RESUMEN
More than 35% of lung adenocarcinoma patients have bone metastases at diagnosis and have a poor survival. Periostin, a carboxylated matrix protein, mediates lung cancer cell dissemination by promoting epithelial-mesenchymal transition, and is involved in bone response to mechanical stress and bone formation regulation. This suggests that periostin may be used as a biomarker to predict survival in lung cancer patients. Serum periostin was assessed at diagnosis in a prospective cohort of 133 patients with lung adenocarcinoma of all stages. Patients were divided into localized and bone metastatic groups. Both groups were matched to healthy controls. Survival analysis and Cox proportional hazards models were conducted in the total population and in bone metastatic group. The median serum periostin level was higher in bone metastatic (nâ¯=â¯67; median: 1752â¯pmol/L) than in the localized group (nâ¯=â¯66; 861â¯pmol/L; pâ¯<â¯0.0001). Patients with high periostin (>median) had a poorer overall survival in the whole population (33.3â¯weeks vs. NR; pâ¯<â¯0.0001) and the bone metastatic group (24.4 vs. 66.1â¯weeks; pâ¯<â¯0.001). In multivariate analysis, patients with high periostin had increased risk of death (HRâ¯=â¯2.09, 95%CI [1.06-4.13]; pâ¯=â¯0.03). This was also found in the bone metastatic group (HRâ¯=â¯3.62, 95%CI [1.74-7.52]; pâ¯=â¯0.0005). Immunohistochemistry on bone metastasis biopsies showed periostin expression in the bone matrix and nuclear and cytoplasmic staining in cancer cells. Serum periostin was an independent survival biomarker in all-stage and in bone metastatic lung adenocarcinoma patients. IHC data suggest that periostin might be induced in cancer cells in bone metastatic niche in addition to bone microenvironment expression.
RESUMEN
The orphan nuclear estrogen receptor-related receptor alpha (ERRalpha), is expressed by many cell types, but is very highly expressed by osteoblastic cells in which it transactivates at least one osteoblast-associated gene, osteopontin. To study the putative involvement of ERRalpha in bone, we first assessed its expression in rat calvaria (RC) in vivo and in RC cells in vitro. ERRalpha mRNA and protein were expressed at all developmental stages from early osteoprogenitors to bone-forming osteoblasts, but protein was most abundant in mature cuboidal osteoblasts. To assess a functional role for ERRalpha in osteoblast differentiation and bone formation, we blocked its expression by antisense oligonucleotides in either proliferating or differentiating RC cell cultures and found inhibition of cell growth and a proliferation-independent inhibition of differentiation. On the other hand, ERRalpha overexpression in RC cells increased differentiation and maturation of progenitors to mature bone-forming cells. Our findings show that ERRalpha is highly expressed throughout the osteoblast developmental sequence and plays a physiological role in differentiation and bone formation at both proliferation and differentiation stages. In addition, we found that manipulation of receptor levels in the absence of known ligand is a fruitful approach for functional analysis of this orphan receptor and identification of potential target genes.
Asunto(s)
Diferenciación Celular , Osteoblastos/citología , Osteoblastos/metabolismo , Osteogénesis , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Estrógenos/metabolismo , Animales , Calcificación Fisiológica , División Celular , Linaje de la Célula , Células Cultivadas , Regulación del Desarrollo de la Expresión Génica , Inmunohistoquímica , Peso Molecular , Oligonucleótidos Antisentido/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , Receptores Citoplasmáticos y Nucleares/química , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Estrógenos/antagonistas & inhibidores , Receptores de Estrógenos/química , Receptores de Estrógenos/genética , Cráneo/citología , Cráneo/embriología , Cráneo/metabolismo , Células Madre/citología , Células Madre/metabolismo , Transfección , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
Bone is the most common metastatic site for breast cancer. Estrogen-related-receptor alpha (ERRα) has been implicated in cancer cell invasiveness. Here, we established that ERRα promotes spontaneous metastatic dissemination of breast cancer cells from primary mammary tumors to the skeleton. We carried out cohort studies, pharmacological inhibition, gain-of-function analyses in vivo and cellular and molecular studies in vitro to identify new biomarkers in breast cancer metastases. Meta-analysis of human primary breast tumors revealed that high ERRα expression levels were associated with bone but not lung metastases. ERRα expression was also detected in circulating tumor cells from metastatic breast cancer patients. ERRα overexpression in murine 4T1 breast cancer cells promoted spontaneous bone micro-metastases formation when tumor cells were inoculated orthotopically, whereas lung metastases occurred irrespective of ERRα expression level. In vivo, Rank was identified as a target for ERRα. That was confirmed in vitro in Rankl stimulated tumor cell invasion, in mTOR/pS6K phosphorylation, by transactivation assay, ChIP and bioinformatics analyses. Moreover, pharmacological inhibition of ERRα reduced primary tumor growth, bone micro-metastases formation and Rank expression in vitro and in vivo. Transcriptomic studies and meta-analysis confirmed a positive association between metastases and ERRα/RANK in breast cancer patients and also revealed a positive correlation between ERRα and BRCA1mut carriers. Taken together, our results reveal a novel ERRα/RANK axis by which ERRα in primary breast cancer promotes early dissemination of cancer cells to bone. These findings suggest that ERRα may be a useful therapeutic target to prevent bone metastases.
Asunto(s)
Neoplasias Óseas/metabolismo , Neoplasias de la Mama/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Mamarias Animales/metabolismo , Proteínas de Neoplasias/metabolismo , Receptor Activador del Factor Nuclear kappa-B/biosíntesis , Receptores de Estrógenos/metabolismo , Animales , Neoplasias Óseas/genética , Neoplasias Óseas/patología , Neoplasias Óseas/secundario , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Femenino , Humanos , Neoplasias Mamarias Animales/genética , Neoplasias Mamarias Animales/patología , Ratones , Ratones Endogámicos BALB C , Proteínas de Neoplasias/genética , Receptor Activador del Factor Nuclear kappa-B/genética , Receptores de Estrógenos/genética , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
OBJECTIVES: Subchondral bone loss is a characteristic feature of inflammatory arthritis. Recently, estrogen receptor-related receptor-alpha (ERR-alpha), an orphan nuclear receptor, has been found to be involved in activation of macrophages. We hypothesized that ERR-alpha which is expressed and also functional in articular chondrocytes, osteoblasts and osteoclasts, may be involved in rodent models of inflammatory arthritis. METHODS: Erosive arthritis was induced in DBA/1 mice by injection of type II collagen in Freund's complete adjuvant. RNA was isolated from the bone and joints and expression of ERR-alpha and cartilage (GDF5 and Col2a1) and bone [bone sialoprotein (BSP) and osteocalcin (OCN)] markers was analysed by semi-quantitative PCR. RESULTS: We report for the first time that the expression of ERR-alpha is dysregulated in bones and joints in a mouse model of inflammatory arthritis. Specifically, we show that ERR-alpha expression is down-regulated early in bone and later in joints of mice with type II CIA. Concomitantly, temporal changes were observed in GDF-5 and Col2a1 expression in joints following both initial injection and booster injection of type II collagen. Similarly, down-regulation of ERR-alpha mRNA expression in subchondral bone in mice with induced joint inflammation was also paralleled by down-regulation of markers of bone formation (BSP, OCN). CONCLUSIONS: These data suggest that dysregulation of ERR-alpha expression may precede and contribute to the destruction of cartilage and bone accompanying inflammatory arthritis.
Asunto(s)
Artritis Experimental/metabolismo , Artritis Reumatoide/metabolismo , Receptores de Estrógenos/metabolismo , Animales , Huesos/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo , Expresión Génica , Articulaciones/metabolismo , Masculino , Ratones , Ratones Endogámicos DBA , Monocitos/metabolismo , ARN Mensajero/genética , Receptores de Estrógenos/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
We report for the first time the expression of estrogen receptor-related receptor (ERR)-alpha in fetal and adult rat chondrocytes in growth plate and articular cartilage and the rat chondrogenic cell line C5.18 cells in vitro. ERRalpha mRNA and protein were expressed from proliferating chondrocyte to mature chondrocyte stages. We show that overexpressing ERRalpha in C5.18 cell cultures induces an increase in Sry-type high-mobility-group box transcription factor (Sox)-9 expression, a master gene in cartilage formation. In parallel, we report Sox9 promoter regulation by ERRalpha in C5.18 cells. To assess a functional role for ERRalpha in chondrogenesis, its expression was blocked by antisense oligonucleotides in C5.18 cell cultures, and this led to inhibition of cartilage formation associated with down-regulation of Sox9 and Indian hedgehog expression and maturation of proliferating chondrocytes into hypertrophic chondrocytes in vitro. Together these results implicate ERRalpha in the formation and maintenance of cartilage and also suggest that agonists and antagonists of ERRalpha may be useful as therapeutic agents in a wide variety of diseases affecting cartilage and joints.
Asunto(s)
Cartílago/fisiología , Condrogénesis/fisiología , Proteínas del Grupo de Alta Movilidad/fisiología , Receptores de Estrógenos/fisiología , Factores de Transcripción/fisiología , Animales , Cartílago/embriología , Cartílago/metabolismo , Proliferación Celular , Células Cultivadas , Condrocitos/metabolismo , Regulación de la Expresión Génica , Proteínas del Grupo de Alta Movilidad/metabolismo , Modelos Biológicos , Ratas , Receptores de Estrógenos/metabolismo , Factor de Transcripción SOX9 , Factores de Transcripción/metabolismo , Transfección , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
The superfamily of nuclear receptors comprises transcription factors that depend on a ligand for their activity. In addition, the superfamily includes a number of orphan receptors, for which no ligand is known. We report here that the orphan receptor estrogen receptor related alpha receptor (ERR alpha) stimulates the expression of the thyroid hormone receptor alpha (TR alpha) gene promoter. We characterized a responsive site that is both necessary and sufficient for ERR alpha-induced transactivation. In addition, we show that both TR alpha and ERR alpha are coexpressed in embryonic intestine, brown fat and heart as well as in the adult gonads. In the testis, expression of both receptors can be found in the seminiferous tubes where it is totally restricted to spermatocytes I. Altogether this suggests that TR alpha is an in vivo target of ERR alpha.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Regiones Promotoras Genéticas , Receptores Citoplasmáticos y Nucleares/fisiología , Receptores de Estrógenos/fisiología , Receptores de Hormona Tiroidea/genética , Activación Transcripcional , Tejido Adiposo Pardo/embriología , Tejido Adiposo Pardo/crecimiento & desarrollo , Tejido Adiposo Pardo/metabolismo , Animales , Secuencia de Bases , Línea Celular , Secuencia de Consenso , Femenino , Corazón Fetal/metabolismo , Fibroblastos , Genes Reporteros , Gónadas/embriología , Gónadas/crecimiento & desarrollo , Gónadas/metabolismo , Células HeLa , Humanos , Mucosa Intestinal/metabolismo , Intestinos/embriología , Luciferasas/biosíntesis , Masculino , Ratones , Datos de Secuencia Molecular , Miocardio/metabolismo , Especificidad de Órganos , Ratas , Receptores Citoplasmáticos y Nucleares/biosíntesis , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Estrógenos/biosíntesis , Receptores de Estrógenos/genética , Receptores de Hormona Tiroidea/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , Secuencias Reguladoras de Ácidos Nucleicos , Alineación de Secuencia , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
Estrogen receptor-related receptor-alpha (ERRalpha) is an orphan nuclear receptor with sequence homology to the estrogen receptors, ERalpha/beta, but it does not bind estrogen. However, several recent studies suggest that ERRalpha not only plays a functional role in osteoblasts but also impinges on the estrogen axis in bone, as it does in at least certain other estrogen target tissues. We summarize here data on ERRalpha and its cellular and molecular modes of action that have broad implications for considering the potential role of this orphan receptor as a new therapeutic target in osteopenic disorders such as osteoporosis as well as other estrogen-responsive conditions.
Asunto(s)
Huesos/efectos de los fármacos , Estrógenos/farmacología , Receptores Citoplasmáticos y Nucleares/fisiología , Receptores de Estrógenos/fisiología , Aromatasa/genética , Huesos/fisiología , Regulación de la Expresión Génica , Humanos , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa de Tipo III , Osteogénesis , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
We studied the expression of the estrogen-related receptor 1 (ERR-1) during mouse embryonic development. ERR-1 is expressed at very early stages in ES cells and at E8.5 in the mesodermal cells of the visceral yolk sac. ERR-1 continues to be expressed later in mesodermal tissues and particularly in heart and in skeletal muscles. This expression persists during all the embryonic development and in adult stage. ERR-1 transcripts level increases during muscle differentiation. Accordingly, we show that ERR-1 expression increases during the myoblast to myotube transition in differentiating C2 myoblastic cells. ERR-1 has also been detected in the nervous system during embryonic development. At E10.5, a high level of ERR-1 transcripts can be observed in differentiated cells of the intermediate zone of the spinal cord which also suggests a role of ERR-1 in the differentiation of the nervous system. The same is observed in the telencephalon vesicules at E13.5. Later, at E15.5 and E17.5, expression persists in the spinal cord but decreases dramatically in the central nervous system. Moreover, ERR-1 expression increases during skin formation and is detected in the stratum spinosum which contains differentiated Malpighian cells. Finally, we also observed ERR-1 in endodermal derivatives such as the epithelium of intestine and urogenital system. The DNA target of ERR-1 has been identified to be the SF-1/FTZ-F1 responsive element (SFRE) and we show in this paper that SF-1/FTZ-F1 and ERR-1 bind to and activate transcription independently through the SFRE element. Our study suggests that ERR-1 may be implicated in numerous physiological or developmental functions, particularly in the muscle, the central and peripheral nervous system and the epidermis. Interestingly, in these various systems ERR-1 expression is correlated with post-mitotic cells stage, suggesting that ERR-1 may play a role in the differentiation process.
Asunto(s)
Desarrollo Embrionario y Fetal , Regulación del Desarrollo de la Expresión Génica , Receptores Citoplasmáticos y Nucleares/biosíntesis , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Estrógenos/biosíntesis , Receptores de Estrógenos/genética , Animales , Femenino , Hibridación in Situ , Ratones , Embarazo , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
Estrogen receptor-related receptor alpha (ERR alpha) is an orphan nuclear receptor closely related to the estrogen receptor (ER), whose expression covers various stages of embryonic development and persists in certain adult tissues. We show that ERR alpha binds as a homodimer on a specific target sequence, the SFRE (SF-1 response element), already known to respond to the orphan nuclear receptor SF-1. Target sequences that are related to the SFRE and that discriminate between ERR alpha and SF-1 were identified. We have also analyzed the transcriptional properties of the ERR alpha originating from various species. All ERR alpha orthologs act as potent transactivators through the consensus SFRE. ERR alpha activity depends on the putative AF2AD domain, as well as on a serum compound that is withdrawn by charcoal treatment, suggesting the existence of a critical regulating factor brought by serum.
Asunto(s)
Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Estrógenos/genética , Receptores de Estrógenos/metabolismo , Animales , Secuencia de Bases , Línea Celular , Carbón Orgánico , Medios de Cultivo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Dimerización , Factores de Transcripción Fushi Tarazu , Proteínas de Homeodominio , Humanos , Ratones , Regiones Promotoras Genéticas , Ratas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Elementos de Respuesta , Factor Esteroidogénico 1 , Transactivadores/genética , Transactivadores/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transcripción Genética , Pez Cebra/genética , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
We studied the expression of estrogen-related receptor ERR-1 during mouse embryonic development. ERR-1 mRNA is present in bones formed by both the endochondral and intramembranous routes, and the onset of its expression coincides with bone formation. By RT-PCR experiments, we found that ERR-1, but not the related receptor ERR-2, is expressed in osteoblastic osteosarcoma cell lines as well as in primary osteoblastic cell populations derived from normal human bone. By gel shift analysis we found that ERR-1 binds as a monomer specifically to the SFRE sequence (SF-1-responsive-element; TCAAGGTCA). Mutation analysis revealed that both the core AGGTCA motif and the TCA 5'-extension are required for efficient ERR-1 binding. In transient transfection assays, ERR-1 acts as a potent transactivator through the SFRE sequence. This effect is cell-specific since ERR-1 activates transcription in the rat osteosarcoma cell line ROS 17.2/8 as well as in HeLa, NB-E, and FREJ4 cells but not in COS1 and HepG2 cells. Notably, the osteopontin (a protein expressed by osteoblasts and released in the bone matrix) gene promoter is a target for ERR-1 transcriptional regulation. Our findings suggest a role for ERR-1 in bone development and metabolism.
Asunto(s)
Desarrollo Óseo/genética , Huesos/embriología , Regulación del Desarrollo de la Expresión Génica/genética , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Estrógenos/genética , Factores de Transcripción/metabolismo , Activación Transcripcional/genética , Animales , Secuencia de Bases , Huesos/metabolismo , Línea Celular , ADN/metabolismo , Cartilla de ADN/química , Embrión de Mamíferos/anatomía & histología , Embrión de Mamíferos/química , Hibridación in Situ , Ratones , Reacción en Cadena de la Polimerasa , Unión Proteica/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Estrógenos/metabolismo , Análisis de Secuencia , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
Prostate cancer (PCa) is one of the major public health problems in Western countries. Recently, the TMPRSS2:ERG gene fusion, which results in the aberrant expression of the transcription factor ERG, has been shown to be the most common gene rearrangement in PCa. Previous studies have determined the contributions of this fusion in PCa disease initiation and/or progression in vitro and in vivo. In this study on TMPRSS2:ERG regulation in PCa, we used an androgen receptor and TMPRSS2:ERG fusion double-negative PCa cell model: PC3c. In three cell clones with different TMPRSS2:ERG expression levels, ectopic expression of the fusion resulted in significant induction of cell migration and invasion in a dose-dependent manner. In agreement with this phenotype, high-throughput microarray analysis revealed that a set of genes, functionally associated with cell motility and invasiveness, were deregulated in a dose-dependent manner in TMPRSS2:ERG-expressing cells. Importantly, we identified increased MMP9 (Metalloproteinase 9) and PLXNA2 (Plexin A2) expression in TMPRSS2:ERG-positive PCa samples, and their expression levels were significantly correlated with ERG expression in a PCa cohort. In line with these findings, there was evidence that TMPRSS2:ERG directly and positively regulates MMP9 and PLXNA2 expression in PC3c cells. Moreover, PLXNA2 upregulation contributed to TMPRSS2:ERG-mediated enhancements of PC3c cell migration and invasion. Furthermore, and importantly, PLXNA2 expression was upregulated in metastatic PCa tumors compared with localized primary PCa tumors. This study provides novel insights into the role of the TMPRSS2:ERG fusion in PCa metastasis.
Asunto(s)
Metaloproteinasa 9 de la Matriz/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Neoplasias de la Próstata/metabolismo , Receptores de Superficie Celular/metabolismo , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Regulación de la Expresión Génica , Humanos , Metástasis Linfática , Masculino , Proteínas de Fusión Oncogénica/genética , Fenotipo , Neoplasias de la Próstata/patología , TranscriptomaRESUMEN
In just 3 years, striking new advances have been made in understanding the molecular mechanisms that govern the crosstalk between osteoblasts/stromal cells and hematopoietic osteoclast precursor cells that leads to osteoclastogenesis. Led first by the discovery of osteoprotegerin (OPG), a naturally occurring protein with potent osteoclastogenesis inhibitory activity, rapid progress was made to the isolation of RANKL, a transmembrane ligand expressed on osteoblasts/stromal cells that binds to RANK, a transmembrane receptor on hematopoietic osteoclast precursor cells. The interaction of RANK and RANKL initiates a signaling and gene expression cascade that results in differentiation and maturation of osteoclast precursor cells to active osteoclasts capable of resorbing bone. OPG acts as a decoy receptor, binding to RANKL and blocking its interaction with RANK, inhibiting osteoclast development. Many of the calciotropic hormones and cytokines, including 1,25(OH)2D3, PTH, PGE2 and IL-11, appear to act through a dual capacity to inhibit production of OPG and stimulate production of RANKL. Estrogen, on the other hand, appears to inhibit production of RANKL and RANKL-stimulated osteoclastogenesis. Recently, the results of the first clinical trial with OPG supported its potential as a therapeutic agent for diseases such as osteoporosis. The new understanding provided by the RANK/RANKL/OPG paradigm for both differentiation of osteoclasts and their activation has had tremendous impact on the field and opened new avenues for development of possible treatments of diseases characterized by excessive bone resorption.
Asunto(s)
Resorción Ósea/fisiopatología , Glicoproteínas/fisiología , Ligandos , Osteoblastos/citología , Osteogénesis/fisiología , Receptores Citoplasmáticos y Nucleares/fisiología , Células del Estroma/citología , Proteínas Portadoras/fisiología , Diferenciación Celular , Glicoproteínas/uso terapéutico , Humanos , Glicoproteínas de Membrana/fisiología , Osteoclastos/citología , Osteoprotegerina , Ligando RANK , Receptor Activador del Factor Nuclear kappa-B , Receptores Citoplasmáticos y Nucleares/uso terapéutico , Receptores del Factor de Necrosis Tumoral/fisiologíaRESUMEN
In just 3 years, striking new advances have been made in understanding the molecular mechanisms that govern the crosstalk between osteoblasts/stromal cells and hemopoietic osteoclast precursor cells that leads to osteoclastogenesis. Led first by the discovery of osteoprotegerin (OPG), a naturally occurring protein with potent osteoclastogenesis inhibitory activity, rapid progress was made to the isolation of RANKL, a transmembrane ligand expressed on osteoblasts/stromal cells, that binds to RANK, a transmembrane receptor on hemopoietic osteoclast precursor cells. The interaction of RANK and RANKL initiates a signaling and gene expression cascade that results in differentiation and maturation of osteoclast precursor cells to active osteoclasts capable of resorbing bone. Osteoprotegerin acts as a decoy receptor; it binds to RANKL and blocks its interaction with RANK, thus inhibiting osteoclast development. Many of the calciotropic hormones and cytokines, including vitamin D3, parathyroid hormone, prostaglandin E2 and interleukin-11, appear to stimulate osteoclastogenesis through the dual action of inhibiting production of OPG and stimulating production of RANKL. Estrogen, on the other hand, appears to inhibit production of RANKL and RANKL-stimulated osteoclastogenesis. Recently, the results of the first clinical trial with OPG supported its potential as a therapeutic agent for osteoporosis. The new understanding provided by the RANK/RANKL/OPG paradigm for both differentiation and activation of osteoclasts has had tremendous impact on the field of bone biology and has opened new avenues for development of possible treatments of diseases characterized by excessive bone resorption.
Asunto(s)
Resorción Ósea/fisiopatología , Glicoproteínas/fisiología , Osteoclastos/fisiología , Receptores Citoplasmáticos y Nucleares , Proteínas Portadoras/fisiología , Femenino , Glicoproteínas/química , Glicoproteínas/uso terapéutico , Humanos , Glicoproteínas de Membrana/fisiología , Osteoclastos/citología , Osteoporosis Posmenopáusica/tratamiento farmacológico , Osteoprotegerina , Ligando RANK , Receptor Activador del Factor Nuclear kappa-B , Receptores del Factor de Necrosis Tumoral/química , Receptores del Factor de Necrosis Tumoral/fisiología , Receptores del Factor de Necrosis Tumoral/uso terapéuticoRESUMEN
We have identified and characterized a new orphan member of the nuclear hormone receptor superfamily in the chicken. This new gene, called Rev-erb beta, exhibits strong homologies with the Rev-erb alpha/ear-1 orphan receptor gene, which partially overlaps the thyroid hormone receptor alpha gene in opposite orientation. We demonstrate that both Rev-erb alpha and Rev-erb beta genes are conserved in their C and E domains. Rev-erb beta binds to DNA as a monomer and recognizes the same binding motif as the alpha gene product. The Rev-erb beta gene product does not interact with retinoid X receptors, as revealed by gel shift experiments. In situ hybridization experiments show that Rev-erb beta is expressed in the central and peripheric nervous system, spleen, and mandibular and maxillar processes, as well as in blood islands. During embryonic development, we noticed a striking specific distribution of Rev-erb beta transcripts in the notochord at 24 h and later on, in the floor plate of the neural tube. We propose that Rev-erb beta may play an important role in the complex network of inductive signals, which control neuron differentiation.
Asunto(s)
Proteínas de Unión al ADN/biosíntesis , Sistema Nervioso/metabolismo , Receptores Citoplasmáticos y Nucleares/biosíntesis , Receptores de Hormona Tiroidea , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Northern Blotting , Sistema Nervioso Central/metabolismo , Embrión de Pollo , Pollos , Clonación Molecular , Secuencia Conservada , ADN/metabolismo , Cartilla de ADN , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Fibroblastos/metabolismo , Biblioteca de Genes , Humanos , Datos de Secuencia Molecular , Músculos/metabolismo , Sistema Nervioso/crecimiento & desarrollo , Nervios Periféricos/metabolismo , Reacción en Cadena de la Polimerasa , Biosíntesis de Proteínas , Receptores Citoplasmáticos y Nucleares/química , Receptores Citoplasmáticos y Nucleares/metabolismo , Reticulocitos/metabolismo , Homología de Secuencia de Aminoácido , Bazo/metabolismo , Transcripción GenéticaRESUMEN
We describe the cloning and functional characterization of Schistosoma mansoni retinoid-X-receptor (SmRXR; NR2B4-B), a novel member of the nuclear receptor superfamily from S. mansoni, a homologue of vertebrate retinoid-X-receptor. The DNA-binding C domain of SmRXR shows 80% sequence identity to both human RXRalpha and Drosophila ultraspiracle (USP), but a much lower level of conservation of the ligand-binding E domain (22-25% identity). Phylogenetic analysis places SmRXR within the RXR group as an early offshoot of this clade. SmRXR mRNA is expressed at all life-cycle stages but at higher levels in the free-living larval stages. However, the SmRXR protein is expressed at markedly different levels, being almost absent from eggs while present at the highest concentration in schistosomula. Recombinant SmRXR fails to bind to the consensus direct repeat response elements, either alone, or as a heterodimer with mouse retinoic acid receptor alpha or the Drosophila ecdysone receptor. However, the use of chimaeric constructions shows that the C domain of SmRXR will bind to conventional response elements as a heterodimer, and that its specificity is modified by the presence of the D and E domains. In accordance with these results, native SmRXR failed to transactivate the transcription of a reporter gene after cotransfection of mammalian cell lines.
Asunto(s)
Isoformas de Proteínas/química , Receptores de Ácido Retinoico/química , Schistosoma mansoni/genética , Factores de Transcripción/química , Empalme Alternativo , Secuencia de Aminoácidos , Animales , Sitios de Unión , Línea Celular , Clonación Molecular , Secuencia de Consenso , ADN/metabolismo , Dimerización , Drosophila melanogaster/química , Huevos/análisis , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Genes Reporteros , Humanos , Larva/química , Ratones , Datos de Secuencia Molecular , Filogenia , Isoformas de Proteínas/genética , Estructura Terciaria de Proteína , Ratas , Ratas Endogámicas F344 , Receptores de Ácido Retinoico/genética , Receptores de Ácido Retinoico/metabolismo , Receptores de Esteroides/química , Proteínas Recombinantes de Fusión/metabolismo , Receptores X Retinoide , Schistosoma mansoni/crecimiento & desarrollo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Especificidad de la Especie , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Activación Transcripcional , TransfecciónRESUMEN
Fibrates are widely used hypolipidemic drugs which activate the nuclear peroxisome proliferator-activated receptor (PPAR) alpha and thereby alter the transcription of genes controlling lipoprotein metabolism. Fibrates influence plasma high density lipoprotein and its major protein, apolipoprotein (apo) A-I, in an opposite manner in man (increase) versus rodents (decrease). In the present study we studied the molecular mechanisms of this species-specific regulation of apoA-I expression by fibrates. In primary rat and human hepatocytes fenofibric acid, respectively, decreased and increased apoA-I mRNA levels. The absence of induction of rat apoA-I gene expression by fibrates is due to 3 nucleotide differences between the rat and the human apoA-I promoter A site, rendering a positive PPAR-response element in the human apoA-I promoter nonfunctional in rats. In contrast, rat, but not human, apoA-I transcription is repressed by the nuclear receptor Rev-erbalpha, which binds to a negative response element adjacent to the TATA box of the rat apoA-I promoter. In rats fibrates increase liver Rev-erbalpha mRNA levels >10-fold. In conclusion, the opposite regulation of rat and human apoA-I gene expression by fibrates is linked to differences in cis-elements in their respective promoters leading to repression by Rev-erbalpha of rat apoA-I and activation by PPARalpha of human apoA-I. Finally, Rev-erbalpha is identified as a novel fibrate target gene, suggesting a role for this nuclear receptor in lipid and lipoprotein metabolism.