RESUMEN
ABSTRACT: Yin Yang 1 (YY1) and structural maintenance of chromosomes 3 (SMC3) are 2 critical chromatin structural factors that mediate long-distance enhancer-promoter interactions and promote developmentally regulated changes in chromatin architecture in hematopoietic stem/progenitor cells (HSPCs). Although YY1 has critical functions in promoting hematopoietic stem cell (HSC) self-renewal and maintaining HSC quiescence, SMC3 is required for proper myeloid lineage differentiation. However, many questions remain unanswered regarding how YY1 and SMC3 interact with each other and affect hematopoiesis. We found that YY1 physically interacts with SMC3 and cooccupies with SMC3 at a large cohort of promoters genome wide, and YY1 deficiency deregulates the genetic network governing cell metabolism. YY1 occupies the Smc3 promoter and represses SMC3 expression in HSPCs. Although deletion of 1 Smc3 allele partially restores HSC numbers and quiescence in YY1 knockout mice, Yy1-/-Smc3+/- HSCs fail to reconstitute blood after bone marrow transplant. YY1 regulates HSC metabolic pathways and maintains proper intracellular reactive oxygen species levels in HSCs, and this regulation is independent of the YY1-SMC3 axis. Our results establish a distinct YY1-SMC3 axis and its impact on HSC quiescence and metabolism.
Asunto(s)
Proteínas de Ciclo Celular , Proteínas Cromosómicas no Histona , Células Madre Hematopoyéticas , Factor de Transcripción YY1 , Animales , Ratones , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/genética , Cohesinas , Regulación de la Expresión Génica , Hematopoyesis , Células Madre Hematopoyéticas/metabolismo , Células Madre Hematopoyéticas/citología , Ratones Noqueados , Regiones Promotoras Genéticas , Factor de Transcripción YY1/metabolismo , Factor de Transcripción YY1/genéticaRESUMEN
Ultra-conserved RNA (ucRNA) is a subset of long non-coding RNA, that is highly conserved among mice, rats and humans. UcRNA has attracted extensive attention in recent years for its potential biological significance in normal physiological function and diseases. However, due to the instability of RNA and the technical limitation, the function and mechanism of ucRNAs are largely unknown. Over the last two decades, researchers have made a lot of efforts to try to lift the veil of ucRNA in nervous, cardiovascular system and other systems as well as cancers. Since the concept of the glymphatic system is relatively new, we summarized here recent findings on the functions, regulation and the underlying mechanisms of ucRNAs in physiology and pathology. Meanwhile, pathology in some diseases is likely to contribute to abnormal expression of ucRNA in turn. We also discuss the technical challenges and bright prospects for future applications of ucRNAs in the diagnosis and treatment of diseases.
RESUMEN
Yin Yang 1 (YY1) is a ubiquitous transcription factor and mammalian Polycomb Group protein (PcG) with important functions for regulating lymphocyte development and stem cell self-renewal. YY1 mediates stable PcG-dependent transcriptional repression via recruitment of PcG proteins that result in histone modifications. Many questions remain unanswered regarding how cell- and tissue-specificity is achieved by PcG proteins. Here, we demonstrate that a conditional knockout of Yy1 in the hematopoietic system results in an early T cell developmental blockage at the double negative (DN) 1 stage with reduced Notch1 signaling. There is a lineage-specific requirement for YY1 PcG function. YY1 PcG domain is required for T and B cell development but not necessary for myeloid cells. YY1 functions in early T cell development are multicomponent and involve both PcG-dependent and -independent regulations. Although YY1 promotes early T cell survival through its PcG function, its function to promote the DN1-to-DN2 transition and Notch1 expression and signaling is independent of its PcG function. Our results reveal how a ubiquitously expressed PcG protein mediates lineage-specific and context-specific functions to control early T cell development.
Asunto(s)
Diferenciación Celular/fisiología , Proteínas del Grupo Polycomb/genética , Proteínas del Grupo Polycomb/metabolismo , Linfocitos T/metabolismo , Factor de Transcripción YY1/genética , Factor de Transcripción YY1/metabolismo , Animales , Supervivencia Celular , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Noqueados , Receptor Notch1 , TranscriptomaRESUMEN
MLN4924 (pevonedistat) is a potent and selective NEDD8-activating enzyme (NAE) inhibitor. The NEDD8-regulated neddylation system is responsible for the regulated degradation of intracellular proteins with important cellular functions in cancer cell growth, apoptosis, angiogenesis and metastasis. In human melanoma, inhibition of NAE results in induction of DNA re-replication, S phase cell cycle arrest, DNA damage and apoptosis. The study aimed to assess the anti-cancer effect of MLN4924 on canine malignant melanoma cell lines and patient samples and to elucidate the underlying mechanisms. Canine melanoma cell lines and primary patient samples were evaluated for cell viability after incubation with varying concentrations of MLN4924 or dimethyl sulfoxide. Apoptosis, cell proliferation and senescence assays were performed to address underlying mechanisms of MLN4924-mediated anti-tumour effects. Gene expression of seven previously identified deregulated genes in human melanoma was compared in sensitive vs resistant samples. MLN4924 treatment significantly reduced the viability of canine melanoma cell lines and primary samples in a dose- and time-dependent manners. MLN4924 promoted cell apoptosis and inhibited cell growth through induction of DNA re-replication and cell senescence. While the majority of canine melanoma samples demonstrated sensitivity at nanomolar ranges, some samples were resistant to the treatment. Modulation of P21 levels correlated with canine melanoma cell sensitivity. These results provided justification for further exploration of MLN4924 as a treatment of canine melanoma.
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Ciclopentanos/farmacología , Enfermedades de los Perros/tratamiento farmacológico , Inhibidores Enzimáticos/farmacología , Melanoma/veterinaria , Neoplasias de la Boca/veterinaria , Pirimidinas/farmacología , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Replicación del ADN/efectos de los fármacos , Perros , Melanoma/tratamiento farmacológico , Melanoma/patología , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/patologíaRESUMEN
Yin yang 1 (YY1) is a ubiquitous transcription factor and mammalian polycomb group protein (PcG) with important functions to regulate embryonic development, lineage differentiation, and cell proliferation. YY1 mediates stable PcG-dependent transcriptional repression via recruitment of PcG proteins that catalyze histone modifications. Many questions remain unanswered regarding how cell- and tissue-specificity is achieved by PcG proteins. Here, we demonstrate that a conditional knockout of Yy1 in hematopoietic stem cells (HSCs) decreases long-term repopulating activity and ectopic YY1 expression expands HSCs. Although the YY1 PcG domain is required for Igκ chain rearrangement in B cells, the YY1 mutant lacking the PcG domain retained the capacity to stimulate HSC self-renewal. YY1 deficiency deregulated the genetic network governing HSC cell proliferation and impaired stem cell factor/c-Kit signaling, disrupting mechanisms conferring HSC quiescence. These results reveal a mechanism for how a ubiquitously expressed transcriptional repressor mediates lineage-specific functions to control adult hematopoiesis.
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Autorrenovación de las Células/fisiología , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/metabolismo , Factor de Transcripción YY1/metabolismo , Animales , Técnicas de Inactivación de Genes , RatonesRESUMEN
Highly up-regulated in liver cancer (HULC) is a long non-coding RNA (lncRNA). We found that HULC up-regulated sphingosine kinase 1 (SPHK1), which is involved in tumor angiogenesis. Levels of HULC were positively correlated with levels of SPHK1 and its product, sphingosine-1-phosphate (S1P), in patients HCC samples. HULC increased SPHK1 in hepatoma cells. Chicken chorioallantoic membrane (CAM) assays revealed that si-SPHK1 remarkably blocked the HULC-enhanced angiogenesis. Mechanistically, HULC activated the promoter of SPHK1 in hepatoma cells through the transcription factor E2F1. Chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA) further showed that E2F1 was capable of binding to the E2F1 element in the SPHK1 promoter. HULC increased the expression of E2F1 in hepatoma cells and levels of HULC were positively correlated with those of E2F1 in HCC tissues. Intriguingly, HULC sequestered miR-107, which targeted E2F1 mRNA 3'UTR, by complementary base pairing. Functionally, si-SPHK1 remarkably abolished the HULC-enhanced tumor angiogenesis in vitro and in vivo. Taken together, we conclude that HULC promotes tumor angiogenesis in liver cancer through miR-107/E2F1/SPHK1 signaling. Our finding provides new insights into the mechanism of tumor angiogenesis.
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Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/genética , Neovascularización Patológica/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , ARN Largo no Codificante/genética , Regiones no Traducidas 3'/genética , Adulto , Anciano , Animales , Western Blotting , Embrión de Pollo , Factor de Transcripción E2F1/genética , Factor de Transcripción E2F1/metabolismo , Femenino , Células Hep G2 , Humanos , Neoplasias Hepáticas/irrigación sanguínea , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , MicroARNs/genética , Persona de Mediana Edad , Neovascularización Fisiológica/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Trasplante Heterólogo , Regulación hacia Arriba/genéticaRESUMEN
MicroRNAs acting as oncogenes or tumor suppressor genes play crucial roles in human cancers. Sphingosine kinase 1 (SPHK1) and its metabolite sphingosine 1-phosphate (S1P) contribute to tumor angiogenesis. We have reported that the down-regulation of miR-506 targeting YAP mRNA results in the hepatocarcinogenesis. In the present study, we report a novel function of miR-506, which suppresses tumor angiogenesis through targeting SPHK1 mRNA in liver cancer. Bioinformatics analysis showed that miR-506 might target 3'-untranslated region (3'UTR) of SPHK1 mRNA. Then, we validated that by luciferase reporter gene assays. MiR-506 was able to reduce the expression of SPHK1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blot analysis in hepatoma HepG2 cells. Functionally, human umbilical vein endothelial cell (HUVEC) tube formation assays demonstrated that the forced miR-506 expression remarkably inhibited the production of S1P in the supernatant of hepatoma cells. The supernatant resulted in the inhibition of tumor angiogenesis. Interestingly, the supernatant with overexpression of SPHK1 could rescue the inhibition of angiogenesis of liver cancer mediated by miR-506. Anti-miR-506 increased the production of S1P in the supernatant of hepatoma cells, but the supernatant with silencing of SPHK1 abolished anti-miR-506-induced acceleration of tumor angiogenesis. Clinically, we observed that the levels of miR-506 were negatively related to those of SPHK1 mRNA in liver cancer tissues. Thus, we conclude that miR-506 depresses the angiogenesis of liver cancer through targeting 3'UTR of SPHK1 mRNA. Our finding provides new insights into the mechanism of tumor angiogenesis.
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Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Hígado/patología , MicroARNs/genética , Neovascularización Patológica/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Regiones no Traducidas 3' , Secuencia de Bases , Carcinoma Hepatocelular/irrigación sanguínea , Carcinoma Hepatocelular/patología , Regulación Neoplásica de la Expresión Génica , Células Hep G2 , Células Endoteliales de la Vena Umbilical Humana , Humanos , Hígado/irrigación sanguínea , Hígado/metabolismo , Neoplasias Hepáticas/irrigación sanguínea , Neoplasias Hepáticas/patología , Neovascularización Patológica/patología , ARN Mensajero/genéticaRESUMEN
AIM: Sphingosine kinase 1 (SPHK1) is involved in various cellular functions, including cell growth, migration, apoptosis, cytoskeleton architecture and calcium homoeostasis, etc. As an oncogenic kinase, SPHK1 is associated with the development and progression of cancers. The aim of this study was to investigate whether SPHK1 was involved in hepatocarcinogenesis induced by the hepatitis B virus X protein (HBx). METHODS: The expression of SPHK1 in hepatocellular carcinoma (HCC) tissue and hepatoma cells were measured using qRT-PCR and Western blot analysis. HBx expression levels in hepatoma cells were modulated by transiently transfected with HBx or psi-HBx plasmids. The SPHK1 promoter activity was measured using luciferase reporter gene assay, and the interaction of the transcription factor AP2α with the SPHK1 promoter was studied with chromatin immunoprecipitation assay. The growth of hepatoma cells was evaluated in vitro using MTT and colony formation assays, and in a tumor xenograft model. RESULTS: A positive correlation was found between the mRNA levels of SPHK1 and HBx in 38 clinical HCC samples (r=+0.727, P<0.01). Moreover, the expression of SPHK1 was markedly increased in the liver cancer tissue of HBx-transgenic mice. Overexpressing HBx in normal liver cells LO2 and hepatoma cells HepG2 dose-dependently increased the expression of SPHK1, whereas silencing HBx in HBx-expressing hepatoma cells HepG2-X and HepG2.2.15 suppressed SPHK1 expression. Furthermore, overexpressing HBx in HepG2 cells dose-dependently increased the SPHK1 promoter activity, whereas silencing HBx in HepG2-X cells suppressed this activity. In HepG2-X cells, AP2α was found to directly interact with the SPHK1 promoter, and silencing AP2α suppressed the SPHK1 promoter activity and SPHK1 expression. Silencing HBx in HepG2-X cells abolished the HBx-enhanced proliferation and colony formation in vitro, and tumor growth in vivo. CONCLUSION: HBx upregulates SPHK1 through the transcription factor AP2α, which promotes the growth of human hepatoma cells.
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Carcinoma Hepatocelular/virología , Virus de la Hepatitis B/fisiología , Hepatitis B/complicaciones , Neoplasias Hepáticas/virología , Hígado/virología , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Transactivadores/genética , Factor de Transcripción AP-2/genética , Animales , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Regulación Viral de la Expresión Génica , Células Hep G2 , Hepatitis B/genética , Hepatitis B/patología , Hepatitis B/virología , Virus de la Hepatitis B/genética , Humanos , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Ratones , Ratones Desnudos , Ratones Transgénicos , Regiones Promotoras Genéticas , ARN Mensajero/genética , Regulación hacia Arriba , Proteínas Reguladoras y Accesorias ViralesRESUMEN
Accumulating evidence indicates that microRNAs are able to act as oncogenes or tumor suppressor genes in human cancer. We previously reported that miR-520b was down-regulated in hepatocellular carcinoma (HCC) and its deregulation was involved in hepatocarcinogenesis. In the present study, we report that miR-520b suppresses cell proliferation in HCC through targeting the ten-eleven translocation 1 (TET1) mRNA. Notably, we identified that miR-520b was able to target 3'-untranslated region (3'UTR) of TET1 mRNA by luciferase reporter gene assays. Then, we revealed that miR-520b was able to reduce the expression of TET1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blotting analysis. In terms of function, 5-ethynyl-2-deoxyuridine (EdU) incorporation and colony formation assays demonstrated that the forced miR-520b expression remarkably inhibited proliferation of hepatoma cells, but TET1 overexpression could rescue the inhibition of cell proliferation mediated by miR-520b. Furthermore, anti-miR-520b enhanced proliferation of hepatoma cells, whereas silencing of TET1 abolished anti-miR-520b-induced acceleration of cell proliferation. Then, we validated that the expression levels of miR-520b were negatively related to those of TET1 mRNA in clinical HCC tissues. Thus, we conclude that miR-520b depresses proliferation of liver cancer cells through targeting 3'UTR of TET1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis.
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Carcinoma Hepatocelular/patología , Proliferación Celular , Proteínas de Unión al ADN/genética , Neoplasias Hepáticas/patología , MicroARNs/fisiología , Proteínas Proto-Oncogénicas/genética , ARN Mensajero/genética , Regiones no Traducidas 3' , Secuencia de Bases , Carcinoma Hepatocelular/genética , Línea Celular Transformada , Cartilla de ADN , Humanos , Neoplasias Hepáticas/genética , MicroARNs/genética , Oxigenasas de Función Mixta , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The central dogma of gene expression is that DNA is transcribed into messenger RNAs, which in turn serve as the template for protein synthesis. Recently, it has been reported that mRNAs display regulatory roles that rely on their ability to compete for microRNA binding, independent of their protein-coding function. However, the regulatory mechanism of mRNAs remains poorly understood. Here, we report that a hairpin within YAP mRNA 3'untranslated region (3'UTR) functions in regulation at post-transcription level through generating endogenous siRNAs (esiRNAs). Bioinformatics analysis for secondary structure showed that YAP mRNA displayed a hairpin structure (termed standard hairpin, S-hairpin) within its 3'UTR. Surprisingly, we observed that the overexpression of S-hairpin derived from YAP 3'UTR (YAP-sh) increased the luciferase reporter activities of transcriptional factor NF-κB and AP-1 in 293T cells. Moreover, we identified that a fragment from YAP-sh, an esiRNA, was able to target mRNA 3'UTR of NF2 (a member of Hippo-signaling pathway) and YAP mRNA 3'UTR itself in hepatoma cells. Thus, we conclude that the YAP-sh within YAP mRNA 3'UTR may serve as a novel regulatory element, which functions in regulation at post-transcription level. Our finding provides new insights into the mechanism of mRNAs in regulatory function.
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Proteínas Adaptadoras Transductoras de Señales/genética , Fosfoproteínas/genética , ARN Mensajero/química , ARN Mensajero/genética , Regiones no Traducidas 3' , Secuencia de Bases , Células HEK293 , Células Hep G2 , Vía de Señalización Hippo , Humanos , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Procesamiento Postranscripcional del ARN , ARN Mensajero/metabolismo , ARN Interferente Pequeño/química , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Homología de Secuencia de Ácido Nucleico , Transducción de Señal , Factores de Transcripción , Proteínas Señalizadoras YAPRESUMEN
BACKGROUND: Hepatitis B virus X protein (HBx) plays crucial roles in hepatocarcinogenesis. However, the underlying mechanism remains elusive. We have reported that HBx is able to up-regulate survivin in hepatocellular carcinoma tissues. The oncopreotein hepatitis B X-interacting protein (HBXIP), a target of miR-520b, is involved in the development of cancer. In this study, we focus on the investigation of hepatocarcinogenesis mediated by HBx. METHODS: The expression of HBx and survivin was examined in the liver tissues of HBx-Tg mice. The effect of HBx/survivin on the growth of LO2-X-S cells was determined by colony formation and transplantation in nude mice. The effect of HBx/survivin on promoter of miR-520b was determined by Western blot analysis, luciferase reporter gene assay, co-immunoprecipitation (co-IP) and chromatin immunoprecipitation (ChIP), respectively. The expression of HBx, survivin and HBXIP was detected by immunohistochemistry and real-time PCR in clinical HCC tissues, respectively. The DNA demethylation of HBXIP promoter was examined. The functional influence of miR-520b and HBXIP on proliferation of hepatoma cells was analyzed by MTT, colony formation, EdU and transplantation in nude mice in vitro and in vivo. RESULTS: In this study, we provided evidence that HBx up-regulated survivin in the liver cancer tissues of HBx-Tg mice aged 18 M. The engineered LO2 cell lines with survivin and/or HBx were successfully established, termed LO2-X-S. MiR-520b was down-regulated in LO2-X-S cells and clinical HCC tissues. Our data revealed that HBx survivin-dependently down-regulated miR-520b through interacting with Sp1 in the cells. HBXIP was highly expressed in LO2-X-S cells, liver cancer tissues of HBx-Tg mice aged 18 M and clinical HCC tissues (75.17%, 112/149). The expression level of HBXIP was positively associated with those of HBx or survivin in clinical HCC tissues. In addition, we showed that HBx survivin-dependently up-regulated HBXIP through inducing demethylation of HBXIP promoter in LO2-X-S cells and clinical HCC tissues. In function, low level miR-520b and high level HBXIP mediated by HBx with partner survivin contributed to the growth of LO2-X-S cells in vitro and in vivo. CONCLUSION: HBx accelerates hepatocarcinogenesis with partner survivin through modulating tumor suppressor miR-520b and oncoprotein HBXIP.
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Proteínas Adaptadoras Transductoras de Señales/genética , Carcinoma Hepatocelular/genética , Hepacivirus/genética , Proteínas Inhibidoras de la Apoptosis/genética , Neoplasias Hepáticas/genética , MicroARNs/genética , Transactivadores/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Carcinogénesis/genética , Carcinoma Hepatocelular/etiología , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Línea Celular , Metilación de ADN , Regulación de la Expresión Génica , Hepacivirus/metabolismo , Hepatitis B/complicaciones , Hepatitis B/genética , Hepatitis B/metabolismo , Hepatitis B/patología , Interacciones Huésped-Patógeno , Humanos , Proteínas Inhibidoras de la Apoptosis/metabolismo , Neoplasias Hepáticas/etiología , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Ratones , Ratones Desnudos , Ratones Transgénicos , MicroARNs/metabolismo , Trasplante de Neoplasias , Regiones Promotoras Genéticas , Transducción de Señal , Factor de Transcripción Sp1/genética , Factor de Transcripción Sp1/metabolismo , Survivin , Transactivadores/metabolismo , Proteínas Reguladoras y Accesorias ViralesRESUMEN
The infection of hepatitis B virus (HBV) is closely associated with the development of hepatocellular carcinoma (HCC), in which HBV X protein (HBx) plays crucial roles. MicroRNAs are involved in diverse biologic functions and in carcinogenesis by regulating gene expression. In the present study, we aim to investigate the underlying mechanism by which HBx enhances hepatocarcinogenesis. We found that miR-205 was downregulated in 33 clinical HCC tissues in comparison with adjacent noncancerous hepatic tissues. The expression levels of miR-205 were inversely correlated with those of HBx in abovementioned tissues. Then, we demonstrated that HBx was able to suppress miR-205 expression in hepatoma and liver cells. We validated that miR-205 directly targeted HBx mRNA. Ectopic expression of miR-205 downregulated HBx, whereas depletion of endogenous miR-205 upregulated HBx in hepatoma cells. Notably, our data revealed that HBx downregulated miR-205 through inducing hypermethylation of miR-205 promoter in the cells. In terms of function, the forced miR-205 expression remarkably inhibited the HBx-enhanced proliferation of hepatoma cells in vitro and in vivo, suggesting that miR-205 is a potential tumor-suppressive gene in HCC. HBx-transgenic mice showed that miR-205 was downregulated in the liver. Importantly, HBx was able to abrogate the effect of miR-205 on tumor suppression in carcinogenesis. Therefore, we conclude that HBx is able to inhibit tumor suppressor miR-205 to enhance hepatocarcinogenesis through inducing hypermethylation of miR-205 promoter during their interaction. Therapeutically, miR-205 may be useful in the treatment of HCC.