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1.
Int J Mol Med ; 53(5)2024 05.
Artigo em Inglês | MEDLINE | ID: mdl-38577935

RESUMO

Histone chaperones serve a pivotal role in maintaining human physiological processes. They interact with histones in a stable manner, ensuring the accurate and efficient execution of DNA replication, repair and transcription. Retinoblastoma binding protein (RBBP)4 and RBBP7 represent a crucial pair of histone chaperones, which not only govern the molecular behavior of histones H3 and H4, but also participate in the functions of several protein complexes, such as polycomb repressive complex 2 and nucleosome remodeling and deacetylase, thereby regulating the cell cycle, histone modifications, DNA damage and cell fate. A strong association has been indicated between RBBP4/7 and some major human diseases, such as cancer, age­related memory loss and infectious diseases. The present review assesses the molecular mechanisms of RBBP4/7 in regulating cellular biological processes, and focuses on the variations in RBBP4/7 expression and their potential mechanisms in various human diseases, thus providing new insights for their diagnosis and treatment.


Assuntos
Histonas , Fatores de Transcrição , Humanos , Ciclo Celular , Chaperonas de Histonas/genética , Chaperonas de Histonas/química , Chaperonas de Histonas/metabolismo , Histonas/genética , Histonas/metabolismo , Proteína 4 de Ligação ao Retinoblastoma/química , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma , Fatores de Transcrição/metabolismo
2.
J Transl Med ; 22(1): 170, 2024 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-38368381

RESUMO

BACKGROUND: Hepatocellular carcinoma (HCC) is characterized by aggressive progression and elevated mortality rates. This study aimed to investigate the regulatory effects of RBBP7 on HCC pathogenesis and the underlying mechanisms. METHODS: The expression and clinical feature of RBBP7 were evaluated using bioinformatics analysis and the assessment of clinical HCC samples. CCK8 and colony formation were employed to estimate cell proliferation function of RBBP7. Aerobic glycolysis levels of RBBP7 were evaluated by measuring ATP levels, lactic acid production, glucose uptake capacity, and the expression of relevant enzymes (PFKM, PKM2, and LDHA). The phosphorylation levels in PI3K/AKT signaling were measured by western blotting. The regulatory effect of transcription factors of specificity protein 1 (SP1) on RBBP7 mRNA expression was confirmed in dual-luciferase reporter assays and chromatin immunoprecipitation experiments. The proliferation- and glycolysis-associated proteins were assessed using immunofluorescence staining in vivo. RESULTS: We found that RBBP7 is expressed at high levels in HCC and predicts poor survival. Functional assays showed that RBBP7 promoted HCC proliferation and glycolysis. Mechanistically, it was demonstrated that RBBP7 activates the PI3K/AKT pathway, a crucial pathway in glycolysis, contributing to the progression of HCC. The outcomes of the dual-luciferase assay further confirmed that SP1 is capable of activating the promoter of RBBP7. CONCLUSIONS: RBBP7, which is up-regulated by SP1, promotes HCC cell proliferation and glycolysis through the PI3K/AKT pathway. The findings of this study suggest that RBBP7 is a potential biomarker for HCC.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/patologia , Luciferases/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo
3.
J Clin Invest ; 133(20)2023 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-37843278

RESUMO

Maturation arrest (MA) is a subtype of non-obstructive azoospermia, and male infertility is a known risk factor for testicular tumors. However, the genetic basis for many affected individuals remains unknown. Here, we identified a deleterious hemizygous variant of X-linked retinoblastoma-binding protein 7 (RBBP7) as a potential key cause of MA, which was also found to be associated with the development of Leydig cell tumors. This mutation resulted in premature protein translation termination, affecting the sixth WD40 domain of the RBBP7 and the interaction of the mutated RBBP7 with histone H4. Decreased BRCA1 and increased γH2AX were observed in the proband. In mouse spermatogonial and pachytene spermatocyte-derived cells, deprivation of rbbp7 led to cell cycle arrest and apoptosis. In Drosophila, knockdown of RBBP7/Caf1-55 in germ cells resulted in complete absence of germ cells and reduced testis size, whereas knockdown of RBBP7/Caf1-55 in cyst cells resulted in hyperproliferative testicular cells. Interestingly, male infertility caused by Caf1-55 deficiency was rescued by ectopic expression of wild-type human RBBP7 but not mutant variants, suggesting the importance of RBBP7 in spermatogenesis. Our study provides insights into the mechanisms underlying the co-occurrence of MA and testicular tumors and may pave the way for innovative genetic diagnostics of these 2 diseases.


Assuntos
Azoospermia , Infertilidade Masculina , Neoplasias Testiculares , Animais , Humanos , Masculino , Camundongos , Azoospermia/genética , Azoospermia/metabolismo , Azoospermia/patologia , Infertilidade Masculina/genética , Infertilidade Masculina/metabolismo , Infertilidade Masculina/patologia , Mutação , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Espermatogênese/genética , Neoplasias Testiculares/genética , Neoplasias Testiculares/metabolismo , Testículo/metabolismo
4.
Cancer Gene Ther ; 30(8): 1124-1133, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37225855

RESUMO

Colorectal cancer (CRC) poses one of the most serious threats to human health worldwide, and abnormally expressed c-Myc and p53 are deemed the pivotal driving forces of CRC progression. In this study, we discovered that the lncRNA FIT, which was downregulated in CRC clinical samples, was transcriptionally suppressed by c-Myc in vitro and promoted CRC cell apoptosis by inducing FAS expression. FAS is a p53 target gene, and we found that FIT formed a trimer with RBBP7 and p53 that facilitated p53 acetylation and p53-mediated FAS gene transcription. Moreover, FIT was capable of retarding CRC growth in a mouse xenograft model, and FIT expression was positively correlated with FAS expression in clinical samples. Thus, our study elucidates the role of the lncRNA FIT in human colorectal cancer growth and provides a potential target for anti-CRC drugs.


Assuntos
Neoplasias Colorretais , RNA Longo não Codificante , Humanos , Animais , Camundongos , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Acetilação , RNA Longo não Codificante/genética , Proliferação de Células/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Carcinogênese/genética , Regulação Neoplásica da Expressão Gênica , Linhagem Celular Tumoral , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo
5.
J Immunol Res ; 2022: 6934744, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35958876

RESUMO

BK virus-associated nephropathy (BKVN) remains a major infectious complication due to powerful immunosuppression in kidney transplant recipients, and its histologic appearance can mimic rejection, leading to diagnostic and treatment dilemmas thus molecular diagnostic methods would be beneficial. We collected gene expression profiles of 169 kidney biopsies taken from BKVN, rejection, and stable functioning allografts, based on single sample gene set enrichment analysis and random forest algorithm, and three hallmark activities associated with DNA damage and proliferation were found to be relatively specific in BKVN. Subsequently, weighted gene co-expression network analysis and support vector machines (SVM) algorithm identified RBBP7 as a robust and promising biomarker with high accuracy in both training and validation cohorts (AUC =0.938, 0.977, respectively). Besides, potential drugs for BKVN treatment such as mepacrine were discovered, which may contribute to targeted antiviral therapy and effective patient management rather than simply reducing the doses of immunosuppressive agents in clinical practice. RBBP7 (retinoblastoma binding protein 7) serves as component of serval complexes that regulate chromatin metabolism and functions in affecting DNA replication and controlling cell proliferation. In this research, upregulation of RBBP7 was found to be associated with the higher infiltration of CD8 naïve T, iTreg, and neutrophil cells and the lower amounts of Th1, central memory T, NKT, CD8 T, and dendritic cells. Moreover, the infiltration of Th1, Th17, and NKT cells was steadily different between BKVN and rejection allografts through immune cell assessment. In conclusion, we identified and verified RBBP7 as a molecular biomarker for BKVN diagnosis, which demonstrated great distinguishing ability with allograft rejection and would support clinical decision-making.


Assuntos
Vírus BK , Nefropatias , Transplante de Rim , Nefrite Intersticial , Infecções por Polyomavirus , Infecções Tumorais por Vírus , Vírus BK/genética , Rejeição de Enxerto/diagnóstico , Humanos , Nefropatias/diagnóstico , Nefropatias/etiologia , Transplante de Rim/efeitos adversos , Infecções por Polyomavirus/diagnóstico , Proteína 7 de Ligação ao Retinoblastoma , Infecções Tumorais por Vírus/diagnóstico
6.
Acta Biochim Biophys Sin (Shanghai) ; 54(2): 179-186, 2022 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-35538026

RESUMO

Hypoxia-induced epigenetic regulation calls for more effective therapeutic targets for esophageal cancer. We used GEPIA and UALCAN databases to screen survival-related and cancer stage-associated genes. Eca109 and KYSE450 esophageal cancer cell lines were cultured under normoxia, hypoxia, or CoCl-induced hypoxia conditions, which were further transfected with plasmids expressing RB binding protein 7 (RBBP7), hypoxia-inducible factor 1 (HIF1)-α, or RBBP7 shRNA. Colony formation and MTT assays were used to detect cell proliferation. Tumor sphere formation and stemness marker detection were applied to assess cell stemness. RT-PCR and western blot analysis were used to detect the relative mRNA level and protein expression, respectively. Luciferase assay was utilized to detect the direct interaction between HIF1α and RBBP7. Up-regulated RBBP7 was identified as one of the most prominent survival-related genes, which is negatively correlated with the overall survival (OS), disease recurrence-free survival (DFS), and tumor stages. Hypoxia-induced HIF1α up-regulates RBBP7 expression, which promotes esophagus cancer cell viability, proliferation, and stemness with increased cyclin-dependent kinase 4 (CDK4) expression. Luciferase reporter assay verified that HIF1α transcriptionally regulates the expression of RBBP7. We conclude that hypoxia induces high expression of RBBP7 which is at least partially mediated by HIF1α, up-regulates the expression of downstream CDK4, and thereby promotes tumor progression in esophageal cancer cells.


Assuntos
Quinase 4 Dependente de Ciclina , Neoplasias Esofágicas , Proteína 7 de Ligação ao Retinoblastoma , Hipóxia Celular , Linhagem Celular Tumoral , Proliferação de Células/genética , Quinase 4 Dependente de Ciclina/biossíntese , Quinase 4 Dependente de Ciclina/genética , Progressão da Doença , Epigênese Genética , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/biossíntese , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo
7.
Epigenetics ; 17(10): 1205-1218, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-34709113

RESUMO

Preimplantation development is critical for reproductive successes in mammals. Thus, it is important to understand how preimplantation embryogenesis is regulated. As a key event of preimplantation development, epigenetic reprogramming has been widely studied, yet how epigenetic complexes regulate preimplantation development remains largely unknown. Retinoblastoma binding protein 4 (RBBP4) and 7 (RBBP7) are integral components of epigenetic complexes including SIN3A, NuRD, and CoREST. Here, we demonstrate that double knockdown of Rbbp4 and 7, but not individually, causes embryonic lethality during the morula-to-blastocyst transition. Mechanistically, depletion of RBBP4 and 7 results in dysregulation of genes related to cell cycle, lineage development, and regulation of transcription, which is accompanied by cell cycle block, disrupted lineage specification and chromatin structure. Interestingly, RBBP4/7 depletion leads to a dramatic increase in H3.3 and H3K27ac abundance during morula-to-blastocyst transition. ChIP-seq analysis in early embryos and embryonic stem cells reveals enrichment of H3.3 at the promoter regions of RBBP4/7 target genes. In summary, our studies demonstrate the compensatory role of RBBP4/7 and reveal its potential mechanisms in preimplantation development.Summary sentence:RBBP4 and RBBP7 play a compensatory role in regulating cell proliferation, apoptosis, and histone H3.3 deposition during preimplantation development.


Assuntos
Histonas , Proteína 4 de Ligação ao Retinoblastoma , Animais , Blastocisto/metabolismo , Proliferação de Células , Cromatina/metabolismo , Metilação de DNA , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Histonas/genética , Histonas/metabolismo , Mamíferos/genética , Mamíferos/metabolismo , Camundongos , Proteína 4 de Ligação ao Retinoblastoma/química , Proteína 4 de Ligação ao Retinoblastoma/genética , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Fatores de Transcrição/genética
8.
Nucleic Acids Res ; 49(11): 6196-6212, 2021 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-34086947

RESUMO

Retinoblastoma-binding proteins 4 and 7 (RBBP4 and RBBP7) are two highly homologous human histone chaperones. They function in epigenetic regulation as subunits of multiple chromatin-related complexes and have been implicated in numerous cancers. Due to their overlapping functions, our understanding of RBBP4 and 7, particularly outside of Opisthokonts, has remained limited. Here, we report that in the ciliate protozoan Tetrahymena thermophila a single orthologue of human RBBP4 and 7 proteins, RebL1, physically interacts with histone H4 and functions in multiple epigenetic regulatory pathways. Functional proteomics identified conserved functional links for Tetrahymena RebL1 protein as well as human RBBP4 and 7. We found that putative subunits of multiple chromatin-related complexes including CAF1, Hat1, Rpd3, and MuvB, co-purified with RebL1 during Tetrahymena growth and conjugation. Iterative proteomics analyses revealed that the cell cycle regulatory MuvB-complex in Tetrahymena is composed of at least five subunits including evolutionarily conserved Lin54, Lin9 and RebL1 proteins. Genome-wide analyses indicated that RebL1 and Lin54 (Anqa1) bind within genic and intergenic regions. Moreover, Anqa1 targets primarily promoter regions suggesting a role for Tetrahymena MuvB in transcription regulation. RebL1 depletion inhibited cellular growth and reduced the expression levels of Anqa1 and Lin9. Consistent with observations in glioblastoma tumors, RebL1 depletion suppressed DNA repair protein Rad51 in Tetrahymena, thus underscoring the evolutionarily conserved functions of RBBP4/7 proteins. Our results suggest the essentiality of RebL1 functions in multiple epigenetic regulatory complexes in which it impacts transcription regulation and cellular viability.


Assuntos
Chaperonas de Histonas/metabolismo , Proteínas de Protozoários/metabolismo , Tetrahymena thermophila/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/metabolismo , Evolução Biológica , Sequência Conservada , DNA/metabolismo , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética , Expressão Gênica , Células HEK293 , Chaperonas de Histonas/química , Chaperonas de Histonas/fisiologia , Histonas/metabolismo , Humanos , Neoplasias/metabolismo , Neoplasias/mortalidade , Oncogenes , Proteínas de Protozoários/química , Proteínas de Protozoários/fisiologia , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Tetrahymena thermophila/genética , Tetrahymena thermophila/crescimento & desenvolvimento
9.
Acta Neuropathol ; 142(2): 279-294, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33978814

RESUMO

Evidence indicates that tau hyper-phosphorylation and subsequent neurofibrillary tangle formation contribute to the extensive neuronal death in Alzheimer's disease (AD) and related tauopathies. Recent work has identified that increased tau acetylation can promote tau phosphorylation. Tau acetylation occurs at lysine 280 resulting from increased expression of the lysine acetyltransferase p300. The exact upstream mechanisms mediating p300 expression remain elusive. Additional work highlights the role of the epigenome in tau pathogenesis, suggesting that dysregulation of epigenetic proteins may contribute to acetylation and hyper-phosphorylation of tau. Here, we identify and focus on the histone-binding subunit of the Nucleosome Remodeling and Deacetylase (NuRD) complex: Retinoblastoma-Binding Protein 7 (Rbbp7). Rbbp7 chaperones chromatin remodeling proteins to their nuclear histone substrates, including histone acetylases and deacetylases. Notably, Rbbp7 binds to p300, suggesting that it may play a role in modulating tau acetylation. We interrogated Rbbp7 in post-mortem brain tissue, cell lines and mouse models of AD. We found reduced Rbbp7 mRNA expression in AD cases, a significant negative correlation with CERAD (neuritic plaque density) and Braak Staging (pathogenic tau inclusions) and a significant positive correlation with post-mortem brain weight. We also found a neuron-specific downregulation of Rbbp7 mRNA in AD patients. Rbbp7 protein levels were significantly decreased in 3xTg-AD and PS19 mice compared to NonTg, but no decreases were found in APP/PS1 mice that lack tau pathology. In vitro, Rbbp7 overexpression rescued TauP301L-induced cytotoxicity in immortalized hippocampal cells and primary cortical neurons. In vivo, hippocampal Rbbp7 overexpression rescued neuronal death in the CA1 of PS19 mice. Mechanistically, we found that increased Rbbp7 reduced p300 levels, tau acetylation at lysine 280 and tau phosphorylation at AT8 and AT100 sites. Collectively, these data identify a novel role of Rbbp7, protecting against tau-related pathologies, and highlight its potential as a therapeutic target in AD and related tauopathies.


Assuntos
Acetilação , Neurônios/patologia , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Tauopatias/patologia , Proteínas tau/metabolismo , Doença de Alzheimer/patologia , Animais , Encéfalo/patologia , Modelos Animais de Doenças , Camundongos , Neurônios/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Proteína 7 de Ligação ao Retinoblastoma/genética
10.
Nat Chem Biol ; 16(11): 1199-1207, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32747809

RESUMO

Targeted protein degradation is a new therapeutic modality based on drugs that destabilize proteins by inducing their proximity to E3 ubiquitin ligases. Of particular interest are molecular glues that can degrade otherwise unligandable proteins by orchestrating direct interactions between target and ligase. However, their discovery has so far been serendipitous, thus hampering broad translational efforts. Here, we describe a scalable strategy toward glue degrader discovery that is based on chemical screening in hyponeddylated cells coupled to a multi-omics target deconvolution campaign. This approach led us to identify compounds that induce ubiquitination and degradation of cyclin K by prompting an interaction of CDK12-cyclin K with a CRL4B ligase complex. Notably, this interaction is independent of a dedicated substrate receptor, thus functionally segregating this mechanism from all described degraders. Collectively, our data outline a versatile and broadly applicable strategy to identify degraders with nonobvious mechanisms and thus empower future drug discovery efforts.


Assuntos
Acetamidas/química , Antibacterianos/farmacologia , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/metabolismo , Doxiciclina/farmacologia , Hidrazinas/química , Indóis/química , Proteólise/efeitos dos fármacos , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Regulação da Expressão Gênica , Humanos , Estrutura Molecular , Ligação Proteica , Conformação Proteica , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/efeitos dos fármacos
11.
J Reprod Dev ; 66(2): 125-133, 2020 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-31956172

RESUMO

Kisspeptin, encoded by Kiss1, is essential for reproduction in mammals. Kiss1 expression is regulated by estrogen via histone acetylation in the Kiss1 promotor region. Thus, elucidation of histone modification factor(s) involved in the regulation of Kiss1 expression is required to gain further understanding of the mechanisms of its control. The RNA-seq analysis of isolated kisspeptin neurons, obtained from the arcuate nucleus (ARC) of female rats, revealed that Rbbp7, encoding retinoblastoma binding protein 7 (RBBP7), a member of histone modification and chromatin remodeling complexes, is highly expressed in the ARC kisspeptin neurons. Thus, the present study aimed to investigate whether RBBP7 is involved in Kiss1 expression. Histological analysis using in situ hybridization (ISH) revealed that Rbbp7 expression was located in several hypothalamic nuclei, including the ARC and the anteroventral periventricular nucleus (AVPV), where kisspeptin neurons are located. Double ISH for Rbbp7 and Kiss1 showed that a majority of kisspeptin neurons (more than 85%) expressed Rbbp7 mRNA in both the ARC and the AVPV of female rats. Further, Rbbp7 mRNA knockdown significantly decreased in vitro expression of Kiss1 in a mouse immortalized kisspeptin neuronal cell line (mHypoA-55). Estrogen treatment significantly decreased and increased Kiss1 mRNA levels in the ARC and AVPV of ovariectomized female rats, respectively, but failed to affect Rbbp7 mRNA levels in both the nuclei. Taken together, these findings suggest that RBBP7 is involved in the upregulation of Kiss1 expression in kisspeptin neurons of rodents in an estrogen-independent manner.


Assuntos
Núcleo Arqueado do Hipotálamo/metabolismo , Hipotálamo Anterior/metabolismo , Kisspeptinas/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Regulação para Cima , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Linhagem Celular , Estradiol/farmacologia , Feminino , Hipotálamo Anterior/efeitos dos fármacos , Kisspeptinas/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Ratos , Proteína 7 de Ligação ao Retinoblastoma/genética
12.
Eur Rev Med Pharmacol Sci ; 24(1): 151-163, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31957828

RESUMO

OBJECTIVE: Esophageal cancer (EC) ranks as the sixth leading cause of cancer-related mortality worldwide. Circular RNAs (circRNAs) are involved in the pathogenesis of different cancers. However, the regulatory mechanism of circ_0006168 in EC progression is still unclear. MATERIALS AND METHODS: The expression of circ_0006168, microRNA (miR)-384, and retinoblastoma binding protein 7 (RBBP7) in tumors and cells was measured by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The stability of circ_0006168 was analyzed after RNase R treatment. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was conducted to evaluate cell viability. Transwell assay was applied to determine cell migration and invasion. Glucose consumption and lactate production were detected using glucose detection and lactic acid detection kits. The interaction between miR-384 and circ_0006168 or RBBP7 was certified by Dual-Luciferase reporter system. Protein expression of pyruvate kinase (PK), RBBP7, S6 ribosomal protein kinase (S6K), phosphorylated S6K (p-S6K), S6, phosphorylated S6 (p-S6) was analyzed by Western blot. RESULTS: Circ_0006168 and RBBP7 were over-expressed while miR-384 was low-expressed in EC tumors and cells. The repression of circ_0006168 attenuated cell proliferation, migration, invasion, and glycolysis in EC. Of note, circ_0006168 functioned as a sponge while RBBP7 acted as a target of miR-384 in EC. Rescue experiment revealed that miR-384 inhibitor abrogated circ_0006168 silencing-induced repression on cell proliferation, migration, and invasion in EC. Meanwhile, upregulation of RBBP7 restored the inhibition of miR-384 on EC cell progression. Moreover, circ_0006168 was able to improve RBBP7 level by interacting with miR-384. Also, circ_0006168 could activate S6K/S6 pathway by regulating RBBP7 expression. CONCLUSIONS: Abundance of circ_0006168 contributes to cell proliferation, migration, invasion, and glycolysis in EC by competitively sponging miR-384 to facilitate RBBP7 expression, representing prospective targets for EC therapy.


Assuntos
Neoplasias Esofágicas/metabolismo , MicroRNAs/metabolismo , RNA Circular/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Proteínas Quinases S6 Ribossômicas/metabolismo , Proteína S6 Ribossômica/metabolismo , Movimento Celular , Proliferação de Células , Células Cultivadas , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Humanos , MicroRNAs/genética , RNA Circular/genética , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína S6 Ribossômica/genética , Proteínas Quinases S6 Ribossômicas/genética
13.
Nat Commun ; 11(1): 24, 2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31911655

RESUMO

The spindle assembly checkpoint (SAC) prevents premature chromosome segregation by inactivating the anaphase promoting complex/cyclosome (APC/C) until all chromosomes are properly attached to mitotic spindles. Here we identify a role for Cullin-RING ubiquitin ligase complex 4 (CRL4), known for modulating DNA replication, as a crucial mitotic regulator that triggers the termination of the SAC and enables chromosome segregation. CRL4 is recruited to chromatin by the replication origin binding protein RepID/DCAF14/PHIP. During mitosis, CRL4 dissociates from RepID and replaces it with RB Binding Protein 7 (RBBP7), which ubiquitinates the SAC mediator BUB3 to enable mitotic exit. During interphase, BUB3 is protected from CRL4-mediated degradation by associating with promyelocytic leukemia (PML) nuclear bodies, ensuring its availability upon mitotic onset. Deficiencies in RepID, CRL4 or RBBP7 delay mitotic exit, increase genomic instability and enhance sensitivity to paclitaxel, a microtubule stabilizer and anti-tumor drug.


Assuntos
Anáfase , Proteínas de Ciclo Celular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Metáfase , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteínas de Ciclo Celular/genética , Linhagem Celular , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Mitose , Proteínas de Ligação a Poli-ADP-Ribose/genética , Proteína da Leucemia Promielocítica/genética , Proteína da Leucemia Promielocítica/metabolismo , Ligação Proteica , Proteólise , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Fuso Acromático/metabolismo , Ubiquitina-Proteína Ligases/genética
14.
J Cell Biochem ; 120(4): 6370-6383, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30390344

RESUMO

INTRODUCTION: Currently, studies have shown that microRNA-93 (miR-93) can be an oncogene or a tumor suppressor in different kinds of cancers. The role of miR-93 in human cancers is inconsistent and the underlying mechanism on the aberrant expression of miR-93 is complicated. METHODS: We first conducted gene enrichment analysis to give insight into the prospective mechanism of miR-93. Second, we performed a meta-analysis to evaluate the clinical value of miR-93. Finally, a validation test based on quantitative polymerase chain reaction (qPCR) was performed to further investigate the role of miR-93 in pan-cancer. RESULTS: Gene Ontology (GO) enrichment analysis results showed that the target genes of miR-93 were closely related to transcription, and MAPK1, RBBP7 and Smad7 became the hub genes. In the diagnostic meta-analysis, the overall sensitivity, specificity, and area under the curve were 0.76 (0.64-0.85), 0.82 (0.64-0.92), and 0.85 (0.82-0.88), respectively, which suggested that miR-93 had excellent performance on the diagnosis for human cancers. In the prognostic meta-analysis, dysregulated miR-93 was found to be associated with poor OS in cancer patients. In the qPCR validation test, the serum levels of miR-93 were upregulated in breast cancer, breast hyperplasia, lung cancer, chronic obstructive pulmonary disease, nasopharyngeal cancer, hepatocellular cancer, gastric ulcer, endometrial cancer, esophageal cancer, laryngeal cancer, and prostate cancer compared with healthy controls. CONCLUSIONS: miR-93 could act as an effective diagnostic and prognostic factor for cancer patients. Its clinical value for cancer early diagnosis and survival prediction is promising.


Assuntos
Biologia Computacional/métodos , Redes Reguladoras de Genes , MicroRNAs/genética , Neoplasias/diagnóstico , Área Sob a Curva , Biomarcadores Tumorais/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Proteína Quinase 1 Ativada por Mitógeno/genética , Neoplasias/genética , Neoplasias/mortalidade , Prognóstico , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína Smad7/genética , Análise de Sobrevida
15.
Biochem Biophys Res Commun ; 501(2): 440-447, 2018 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-29738775

RESUMO

The E3 ubiquitin ligase HUWE1/Mule/ARF-BP1 plays an important role in diverse biological processes including DNA damage repair and apoptosis. Our previous study has shown that in response to DNA damage HUWE1 was downregulated in CUL4B-mediated ubiquitination and subsequent proteasomal degradation, and CUL4B-mediated regulation of HUWE1 was important for cell survival upon DNA damage. CUL4B is a core component of the CUL4B Ring ligase complexes containing ROC1, DDB1 and a DDB1-Cullin Associated Factors (DCAFs), the latter of which are DDB1-binding WD40 adaptors critical for substrate recognition and recruitment. However, the identity of DCAF in CRL4B that mediates degradation of HUWE1 remains elusive. Here we report that RBBP7 is the DCAF in the CRL4B complex bridging the DDB1-CUL4B-ROC1 to HUWE1. Loading of HUWE1 to the E3 ubiquitin ligase complex resulted in its polyubiquitination, and consequently its proteasome mediated degradation. Overexpression of RBBP7 promoted HUWE1 protein degradation, while depletion of RBBP7 stabilized HUWE1, and hence accelerated the degradation of MCL-1 and BRCA1, two substrates of HUWE1 that are critical in apoptosis and DNA damage repair. Taken together, these data reveal CRL4BRBBP7 is the E3 ligase responsible for the proteasomal degradation of HUWE1, and further provide a potential strategy for cancer therapy by targeting HUWE1 and the CUL4B E3 ligase complex.


Assuntos
Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteína BRCA1/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas Culina/genética , Proteínas Culina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células HEK293 , Células HeLa , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Estabilidade Proteica , Proteólise , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
16.
Mol Oncol ; 12(4): 476-494, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29377600

RESUMO

Cullin 4B, a member of the Cullins, which serve as scaffolds to facilitate the assembly of E3 ligase complexes, is aberrantly expressed in many cancers, including osteosarcoma. Recently, we observed that CUL4B forms the CRL4BDCAF11 E3 ligase, which specifically ubiquitinates and degrades the cyclin-dependent kinase (CDK) inhibitor p21Cip1 in human osteosarcoma cells. However, the underlying mechanisms regarding the aberrant expression of CUL4B and the upstream members of this signaling pathway are mostly unknown. In this study, we demonstrate that nuclear factor kappaB (NF-κB) is a direct modulator of CUL4B expression. The CUL4B promoter is responsive to several NF-κB subunits, including RelA, RelB, and c-Rel, but not to p50 or p52. Additional studies reveal that the tumor necrosis factor alpha (TNF-α)/NF-κB axis pathway is activated in human osteosarcoma cells. This activation causes both CUL4B and NF-κB subunits to become abundant in the nucleus of human osteosarcoma cells. The down-regulation of individual genes, including TNFR1, RelA, RelB, c-Rel, and CUL4B, or pairs of them, including TNFR1 + RelA, TNFR1 + RelB, TNFR1 + c-Rel, and RelA+CUL4B, has similar effects on cell growth inhibition, colony formation, cell invasion, and in vivo tumor formation, whereas the overexpression of CUL4B in these knockdown cells significantly reverses their phenotypes. The inhibition of the TNF-α/NF-κB pathway greatly attenuates CRL4BDCAF11 E3 ligase activity and causes the accumulation of p21Cip1 , thereby leading to cell cycle arrest at the S phase. Taken together, our results support a model in which the activation of the TNF-α/NF-κB axis contributes to an increase in CRL4BDCAF11 activity and a decrease in p21Cip1 protein levels, thereby controlling cell cycle progression in human osteosarcoma cells.


Assuntos
Neoplasias Ósseas/metabolismo , Proteínas de Transporte/metabolismo , Ciclo Celular , Modelos Biológicos , NF-kappa B/metabolismo , Proteínas de Neoplasias/metabolismo , Osteossarcoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , Animais , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Feminino , Humanos , Masculino , Camundongos , Camundongos Nus , NF-kappa B/genética , Proteínas de Neoplasias/genética , Osteossarcoma/genética , Osteossarcoma/patologia , Proteína 7 de Ligação ao Retinoblastoma/genética , Fator de Necrose Tumoral alfa/genética , Complexos Ubiquitina-Proteína Ligase
17.
J Biol Chem ; 293(6): 2125-2136, 2018 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-29263092

RESUMO

The transcription factor BCL11A has recently been reported to be a driving force in triple-negative breast cancer (TNBC), contributing to the maintenance of a chemoresistant breast cancer stem cell (BCSC) population. Although BCL11A was shown to suppress γ-globin and p21 and to induce MDM2 expression in the hematopoietic system, its downstream targets in TNBC are still unclear. For its role in transcriptional repression, BCL11A was found to interact with several corepressor complexes; however, the mechanisms underlying these interactions remain unknown. Here, we reveal that BCL11A interacts with histone methyltransferase (PRC2) and histone deacetylase (NuRD and SIN3A) complexes through their common subunit, RBBP4/7. In fluorescence polarization assays, we show that BCL11A competes with histone H3 for binding to the negatively charged top face of RBBP4. To define that interaction, we solved the crystal structure of RBBP4 in complex with an N-terminal peptide of BCL11A (residues 2-16, BCL11A(2-16)). The crystal structure identifies novel interactions between BCL11A and the side of the ß-propeller of RBBP4 that are not seen with histone H3. We next show that BCL11A(2-16) pulls down RBBP4, RBBP7, and other components of PRC2, NuRD, and SIN3A from the cell lysate of the TNBC cell line SUM149. Furthermore, we demonstrate the therapeutic potential of targeting the RBBP4-BCL11A binding by showing that a BCL11A peptide can decrease aldehyde dehydrogenase-positive BCSCs and mammosphere formation capacity in SUM149. Together, our findings have uncovered a previously unidentified mechanism that BCL11A may use to recruit epigenetic complexes to regulate transcription and promote tumorigenesis.


Assuntos
Proteínas de Transporte/metabolismo , Proteínas Nucleares/metabolismo , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Carcinogênese , Proteínas de Transporte/química , Linhagem Celular , Cristalografia por Raios X , Epigenômica , Histona Desacetilases/metabolismo , Histona Metiltransferases/metabolismo , Humanos , Proteínas Nucleares/química , Ligação Proteica , Proteínas Repressoras , Proteína 4 de Ligação ao Retinoblastoma/química , Proteína 7 de Ligação ao Retinoblastoma/química , Fatores de Transcrição/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
18.
Sci Signal ; 10(464)2017 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-28143904

RESUMO

Adenosine monophosphate (AMP)-activated protein kinase (AMPK) acts as a master regulator of cellular energy homeostasis by directly phosphorylating metabolic enzymes and nutrient transporters and by indirectly promoting the transactivation of nuclear genes involved in mitochondrial biogenesis and function. We explored the mechanism of AMPK-mediated induction of gene expression. We identified AMPK consensus phosphorylation sequences in three proteins involved in nucleosome remodeling: DNA methyltransferase 1 (DNMT1), retinoblastoma binding protein 7 (RBBP7), and histone acetyltransferase 1 (HAT1). DNMT1 mediates DNA methylation that limits transcription factor access to promoters and is inhibited by RBBP7. Acetylation of histones by HAT1 creates a more relaxed chromatin-DNA structure that favors transcription. AMPK-mediated phosphorylation resulted in the activation of HAT1 and inhibition of DNMT1. For DNMT1, this inhibition was both a direct effect of phosphorylation and the result of increased interaction with RBBP7. In human umbilical vein cells, pharmacological AMPK activation or pulsatile shear stress triggered nucleosome remodeling and decreased cytosine methylation, leading to increased expression of nuclear genes encoding factors involved in mitochondrial biogenesis and function, such as peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α), transcription factor A (Tfam), and uncoupling proteins 2 and 3 (UCP2 and UCP3). Similar effects were seen in the aortas of mice given pharmacological AMPK activators, and these effects required AMPK2α. These results enhance our understanding of AMPK-mediated mitochondrial gene expression through nucleosome remodeling.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Histona Acetiltransferases/metabolismo , Biogênese de Organelas , Proteína 7 de Ligação ao Retinoblastoma/metabolismo , Células Cultivadas , Montagem e Desmontagem da Cromatina/genética , DNA (Citosina-5-)-Metiltransferase 1/genética , Metilação de DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Histona Acetiltransferases/genética , Humanos , Immunoblotting , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Nucleossomos/genética , Nucleossomos/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Ligação Proteica , Proteína 7 de Ligação ao Retinoblastoma/genética , Espectrometria de Massas em Tandem , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Desacopladora 2/genética , Proteína Desacopladora 2/metabolismo , Proteína Desacopladora 3/genética , Proteína Desacopladora 3/metabolismo
19.
Biochim Biophys Acta Proteins Proteom ; 1865(5): 531-538, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28179136

RESUMO

The repressive Nucleosome Remodeling and histone Deacetylation (NuRD) complex remodels the chromatin structure by coupling ATP-dependent remodeling activity with histone deacetylase function and plays important roles in regulating gene transcription, DNA damage repair and chromatin assembly. The complex is composed of six subunits: Metastasis Associated proteins MTA1/2/3 initially recruit histone chaperones RBBP4/7 followed by the histone deacetylases HDAC1/2 forming a core complex. Further association of the CpG-binding protein MBD2/3, p66α/ß and the ATP-dependent helicase CDH3/4 constitutes the NuRD complex. Recent structural studies on truncated human proteins or orthologous have revealed that the stoichiometry of the MTA1-RBBP4 complex is 2:4. This study reports expression and purification of the intact human MTA2-RBBP7 complex using HEK293F cells as expression system. In analogy with findings on the Drosophila NuRD complex, we find that also the human MTA-RBBP can be isolated in vitro. Taken together with previous findings this suggests, that MTA-RBBP is a stable complex, with a central role in the initial assembly of the human NuRD complex. Refined 3D volumes of the complex generated from negative stain electron microscopy (EM) data reveals an elongated architecture that is capable of hinge like motion around the center of the particle.


Assuntos
Montagem e Desmontagem da Cromatina/genética , Histona Desacetilases/química , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/química , Proteínas Repressoras/química , Proteína 7 de Ligação ao Retinoblastoma/química , Sequência de Aminoácidos/genética , Regulação da Expressão Gênica , Células HEK293 , Chaperonas de Histonas/química , Chaperonas de Histonas/isolamento & purificação , Chaperonas de Histonas/metabolismo , Histona Desacetilase 1/química , Histona Desacetilase 1/genética , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/química , Histona Desacetilase 2/genética , Histona Desacetilase 2/metabolismo , Histona Desacetilases/genética , Histona Desacetilases/isolamento & purificação , Humanos , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/genética , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/isolamento & purificação , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/isolamento & purificação
20.
Nat Commun ; 7: 13465, 2016 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-27811920

RESUMO

Centromeres are specified epigenetically through the deposition of the centromere-specific histone H3 variant CENP-A. However, how additional epigenetic features are involved in centromere specification is unknown. Here, we find that histone H4 Lys5 and Lys12 acetylation (H4K5ac and H4K12ac) primarily occur within the pre-nucleosomal CENP-A-H4-HJURP (CENP-A chaperone) complex, before centromere deposition. We show that H4K5ac and H4K12ac are mediated by the RbAp46/48-Hat1 complex and that RbAp48-deficient DT40 cells fail to recruit HJURP to centromeres and do not incorporate new CENP-A at centromeres. However, C-terminally-truncated HJURP, that does not bind CENP-A, does localize to centromeres in RbAp48-deficient cells. Acetylation-dead H4 mutations cause mis-localization of the CENP-A-H4 complex to non-centromeric chromatin. Crucially, CENP-A with acetylation-mimetic H4 was assembled specifically into centromeres even in RbAp48-deficient DT40 cells. We conclude that H4K5ac and H4K12ac, mediated by RbAp46/48, facilitates efficient CENP-A deposition into centromeres.


Assuntos
Proteína Centromérica A/metabolismo , Centrômero/metabolismo , Histonas/metabolismo , Chaperonas Moleculares/metabolismo , Nucleossomos/metabolismo , Acetilação , Animais , Linhagem Celular Tumoral , Centrômero/genética , Proteína Centromérica A/genética , Galinhas , Cromatina/metabolismo , Epigênese Genética , Histonas/genética , Humanos , Lisina/metabolismo , Chaperonas Moleculares/genética , Mutação , Nucleossomos/genética , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/metabolismo
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