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1.
Genes Dev ; 37(21-24): 984-997, 2023 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-37993255

RESUMO

The RING-type E3 ligase has been known for over two decades, yet its diverse modes of action are still the subject of active research. Plant homeodomain (PHD) finger protein 7 (PHF7) is a RING-type E3 ubiquitin ligase responsible for histone ubiquitination. PHF7 comprises three zinc finger domains: an extended PHD (ePHD), a RING domain, and a PHD. While the function of the RING domain is largely understood, the roles of the other two domains in E3 ligase activity remain elusive. Here, we present the crystal structure of PHF7 in complex with the E2 ubiquitin-conjugating enzyme (E2). Our structure shows that E2 is effectively captured between the RING domain and the C-terminal PHD, facilitating E2 recruitment through direct contact. In addition, through in vitro binding and functional assays, we demonstrate that the N-terminal ePHD recognizes the nucleosome via DNA binding, whereas the C-terminal PHD is involved in histone H3 recognition. Our results provide a molecular basis for the E3 ligase activity of PHF7 and uncover the specific yet collaborative contributions of each domain to the PHF7 ubiquitination activity.


Assuntos
Histonas , Ubiquitina-Proteína Ligases , Histonas/metabolismo , Ubiquitinação , Ubiquitina-Proteína Ligases/metabolismo , Proteínas de Ligação a DNA/metabolismo , Dedos de Zinco , Enzimas de Conjugação de Ubiquitina/metabolismo
2.
Mol Cell ; 69(3): 398-411.e6, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29395062

RESUMO

The inflammatory response mediated by nuclear factor κB (NF-κB) signaling is essential for host defense against pathogens. Although the regulatory mechanism of NF-κB signaling has been well studied, the molecular basis for epigenetic regulation of the inflammatory response is poorly understood. Here we identify a new signaling axis of PKCα-LSD1-NF-κB, which is critical for activation and amplification of the inflammatory response. In response to excessive inflammatory stimuli, PKCα translocates to the nucleus and phosphorylates LSD1. LSD1 phosphorylation is required for p65 binding and facilitates p65 demethylation, leading to enhanced stability. In vivo genetic analysis using Lsd1SA/SA mice with ablation of LSD1 phosphorylation and chemical approaches in wild-type mice with inhibition of PKCα or LSD1 activity show attenuated sepsis-induced inflammatory lung injury and mortality. Together, we demonstrate that the PKCα-LSD1-NF-κB signaling cascade is crucial for epigenetic control of the inflammatory response, and targeting this signaling could be a powerful therapeutic strategy for systemic inflammatory diseases, including sepsis.


Assuntos
Histona Desmetilases/metabolismo , Proteína Quinase C/metabolismo , Animais , Núcleo Celular/metabolismo , Epigênese Genética/genética , Histona Desmetilases/genética , Inflamação/metabolismo , Metilação , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fosforilação , Proteína Quinase C/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/genética , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
3.
Mol Cell ; 65(5): 781-785, 2017 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-28257699

RESUMO

Autophagy is an evolutionarily conserved catabolic process. Although the components of autophagy in cytoplasm have been well-studied, the molecular basis for the epigenetic regulation of autophagy is poorly understood. It is becoming more important to propose a "whole-cell view" of autophagy embracing both cytoplasmic and nuclear events. Thus, it is great timing to summarize current status and discuss future direction.


Assuntos
Autofagia , Núcleo Celular/metabolismo , Cromatina/metabolismo , Epigênese Genética , Histonas/metabolismo , Acetilação , Animais , Autofagia/genética , Cromatina/genética , Montagem e Desmontagem da Cromatina , Humanos , Metilação , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Metiltransferases/metabolismo
4.
Nucleic Acids Res ; 50(13): 7298-7309, 2022 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-35801910

RESUMO

Autophagy, a catabolic process to remove unnecessary or dysfunctional organelles, is triggered by various signals including nutrient starvation. Depending on the types of the nutrient deficiency, diverse sensing mechanisms and signaling pathways orchestrate for transcriptional and epigenetic regulation of autophagy. However, our knowledge about nutrient type-specific transcriptional regulation during autophagy is limited. To understand nutrient type-dependent transcriptional mechanisms during autophagy, we performed single cell RNA sequencing (scRNAseq) in the mouse embryonic fibroblasts (MEFs) with or without glucose starvation (GS) as well as amino acid starvation (AAS). Trajectory analysis using scRNAseq identified sequential induction of potential transcriptional regulators for each condition. Gene regulatory rules inferred using TENET newly identified CCAAT/enhancer binding protein γ (C/EBPγ) as a regulator of autophagy in AAS, but not GS, condition, and knockdown experiment confirmed the TENET result. Cell biological and biochemical studies validated that activating transcription factor 4 (ATF4) is responsible for conferring specificity to C/EBPγ for the activation of autophagy genes under AAS, but not under GS condition. Together, our data identified C/EBPγ as a previously unidentified key regulator under AAS-induced autophagy.


Assuntos
Aminoácidos , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Transcriptoma , Fator 4 Ativador da Transcrição/metabolismo , Aminoácidos/genética , Aminoácidos/metabolismo , Animais , Autofagia/genética , Epigênese Genética , Fibroblastos/metabolismo , Camundongos , Análise de Célula Única
5.
Nucleic Acids Res ; 50(14): 7856-7872, 2022 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-35821310

RESUMO

Autophagy is a catabolic pathway that maintains cellular homeostasis under various stress conditions, including conditions of nutrient deprivation. To elevate autophagic flux to a sufficient level under stress conditions, transcriptional activation of autophagy genes occurs to replenish autophagy components. Thus, the transcriptional and epigenetic control of the genes regulating autophagy is essential for cellular homeostasis. Here, we applied integrated transcriptomic and epigenomic profiling to reveal the roles of plant homeodomain finger protein 20 (PHF20), which is an epigenetic reader possessing methyl binding activity, in controlling the expression of autophagy genes. Phf20 deficiency led to impaired autophagic flux and autophagy gene expression under glucose starvation. Interestingly, the genome-wide characterization of chromatin states by Assay for Transposase-Accessible Chromatin (ATAC)-sequencing revealed that the PHF20-dependent chromatin remodelling occurs in enhancers that are co-occupied by dimethylated lysine 36 on histone H3 (H3K36me2). Importantly, the recognition of H3K36me2 by PHF20 was found to be highly correlated with increased levels of H3K4me1/2 at the enhancer regions. Collectively, these results indicate that PHF20 regulates autophagy genes through enhancer activation via H3K36me2 recognition as an epigenetic reader. Our findings emphasize the importance of nuclear events in the regulation of autophagy.


Assuntos
Epigenômica , Inanição , Autofagia/genética , Cromatina/genética , Proteínas de Ligação a DNA/genética , Epigênese Genética , Proteínas de Homeodomínio/genética , Humanos , Inanição/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
7.
Nature ; 534(7608): 553-7, 2016 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-27309807

RESUMO

Autophagy is a highly conserved self-digestion process, which is essential for maintaining homeostasis and viability in response to nutrient starvation. Although the components of autophagy in the cytoplasm have been well studied, the molecular basis for the transcriptional and epigenetic regulation of autophagy is poorly understood. Here we identify co-activator-associated arginine methyltransferase 1 (CARM1) as a crucial component of autophagy in mammals. Notably, CARM1 stability is regulated by the SKP2-containing SCF (SKP1-cullin1-F-box protein) E3 ubiquitin ligase in the nucleus, but not in the cytoplasm, under nutrient-rich conditions. Furthermore, we show that nutrient starvation results in AMP-activated protein kinase (AMPK)-dependent phosphorylation of FOXO3a in the nucleus, which in turn transcriptionally represses SKP2. This repression leads to increased levels of CARM1 protein and subsequent increases in histone H3 Arg17 dimethylation. Genome-wide analyses reveal that CARM1 exerts transcriptional co-activator function on autophagy-related and lysosomal genes through transcription factor EB (TFEB). Our findings demonstrate that CARM1-dependent histone arginine methylation is a crucial nuclear event in autophagy, and identify a new signalling axis of AMPK-SKP2-CARM1 in the regulation of autophagy induction after nutrient starvation.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Autofagia/genética , Proteína-Arginina N-Metiltransferases/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Transdução de Sinais , Transcrição Gênica , Animais , Arginina/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Privação de Alimentos , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/deficiência , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Histonas/metabolismo , Humanos , Lisossomos/genética , Metilação , Camundongos , Fosforilação , Proteínas Quinases Associadas a Fase S/antagonistas & inibidores , Proteínas Ligases SKP Culina F-Box/química , Proteínas Ligases SKP Culina F-Box/metabolismo
8.
Mol Cell ; 56(2): 261-274, 2014 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-25219498

RESUMO

Biological roles for UFM1, a ubiquitin-like protein, are largely unknown, and therefore we screened for targets of ufmylation. Here we show that ufmylation of the nuclear receptor coactivator ASC1 is a key step for ERα transactivation in response to 17ß-estradiol (E2). In the absence of E2, the UFM1-specific protease UfSP2 was bound to ASC1, which maintains ASC1 in a nonufmylated state. In the presence of E2, ERα bound ASC1 and displaced UfSP2, leading to ASC1 ufmylation. Polyufmylation of ASC1 enhanced association of p300, SRC1, and ASC1 at promoters of ERα target genes. ASC1 overexpression or UfSP2 knockdown promoted ERα-mediated tumor formation in vivo, which could be abrogated by treatment with the anti-breast cancer drug tamoxifen. In contrast, expression of ufmylation-deficient ASC1 mutant or knockdown of the UFM1-activating E1 enzyme UBA5 prevented tumor growth. These findings establish a role for ASC1 ufmylation in breast cancer development by promoting ERα transactivation.


Assuntos
Sistema y+ de Transporte de Aminoácidos/metabolismo , Neoplasias da Mama/patologia , Receptor alfa de Estrogênio/metabolismo , Proteínas/química , Sistema y+ de Transporte de Aminoácidos/química , Sistema y+ de Transporte de Aminoácidos/genética , Animais , Neoplasias da Mama/metabolismo , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Cisteína Endopeptidases/metabolismo , Proteína p300 Associada a E1A/genética , Ativação Enzimática/genética , Estradiol/genética , Estradiol/metabolismo , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/genética , Feminino , Células HEK293 , Humanos , Células MCF-7 , Camundongos , Camundongos Nus , Coativador 1 de Receptor Nuclear/genética , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética , Proteínas/metabolismo , Tamoxifeno/farmacologia , Ativação Transcricional , Ubiquitina/metabolismo , Enzimas Ativadoras de Ubiquitina/genética , Ubiquitina-Proteína Ligases/metabolismo
9.
Mol Cell ; 53(5): 791-805, 2014 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-24582500

RESUMO

The circadian clock is a self-sustaining oscillator that controls daily rhythms. For the proper circadian gene expression, dynamic changes in chromatin structure are important. Although chromatin modifiers have been shown to play a role in circadian gene expression, the in vivo role of circadian signal-modulated chromatin modifiers at an organism level remains to be elucidated. Here, we provide evidence that the lysine-specific demethylase 1 (LSD1) is phosphorylated by protein kinase Cα (PKCα) in a circadian manner and the phosphorylated LSD1 forms a complex with CLOCK:BMAL1 to facilitate E-box-mediated transcriptional activation. Knockin mice bearing phosphorylation-defective Lsd1(SA/SA) alleles exhibited altered circadian rhythms in locomotor behavior with attenuation of rhythmic expression of core clock genes and impaired phase resetting of circadian clock. These data demonstrate that LSD1 is a key component of the molecular circadian oscillator, which plays a pivotal role in rhythmicity and phase resetting of the circadian clock.


Assuntos
Ritmo Circadiano , Regulação da Expressão Gênica , Oxirredutases N-Desmetilantes/metabolismo , Proteína Quinase C-alfa/metabolismo , Fatores de Transcrição ARNTL/metabolismo , Sequência de Aminoácidos , Animais , Comportamento Animal , Proteínas CLOCK/metabolismo , Imunoprecipitação da Cromatina , Histona Desmetilases , Luz , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência Molecular , Oscilometria , Oxirredutases N-Desmetilantes/genética , Fosforilação , Regiões Promotoras Genéticas , Homologia de Sequência de Aminoácidos , Núcleo Supraquiasmático/metabolismo , Fatores de Tempo
10.
Nucleic Acids Res ; 48(16): 9037-9052, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32735658

RESUMO

Epigenetic regulation is important for establishing lineage-specific gene expression during early development. Although signaling pathways have been well-studied for regulation of trophectoderm reprogramming, epigenetic regulation of trophectodermal genes with histone modification dynamics have been poorly understood. Here, we identify that plant homeodomain finger protein 6 (PHF6) is a key epigenetic regulator for activation of trophectodermal genes using RNA-sequencing and ChIP assays. PHF6 acts as an E3 ubiquitin ligase for ubiquitination of H2BK120 (H2BK120ub) via its extended plant homeodomain 1 (PHD1), while the extended PHD2 of PHF6 recognizes acetylation of H2BK12 (H2BK12Ac). Intriguingly, the recognition of H2BK12Ac by PHF6 is important for exerting its E3 ubiquitin ligase activity for H2BK120ub. Together, our data provide evidence that PHF6 is crucial for epigenetic regulation of trophectodermal gene expression by linking H2BK12Ac to H2BK120ub modification.


Assuntos
Cromatina/genética , Proteínas Repressoras/genética , Ubiquitina-Proteína Ligases/genética , Acetilação , Animais , Reprogramação Celular/genética , Histonas/genética , Proteínas de Homeodomínio/genética , Humanos , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Ligação Proteica/genética , Processamento de Proteína Pós-Traducional/genética , Ubiquitinação/genética
11.
Proc Natl Acad Sci U S A ; 116(42): 21140-21149, 2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31570593

RESUMO

Retinoic acid-related orphan receptor α (RORα) functions as a transcription factor for various biological processes, including circadian rhythm, cancer, and metabolism. Here, we generate intestinal epithelial cell (IEC)-specific RORα-deficient (RORαΔIEC) mice and find that RORα is crucial for maintaining intestinal homeostasis by attenuating nuclear factor κB (NF-κB) transcriptional activity. RORαΔIEC mice exhibit excessive intestinal inflammation and highly activated inflammatory responses in the dextran sulfate sodium (DSS) mouse colitis model. Transcriptome analysis reveals that deletion of RORα leads to up-regulation of NF-κB target genes in IECs. Chromatin immunoprecipitation analysis reveals corecruitment of RORα and histone deacetylase 3 (HDAC3) on NF-κB target promoters and subsequent dismissal of CREB binding protein (CBP) and bromodomain-containing protein 4 (BRD4) for transcriptional repression. Together, we demonstrate that RORα/HDAC3-mediated attenuation of NF-κB signaling controls the balance of inflammatory responses, and therapeutic strategies targeting this epigenetic regulation could be beneficial to the treatment of chronic inflammatory diseases, including inflammatory bowel disease (IBD).


Assuntos
Homeostase/fisiologia , Inflamação/metabolismo , Intestinos/fisiologia , Receptores Nucleares Órfãos/metabolismo , Animais , Epigênese Genética/fisiologia , Células Epiteliais/metabolismo , Células Epiteliais/fisiologia , Feminino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica/fisiologia , Transcriptoma/fisiologia
12.
J Biomed Sci ; 28(1): 41, 2021 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-34082769

RESUMO

Lysine-specific demethylase 1 (LSD1) targets mono- or di-methylated histone H3K4 and H3K9 as well as non-histone substrates and functions in the regulation of gene expression as a transcriptional repressor or activator. This enzyme plays a pivotal role in various physiological processes, including development, differentiation, inflammation, thermogenesis, neuronal and cerebral physiology, and the maintenance of stemness in stem cells. LSD1 also participates in pathological processes, including cancer as the most representative disease. It promotes oncogenesis by facilitating the survival of cancer cells and by generating a pro-cancer microenvironment. In this review, we discuss the role of LSD1 in several aspects of cancer, such as hypoxia, epithelial-to-mesenchymal transition, stemness versus differentiation of cancer stem cells, as well as anti-tumor immunity. Additionally, the current understanding of the involvement of LSD1 in various other pathological processes is discussed.


Assuntos
Histona Desmetilases/genética , Homeostase/genética , Neoplasias/genética , Animais , Diferenciação Celular/genética , Transição Epitelial-Mesenquimal/genética , Histona Desmetilases/imunologia , Histona Desmetilases/metabolismo , Homeostase/imunologia , Humanos , Hipóxia/enzimologia , Hipóxia/genética , Hipóxia/imunologia , Camundongos , Neoplasias/enzimologia , Neoplasias/imunologia , Células-Tronco Neoplásicas/fisiologia , Evasão Tumoral/genética
13.
Adv Exp Med Biol ; 1187: 103-119, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33983575

RESUMO

Aberrant epigenetic alteration has been associated with development of various cancers, including breast cancer. Since epigenetic modifications such as DNA methylation and histone modification are reversible, epigenetic enzymes, including histone modifying enzymes and DNA methyltransferases, emerge as attractive targets for cancer therapy. Although epi-drugs targeting histone deacetylation or DNA methylation have received FDA approval for cancer therapy, a very modest anti-tumor activity has been observed with monotherapy in clinical studies of breast cancer. To improve efficacy of epi-drugs in breast cancer, combination of epi-drugs with other therapies currently has been investigated. Additionally, basic researches to elucidate molecular causes of cancer should be extensively and intensively conducted in order to find novel epigenetic druggable targets. In this chapter, we summarize how epigenetic regulation affects the development of breast cancer and how to control cancer phenotype by modulating abnormal epigenetic modifications, and then suggest future research directions in epigenetics for breast cancer treatment.


Assuntos
Neoplasias da Mama , Epigênese Genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Metilação de DNA , Histonas/genética , Histonas/metabolismo , Humanos , Processamento de Proteína Pós-Traducional
14.
Proc Natl Acad Sci U S A ; 115(46): 11766-11771, 2018 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-30377265

RESUMO

Janus tyrosine kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) signaling pathway is essential for modulating cellular development, differentiation, and homeostasis. Thus, dysregulation of JAK2-STAT3 signaling pathway is frequently associated with human malignancies. Here, we provide evidence that lysine-specific demethylase 3A (KDM3A) functions as an essential epigenetic enzyme for the activation of JAK2-STAT3 signaling pathway. KDM3A is tyrosine-phosphorylated by JAK2 in the nucleus and functions as a STAT3-dependent transcriptional coactivator. JAK2-KDM3A signaling cascade induced by IL-6 leads to alteration of histone H3K9 methylation as a predominant epigenetic event, thereby providing the functional and mechanistic link between activation of JAK2-STAT3 signaling pathway and its epigenetic control. Together, our findings demonstrate that inhibition of KDM3A phosphorylation could be a potent therapeutic strategy to control oncogenic effect of JAK2-STAT3 signaling pathway.


Assuntos
Histona Desmetilases com o Domínio Jumonji/metabolismo , Epigênese Genética , Células HEK293/metabolismo , Células HeLa , Histonas/metabolismo , Humanos , Interleucina-6/metabolismo , Janus Quinase 2/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Fosforilação , Proteínas Tirosina Quinases/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Ativação Transcricional
15.
Biochem Biophys Res Commun ; 526(1): 176-183, 2020 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-32201075

RESUMO

Autophagy is an essential process to maintain cell survival and homeostasis under various stress conditions. Here, we report that lysine-specific demethylase 3A (KDM3A) plays an important role in starvation-induced autophagy. Using Kdm3a knockout mice, we demonstrate that KDM3A is crucial for proper hepatic autophagy in vivo. Hepatic mRNA expression analysis and ChIP assay in WT and Kdm3a knockout mouse livers reveal that KDM3A activates autophagy genes by reducing histone H3K9me2 levels upon fasting. Together, our finding represents previously unidentified function of KDM3A as a key regulator of autophagy, implicating potential therapeutic approaches for autophagy-related diseases.


Assuntos
Autofagia , Histona Desmetilases com o Domínio Jumonji/metabolismo , Animais , Autofagossomos/metabolismo , Jejum , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Células Hep G2 , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Fígado/citologia , Fígado/metabolismo , Lisossomos/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
16.
Mol Cell ; 46(3): 260-73, 2012 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-22516971

RESUMO

The Mis18 complex has been identified as a critical factor for the centromeric localization of a histone H3 variant, centromeric protein A (CENP-A), which is responsible for the specification of centromere identity in the chromosome. However, the functional role of Mis18 complex is largely unknown. Here, we generated Mis18α conditional knockout mice and found that Mis18α deficiency resulted in lethality at early embryonic stage with severe defects in chromosome segregation caused by mislocalization of CENP-A. Further, we demonstrate Mis18α's crucial role for epigenetic regulation of centromeric chromatin by reinforcing centromeric localization of DNMT3A/3B. Mis18α interacts with DNMT3A/3B, and this interaction is critical for maintaining DNA methylation and hence regulating epigenetic states of centromeric chromatin. Mis18α deficiency led to reduced DNA methylation, altered histone modifications, and uncontrolled noncoding transcripts in centromere region by decreased DNMT3A/3B enrichment. Together, our findings uncover the functional mechanism of Mis18α and its pivotal role in mammalian cell cycle.


Assuntos
Autoantígenos/metabolismo , Centrômero/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/fisiologia , Segregação de Cromossomos/genética , Epigênese Genética , Animais , Autoantígenos/análise , Sítios de Ligação , Centrômero/metabolismo , Proteína Centromérica A , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/análise , Proteínas Cromossômicas não Histona/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA , DNA Metiltransferase 3A , Células HeLa , Histonas/metabolismo , Humanos , Camundongos , Camundongos Knockout , Mapeamento de Interação de Proteínas
17.
Mol Cell ; 48(4): 572-86, 2012 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-23063525

RESUMO

Ubiquitination plays a major role in protein degradation. Although phosphorylation-dependent ubiquitination is well known for the regulation of protein stability, methylation-dependent ubiquitination machinery has not been characterized. Here, we provide evidence that methylation-dependent ubiquitination is carried out by damage-specific DNA binding protein 1 (DDB1)/cullin4 (CUL4) E3 ubiquitin ligase complex and a DDB1-CUL4-associated factor 1 (DCAF1) adaptor, which recognizes monomethylated substrates. Molecular modeling and binding affinity studies reveal that the putative chromo domain of DCAF1 directly recognizes monomethylated substrates, whereas critical binding pocket mutations of the DCAF1 chromo domain ablated the binding from the monomethylated substrates. Further, we discovered that enhancer of zeste homolog 2 (EZH2) methyltransferase has distinct substrate specificities for histone H3K27 and nonhistones exemplified by an orphan nuclear receptor, RORα. We propose that EZH2-DCAF1/DDB1/CUL4 represents a previously unrecognized methylation-dependent ubiquitination machinery specifically recognizing "methyl degron"; through this, nonhistone protein stability can be dynamically regulated in a methylation-dependent manner.


Assuntos
Proteínas de Transporte/metabolismo , Proteínas Culina/metabolismo , Proteínas de Ligação a DNA/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste , Humanos , Células MCF-7 , Metilação , Membro 1 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Proteínas Serina-Treonina Quinases , Especificidade por Substrato
18.
Mol Cell ; 42(3): 274-84, 2011 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-21549306

RESUMO

Signaling pathways involve cascades of protein phosphorylation and ultimately affect regulation of transcription in the nucleus. However, most of the kinases in these pathways have not been generally considered to directly modulate transcription thus far. Here, recent significant progress in the field elucidating direct modifications of histones and histone modifiers by upstream kinases is summarized, and future directions are discussed.


Assuntos
Núcleo Celular/metabolismo , Histonas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Animais , Núcleo Celular/genética , Regulação da Expressão Gênica , Humanos , Modelos Genéticos , Fosforilação , Processamento de Proteína Pós-Traducional
19.
Mol Cell ; 44(5): 797-810, 2011 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-22152482

RESUMO

A critical component of the DNA damage response is the p53 tumor suppressor, and aberrant p53 function leads to uncontrolled cell proliferation and malignancy. Several molecules have been shown to regulate p53 stability; however, genome-wide systemic approaches for determining the affected, specific downstream target genes have not been extensively studied. Here, we first identified an orphan nuclear receptor, RORα, as a direct target gene of p53, which contains functional p53 response elements. The functional consequences of DNA damage-induced RORα are to stabilize p53 and activate p53 transcription in a HAUSP/Usp7-dependent manner. Interestingly, microarray analysis revealed that RORα-mediated p53 stabilization leads to the activation of a subset of p53 target genes that are specifically involved in apoptosis. We further confirmed that RORα enhances p53-dependent, in vivo apoptotic function in the Drosophila model system. Together, we determined that RORα is a p53 regulator that exerts its role in increased apoptosis via p53.


Assuntos
Apoptose , Dano ao DNA , Membro 1 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 1 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Estabilidade Proteica , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/genética , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Regiões Promotoras Genéticas/genética , Elementos de Resposta/genética , Ubiquitina Tiolesterase/metabolismo , Ubiquitinação
20.
J Immunol ; 196(9): 3887-95, 2016 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-27016605

RESUMO

A balance between bone formation and bone resorption is critical for the maintenance of bone mass. In many pathological conditions, including chronic inflammation, uncontrolled activation of osteoclast differentiation often causes excessive bone resorption that results in osteoporosis. In this study, we identified the osteopenia phenotype of mice lacking Usp18 (also called Ubp43), which is a deISGylating enzyme and is known as a negative regulator of type I IFN signaling. The expression of Usp18 was induced in preosteoclasts upon receptor activator of NF-κB ligand (RANKL) treatment. In an in vitro osteoclast-differentiation assay, bone marrow macrophages from Usp18-deficient mice exhibited an enhanced differentiation to multinucleated cells, elevated activation of NFATc1, and an increased expression of osteoclast marker genes upon RANKL treatment. Furthermore, in vitro quantification of bone resorption revealed a great increase in osteoclastic activities in Usp18-deficient cells. Interestingly, proinflammatory cytokine genes, such as IP-10 (CXCL10), were highly expressed in Usp18-deficient bone marrow macrophages upon RANKL treatment compared with wild-type cells. In addition, serum cytokine levels, especially IP-10, were significantly high in Usp18-knockout mice. In sum, we suggest that, although type I IFN is known to restrict osteoclast differentiation, the exaggerated activation of the type I IFN response in Usp18-knockout mice causes an osteopenia phenotype in mice.


Assuntos
Macrófagos/fisiologia , Osteoclastos/fisiologia , Osteogênese , Osteoporose/imunologia , Ubiquitina Tiolesterase/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Quimiocina CXCL10/metabolismo , Interferon Tipo I/metabolismo , Camundongos , Camundongos da Linhagem 129 , Camundongos Knockout , Fatores de Transcrição NFATC/metabolismo , Osteogênese/genética , Osteogênese/imunologia , Ligante RANK/metabolismo , Ubiquitina Tiolesterase/genética
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