RESUMEN
SET/TAF-Iß, a subunit of the inhibitor of acetyltransferases (INHAT) complex, exhibits transcriptional repression activity by inhibiting histone acetylation. We find that SET/TAF-Iß regulates mono-ubiquitination of histone H2A at lysine 119 (H2AK119ub), which is involved in polycomb-mediated transcriptional repression, in HCT116 cells. In this report, we demonstrate that SET/TAF-Iß acts as an E2 ubiquitin-conjugating enzyme for PRC1-independent H2AK119ub. Furthermore, we identify that MIB1 is the E3 ligase partner for SET/TAF-Iß using LC-MS/MS and in vitro ubiquitination assays. Transcriptome analysis reveals that SET/TAF-Iß and MIB1 regulate the expression of genes related to DNA replication and cell cycle progression in HCT116 cells, and knockdown of either protein reduces proliferation of HCT116 cells by impeding cell cycle progression. Together, our study reveals a novel PRC1-independent epigenetic regulatory mechanism for H2AK119ub by SET/TAF-Iß and MIB1 in colon cancer.
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Histone modifications are a key mechanism underlying the epigenetic regulation of gene expression, which is critically involved in the consolidation of multiple forms of memory. However, the roles of histone modifications in cerebellum-dependent motor learning and memory are not well understood. To test whether changes in histone methylation are involved in cerebellar learning, we used heterozygous Kdm3b knockout (Kdm3b+/-) mice, which show reduced lysine 9 on histone 3 (H3K9) demethylase activity. H3K9 di-methylation is significantly increased selectively in the granule cell layer of the cerebellum of Kdm3b+/- mice. In the cerebellum-dependent optokinetic response (OKR) learning, Kdm3b+/- mice show deficits in memory consolidation, whereas they are normal in basal oculomotor performance and OKR acquisition. In addition, RNA-seq analyses revealed that the expression levels of several plasticity-related genes were altered in the mutant cerebellum. Our study suggests that active regulation of histone methylation is critical for the consolidation of cerebellar motor memory.
Asunto(s)
Cerebelo/fisiología , Haploinsuficiencia/genética , Histona Demetilasas con Dominio de Jumonji/genética , Consolidación de la Memoria/fisiología , Actividad Motora/fisiología , Animales , Regulación de la Expresión Génica , Histonas/metabolismo , Histona Demetilasas con Dominio de Jumonji/metabolismo , Lisina/metabolismo , Masculino , Metilación , Ratones Endogámicos C57BLRESUMEN
Acute myeloid leukemia (AML) is the most common type of leukemia in adults. Owing to the chemotherapy associated side effects and toxicity, it is necessary to find a new mechanism, which can identify new potential therapeutic targets at the molecular level. Here, we identified new target genes that are induced during the TPA-induced HL-60 cell differentiation by ChIP-seq and microarray data analysis. Using q-PCR and ChIP assay, we confirmed that the target genes including USP3, USP35, TCF4, and SGK1 are upregulated during TPA-mediated HL-60 cell differentiation. Levels of USP3, one of the deubiquitinating enzymes (DUBs), increased by TPA treatment, resulting in the reduction of H2AK119ub levels. In addition, we revealed that depletion of USP3 inhibits TPA-mediated leukemia cell differentiation q-PCR and FACS analysis. Taken together, our data indicate that USP3 promotes TPA-mediated leukemia cell differentiation via regulating H2AK119ub levels.
RESUMEN
Posttranslational modifications of the Forkhead family transcription factor, FOXO1, have been known to have important regulatory implications in its diverse activities. Several types of modifications of FOXO1, including acetylation, phosphorylation, and ubiquitination, have been reported. However, lysine methylation of FOXO1 has not yet been identified. Here, we reported that FOXO1 is methylated by G9a at K273 residue in vitro and in vivo. Methylation of FOXO1 by G9a increased interaction between FOXO1 and a specific E3 ligase, SKP2, and decreased FOXO1 protein stability. In addition, G9a expression was increased by insulin and resulted in insulin-mediated FOXO1 degradation by K273 methylation. Tissue array analysis indicated that G9a was overexpressed and FOXO1 levels decreased in human colon cancer. Cell proliferation assays revealed that G9a-mediated FOXO1 methylation increased colon cancer cell proliferation. Fluorescence-activated cell sorting (FACS) analysis indicated that apoptosis rates were higher in the presence of FOXO1 than in FOXO1 knock-out cells. Furthermore, we found that G9a protein levels were elevated and FOXO1 protein levels were decreased in human colon cancer patients tissue samples. Here, we report that G9a specific inhibitor, BIX-01294, can regulate cell proliferation and apoptosis by inhibiting G9a-mediated FOXO1 methylation.
Asunto(s)
Neoplasias del Colon/genética , Proteína Forkhead Box O1/genética , Antígenos de Histocompatibilidad/genética , N-Metiltransferasa de Histona-Lisina/genética , Proteínas Quinasas Asociadas a Fase-S/genética , Apoptosis/genética , Azepinas/farmacología , Sistemas CRISPR-Cas/genética , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/patología , Metilación de ADN/genética , Femenino , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Inactivación de Genes , Células HCT116 , N-Metiltransferasa de Histona-Lisina/antagonistas & inhibidores , Humanos , Masculino , Quinazolinas/farmacología , Análisis de Matrices Tisulares , Ubiquitinación/genéticaRESUMEN
A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.
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The human myeloid leukemia cell line HL-60 differentiate into monocytes following treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA). However, the mechanism underlying the differentiation of these cells in response to TPA has not been fully elucidated. In this study, we performed ChIP-seq profiling of RNA Pol II, HDAC2, Acetyl H3 (AcH3), and H3K27me3 and analyzed differential chromatin state changes during TPA-induced differentiation of HL-60 cells. We focused on atypically active genes, which showed enhanced H3 acetylation despite increased HDAC2 recruitment. We found that HDAC2 positively regulates the expression of these genes in a histone deacetylase activity-independent manner. HDAC2 interacted with and recruited paired box 5 (PAX5) to the promoters of the target genes and regulated HL-60 cell differentiation by PAX5-mediated gene activation. Taken together, these data elucidated the specific-chromatin status during HL-60 cell differentiation following TPA exposure and suggested that HDAC2 can activate transcription of certain genes through interactions with PAX5 in a deacetylase activity-independent pathway.
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Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Histona Desacetilasa 2/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Activación Transcripcional/efectos de los fármacos , Cromatina/efectos de los fármacos , Cromatina/metabolismo , Células HL-60 , Humanos , Factor de Transcripción PAX5/metabolismoRESUMEN
The histone methyltransferase multiple myeloma SET domain protein (MMSET/WHSC1) is highly expressed in diverse tumor types, and its expression appears to be involved in cell proliferation. In this study, we report that MMSET interacts with and methylates Aurora kinase A (AURKA). We show that MMSET-mediated methylation of AURKA induces interaction with p53 as well as enhanced kinase activity of AURKA, which results in the proteasomal degradation of p53. MMSET-mediated p53 degradation increases cell proliferation and results in oncogenic activity. Furthermore, knockdown of MMSET potently inhibits tumorigenic cells and renders them sensitive to growth inhibition by the therapeutic drug, alisertib (AURKA inhibitor). Taken together, our results suggest that MMSET is a regulator of p53 stability via methylation of AURKA in proliferating cells and might be a potential therapeutic target in solid tumors.
Asunto(s)
Apoptosis/genética , Aurora Quinasa A/genética , Proliferación Celular/genética , N-Metiltransferasa de Histona-Lisina/genética , Proteínas Represoras/genética , Proteína p53 Supresora de Tumor/genética , Carcinogénesis/genética , Carcinogénesis/patología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/genética , Células HCT116 , Humanos , Metilación , Neoplasias/genética , Neoplasias/patologíaRESUMEN
The methylation of histone H3 lysine 79 (H3K79) is an active chromatin marker and is prominent in actively transcribed regions of the genome; however, demethylase of H3K79 remains unknown despite intensive research. Here, we show that KDM2B, also known as FBXL10 and a member of the Jumonji C family of proteins known for its histone H3K36 demethylase activity, is a di- and trimethyl H3K79 demethylase. We demonstrate that KDM2B induces transcriptional repression of HOXA7 and MEIS1 via occupancy of promoters and demethylation of H3K79. Furthermore, genome-wide analysis suggests that H3K79 methylation levels increase when KDM2B is depleted, which indicates that KDM2B functions as an H3K79 demethylase in vivo. Finally, stable KDM2B-knockdown cell lines exhibit displacement of NAD+-dependent deacetylase sirtuin-1 (SIRT1) from chromatin, with concomitant increases in H3K79 methylation and H4K16 acetylation. Our findings identify KDM2B as an H3K79 demethylase and link its function to transcriptional repression via SIRT1-mediated chromatin silencing.-Kang, J.-Y., Kim, J.-Y., Kim, K.-B., Park, J. W., Cho, H., Hahm, J. Y., Chae, Y.-C., Kim, D., Kook, H., Rhee, S., Ha, N.-C., Seo, S.-B. KDM2B is a histone H3K79 demethylase and induces transcriptional repression via sirtuin-1-mediated chromatin silencing.
Asunto(s)
Cromatina/metabolismo , Proteínas F-Box/metabolismo , Silenciador del Gen , Proteínas de Homeodominio/biosíntesis , Histona Demetilasas con Dominio de Jumonji/metabolismo , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/biosíntesis , Sirtuina 1/metabolismo , Transcripción Genética , Cromatina/genética , Proteínas F-Box/genética , Proteínas de Homeodominio/genética , Humanos , Histona Demetilasas con Dominio de Jumonji/genética , Células K562 , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/genética , Sirtuina 1/genéticaRESUMEN
Recent studies have reported the ectopic expression of olfactory receptors (ORs) in non-olfactory tissues, however, their physiological roles were not well elucidated. ORs are expressed in and function in different types of cancers. Here, we identified that the H3K9me2 levels of several OR promoters decreased during differentiation in the HL-60, human myeloid leukaemia cell line, by all-trans-retinoic acid (ATRA). We found that the differential OR promoters H3K9me2 levels were regulated by G9a and LSD1, resulting in the decrease of ORs transcription during HL-60 differentiation. G9a and LSD1 could regulate the expression of ORs in several non-olfactory cells via the methylation and demethylation of H3K9me2. In addition, we demonstrated that knockdown of OR significantly reduced cell proliferation. Therefore, the epigenetic regulation of ORs transcription is critical for carcinogenesis.
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Regulación Leucémica de la Expresión Génica , Antígenos de Histocompatibilidad/metabolismo , Histona Demetilasas/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Leucemia/genética , Leucemia/metabolismo , Receptores Odorantes/genética , Transcripción Genética , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Línea Celular Tumoral , Proliferación Celular , Metilación de ADN , Técnicas de Silenciamiento del Gen , Células HL-60 , Histonas/metabolismo , Humanos , Leucemia/patología , Tretinoina/farmacologíaRESUMEN
Data derived from genomic and transcriptomic analyses have revealed that long noncoding RNAs (lncRNAs) have important roles in the transcriptional regulation of various genes. Recent studies have identified the mechanism underlying this function. To date, a variety of noncoding transcripts have been reported to function in conjunction with epigenetic regulator proteins. In this study, we investigated the function of linc00598, which is transcribed by a genomic sequence on chromosome 13, downstream of FoxO1 and upstream of COG6. Microarray analysis showed that linc00598 regulates the transcription of specific target genes, including those for cell cycle regulators. We discovered that linc00598 regulates CCND2 transcription through modulation of the transcriptional regulatory effect of FoxO1 on the CCND2 promoter. Moreover, we observed that knockdown of linc00598 induced G0/G1 cell cycle arrest and inhibited proliferation. These data indicate that linc00598 plays an important role in cell cycle regulation and proliferation through its ability to regulate the transcription of CCND2.
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Ciclina D2/biosíntesis , Puntos de Control de la Fase G1 del Ciclo Celular/fisiología , ARN Largo no Codificante/metabolismo , Elementos de Respuesta/fisiología , Transcripción Genética/fisiología , Ciclina D2/genética , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Células HEK293 , Células HL-60 , Células HeLa , Células Hep G2 , Humanos , Células K562 , Células MCF-7 , ARN Largo no Codificante/genética , Fase de Descanso del Ciclo Celular/fisiología , Células THP-1RESUMEN
Histone H3S10 phosphorylation has been known as a cell cycle-specific marker and has a role in transcriptional activation. Various kinases phosphorylate H3S10 in different species, however, the role of the mitotic serine/threonine protein kinase Aurora A (AURKA) is largely unknown. Here we present evidence that AURKA phosphorylates H3S10 and activates target gene transcription. We show that down-regulation of AURKA level during leukemia cell differentiation results in decreased H3S10 phosphorylation level. We further show that AURKA is recruited to target gene promoters and activates transcription via H3S10 phosphorylation. Furthermore, this recruitment can be disrupted by the AURKA inhibitor Alisertib and results in H3K9-me2 recruitment by G9a.
Asunto(s)
Aurora Quinasa A/genética , Código de Histonas/genética , Histonas/genética , Neoplasias Experimentales/genética , Regiones Promotoras Genéticas/genética , Activación Transcripcional/genética , Línea Celular Tumoral , Regulación de la Expresión Génica/genética , Células HL-60 , Humanos , Fosforilación , Transcripción Genética/genéticaRESUMEN
Histone lysine methylation contributes to transcriptional regulation by serving as a platform for the recruitment of various cofactors. Intense studies have been conducted for elucidating the functional meaning of H3K79 methylation, and to date, the only known HMTase responsible for the modification was DOT1L. In this study, we report that the MMSET isoform RE-IIBP has HMTase activity for H3K79. It was uncovered that RE-IIBP up-regulates MEIS1 transcription through H3K79 methylation via recruitment to the MEIS1 promoter. By means of proteomic and biochemical analysis, association of RE-IIBP with the E3 ubiquitin ligase RNF20 was demonstrated for synergistic activation of MEIS1 transcription via H3K79 HMTase activity. Furthermore, It was observed that RE-IIBP induces MEIS1-mediated apoptosis, which was dependent on H2BK120 ubiquitination by RNF20. These findings suggest RE-IIBP as another candidate for further studies to elucidate the mechanism of H3K79 methylation and its biological functions.
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Apoptosis/fisiología , N-Metiltransferasa de Histona-Lisina/metabolismo , Histonas/metabolismo , Proteínas de Homeodominio/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Represoras/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación/fisiología , Línea Celular , N-Metiltransferasa de Histona-Lisina/genética , Histonas/genética , Proteínas de Homeodominio/genética , Humanos , Metilación , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide , Proteínas de Neoplasias/genética , Proteínas Represoras/genética , Transcripción Genética/fisiología , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Here we report that the H3K9 demethylase KDM3B represses transcription of the angiogenesis regulatory gene, ANGPT1. Negative regulation of ANGPT1 by KDM3B is independent of its Jumonji (JmjC) domain-mediated H3K9 demethylase activity. We demonstrate that KDM3B downregulates ANGPT1 via interaction with SMRT, and suggest that the repressor complex is formed at the promoter area of ANGPT1. Using MTT and wound healing assays, depletion of KDM3B was found to increase cell proliferation and cell motility, indicating that KDM3B has a role in angiogenesis.
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Angiopoyetina 1/genética , Histonas/metabolismo , Histona Demetilasas con Dominio de Jumonji/metabolismo , Lisina/metabolismo , Proteínas Represoras/metabolismo , Transcripción Genética , Angiopoyetina 1/metabolismo , Movimiento Celular , Proliferación Celular , Proteínas Co-Represoras/metabolismo , Regulación de la Expresión Génica , Células HCT116 , Humanos , Células K562 , Células MCF-7 , Co-Represor 2 de Receptor Nuclear/metabolismo , Unión ProteicaRESUMEN
Post-translational modification of forkhead family transcription factor, FoxO1, is an important regulatory mode for its diverse activities. FoxO1 is acetylated by HAT coactivators and its transcriptional activity is decreased via reduced DNA binding affinity. Here, we report that SET/TAF-Iß inhibited p300-mediated FoxO1 acetylation in an INHAT domain-dependent manner. SET/TAF-Iß interacted with FoxO1 and activated transcription of FoxO1 target gene, p21. Moreover, SET/TAF-Iß inhibited acetylation of FoxO1 and increased p21 transcription induced by oxidative stress. Our results suggest that SET/TAF-Iß inhibits FoxO1 acetylation and activates its transcriptional activity toward p21.
Asunto(s)
Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Factores de Transcripción Forkhead/metabolismo , Chaperonas de Histonas/metabolismo , Subunidades de Proteína/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética , Acetilación , Apoptosis , Proteínas de Unión al ADN , Proteína Forkhead Box O1 , Regulación de la Expresión Génica , Células HCT116 , Células HEK293 , Humanos , Estrés OxidativoRESUMEN
We report that H3K9 HMTase G9a activates transcription of the cell cycle regulatory gene, p21, in p53-null H1299 cells. Positive regulation of p21 by G9a is independent of its HMTase activity. We demonstrate that G9a upregulates p21 via interaction with PCAF, and provide evidence that the activating complex is recruited to the p21 promoter upon DNA damage-inducing agent etoposide treatment. Our study suggests that G9a decreases proliferation and cell viability by increasing the level of p21-mediated apoptosis. Our results suggest that G9a functions as a coactivator for p21 transcription, and directs cells to undergo apoptosis.
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Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Antígenos de Histocompatibilidad/fisiología , N-Metiltransferasa de Histona-Lisina/fisiología , Activación Transcripcional , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Daño del ADN , Células HEK293 , Humanos , Regiones Promotoras Genéticas , Unión Proteica , ARN Mensajero/genética , ARN Mensajero/metabolismo , Regulación hacia Arriba , Factores de Transcripción p300-CBP/metabolismoRESUMEN
Significant progress has been made in understanding the relationship between histone modifications and 'reader' molecules and their effects on transcriptional regulation. A previously identified INHAT complex subunit, SET/TAF-Iß, binds to histones and inhibits histone acetylation. To investigate the binding specificities of SET/TAF-Iß to various histone modifications, we employed modified histone tail peptide array analyses. SET/TAF-Iß strongly recognized PRC2-mediated H3K27me1/2/3; however, the bindings were completely disrupted by H3S28 phosphorylation. We have demonstrated that SET/TAF-Iß is sequentially recruited to the target gene promoter ATF3 after the PRC2 complex via H3K27me recognition and may offer additive effects in the repression of the target gene.
