor. Olanzapine was found to enhance the formation of this complex. Overexpression of PHF2 led to upregulated protein levels of FASN/ACC1 and intracellular lipid accumulation, whereas knockdown of PHF2 using siRNA attenuated these effects. Notably, the upregulation of FASN/ACC1 expression induced by olanzapine was markedly diminished in PHF2-deficient AML12 cells via ChREBP-PHF2-mediated H3K9me2 demethylation. Additionally, olanzapine inhibited the nuclear translocation of FOXA2, a PHF2 transcriptional regulator, thereby augmenting PHF2 expression. These findings uncover a novel epigenetic mechanism underlying olanzapine-induced dyslipidemia, positioning the FOXA2-PHF2-ChREBP axis as a potential therapeutic target through modulation of hepatic histone methylation.

Animals↗

PubMed2006

Androgen receptor coactivators lysine-specific histone demethylase 1 and four and a half LIM domain protein 2 predict risk of prostate cancer recurrence.

Prostate cancer biology varies from locally confined tumors with low risk for relapse to tumors with high risk for progression even after radical prostatectomy. Currently, there are no reliable biomarkers to predict tumor relapse and poor clinical outcome. In this study, we correlated expression patterns of the androgen receptor (AR) coactivators lysine-specific histone demethylase 1 (LSD1) and four and a half LIM-domain protein 2 (FHL2), AR, Gleason score, Gleason grade, and p53 expression in clinically organ confined prostate cancers with relapse after radical prostatectomy. Our data reveal that high levels of LSD1, nuclear expression of the FHL2 coactivator, high Gleason score and grade, and very strong staining of nuclear p53 correlate significantly with relapse during follow-up. No correlation exists with relapse and the expression of AR and cytoplasmic expression of FHL2. To confirm these data, we did quantitative reverse transcription-PCR and Western blot analyses in a subset of tumor specimens. Consistently, both LSD1 mRNA and protein levels were significantly up-regulated in high-risk tumors. We previously identified LSD1 and FHL2 as nuclear cofactors interacting specifically with the AR in prostate cells and showed that both stimulate androgen-dependent gene transcription. Our present study suggests that LSD1 and nuclear FHL2 may serve as novel biomarkers predictive for prostate cancer with aggressive biology and point to a role of LSD1 and FHL2 in constitutive activation of AR-mediated growth signals.

Aged↗

PubMed2006

A highly specific mechanism of histone H3-K4 recognition by histone demethylase LSD1.

Human lysine-specific demethylase (LSD1) is a chromatin-modifying enzyme that specifically removes methyl groups from mono- and dimethylated Lys4 of histone H3 (H3-K4). We used a combination of in vivo and in vitro experiments to characterize the substrate specificity and recognition by LSD1. Biochemical assays on histone peptides show that essentially all epigenetic modifications on the 21 N-terminal amino acids of histone H3 cause a significant reduction in enzymatic activity. Replacement of Lys4 with Arg greatly enhances binding affinity, and a histone peptide incorporating this mutation has a strong inhibitory power. Conversely, a peptide bearing a trimethylated Lys4 is only a weak inhibitor of the enzyme. Rapid kinetics measurements evidence that the enzyme is efficiently reoxidized by molecular oxygen with a second-order rate constant of 9.6x10(3) M-1 s-1, and that the presence of the reaction product does not greatly influence the rate of flavin reoxidation. In vivo experiments provide a correlation between the in vitro inhibitory properties of the tested peptides and their ability of affecting endogenous LSD1 activity. Our results show that epigenetic modifications on histone H3 need to be removed before Lys4 demethylation can efficiently occur. The complex formed by LSD1 with histone deacetylases 1/2 may function as a "double-blade razor" that first eliminates the acetyl groups from acetylated Lys residues and then removes the methyl group from Lys4. We suggest that after H3-K4 demethylation, LSD1 recruits the forthcoming chromatin remodelers leading to the introduction of gene repression marks.

Cell Line↗

PubMed2004

Histone demethylation mediated by the nuclear amine oxidase homolog LSD1.

Posttranslational modifications of histone N-terminal tails impact chromatin structure and gene transcription. While the extent of histone acetylation is determined by both acetyltransferases and deacetylases, it has been unclear whether histone methylation is also regulated by enzymes with opposing activities. Here, we provide evidence that LSD1 (KIAA0601), a nuclear homolog of amine oxidases, functions as a histone demethylase and transcriptional corepressor. LSD1 specifically demethylates histone H3 lysine 4, which is linked to active transcription. Lysine demethylation occurs via an oxidation reaction that generates formaldehyde. Importantly, RNAi inhibition of LSD1 causes an increase in H3 lysine 4 methylation and concomitant derepression of target genes, suggesting that LSD1 represses transcription via histone demethylation. The results thus identify a histone demethylase conserved from S. pombe to human and reveal dynamic regulation of histone methylation by both histone methylases and demethylases.

Conserved Sequence↗

PubMed2006

Histone lysine demethylases and their impact on epigenetics.

Methylation marks on the lysine residues of histone proteins are thought to contribute to epigenetic phenomena in part because of their apparent irreversibility. Will this view change with the recent discovery of histone lysine demethylases that reversibly remove methyl marks?

Animals↗

PubMed2006

The transcriptional repressor JHDM3A demethylates trimethyl histone H3 lysine 9 and lysine 36.

Post-translational modification of chromatin has profound effects on many biological processes including transcriptional regulation, heterochromatin organization, and X-chromosome inactivation. Recent studies indicate that methylation on specific histone lysine (K) residues participates in many of these processes. Lysine methylation occurs in three distinct states, having either one (me1), two (me2) or three (me3) methyl groups attached to the amine group of the lysine side chain. These differences in modification state have an important role in defining how methylated chromatin is recognized and interpreted. Until recently, histone lysine methylation was considered a stable modification, but the identification of histone demethylase enzymes has demonstrated the reversibility of this epigenetic mark. So far, all characterized histone demethylases show enzymatic activity towards lysine residues modified in the me1 or me2 state, leaving open the possibility that me3 constitutes an irreversible modification. Here we demonstrate that JHDM3A (jumonji C (JmjC)-domain-containing histone demethylase 3A; also known as JMJD2A) is capable of removing the me3 group from modified H3 lysine 9 (H3K9) and H3 lysine 36 (H3K36). Overexpression of JHDM3A abrogates recruitment of HP1 (heterochromatin protein 1) to heterochromatin, indicating a role for JHDM3A in antagonizing methylated H3K9 nucleated events. siRNA-mediated knockdown of JHDM3A leads to increased levels of H3K9 methylation and upregulation of a JHDM3A target gene, ASCL2, indicating that JHDM3A may function in euchromatin to remove histone methylation marks that are associated with active transcription.

Animals↗