RESUMO
BACKGROUND: Breast cancer (BRCA) is a malignant cancer that threatened the life of female with unsatisfactory prognosis. The aim of this study was to identify prognostic nuclear receptors (NRs) signature of BRCA. METHODS: BRCA patient samples were collected from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) database. Consensus clustering analysis, univariate Cox regression analysis and the least absolute shrinkage and selection operator (LASSO) Cox regression analysis were performed to evaluate, select NRs as prognostic factors and build Risk Score model. GSEA analysis was explored to check signaling differences between High- and Low-Risk group. Nomogram model basing on age and Risk Score was established to predict the 1-, 3- and 5-year survival. Model performance was assessed by a time-dependent receiver operating characteristic (ROC) curve and calibration plot. CIBERSORT, ESTIMATE and TIMER algorithm were introduced to evaluate the immune landscape. RESULTS: NR3C1, NR4A3, THRA, RXRG, NR2F6, NR1D2 and RORB were optimized as a prognostic signature for BRCA. This seven-NR-based Risk Score could effectively predict overall survival status. The area under the curve (AUC) of 1-, 3- and 5-year overall survival are 0.702, 0.734 and 0.722 in TCGA training cohort, and 0.630, 0.721 and 0.823 in GEO validation cohort, respectively. Calibration plot demonstrated satisfactory agreement between predictive and observed outcomes. Nomogram model worked well on predicting survival probabilities. Multiple cancer-related pathways were highly enriched in High-Risk group. High- and Low-Risk groups showed significant differed immune cell infiltration. There exists an obvious connection between Risk Score and immune checkpoints LAG3, PD1 and TIM3. CONCLUSION: The seven-NR-based Risk Score represents a promising signature for estimating overall survival in patients with BRCA, and is correlated with the immune microenvironment.
Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Receptores Citoplasmáticos e Nucleares/biossíntese , Neoplasias da Mama/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Receptores Citoplasmáticos e Nucleares/genética , Taxa de SobrevidaRESUMO
The estrogen receptor alpha (ERα) is a ligand-activated transcription factor whose activity is modulated by its interaction with multiple protein complexes. In this work, we have identified the protein interferon alpha inducible protein 27 (IFI27/ISG12) as a novel ERα-associated protein. IFI27/ISG12 transcription is regulated by interferon and estradiol and its overexpression is associated to reduced overall survival in ER+ breast cancer patients but its function in mammary gland tissue remains elusive. In this study we showed that overexpression of IFI27/ISG12 in breast cancer cells attenuates ERα transactivation activity and the expression of ERα-dependent genes. Our results demonstrated that IFI27/ISG12 overexpression in MCF-7 cells reduced their proliferation rate in 2-D and 3-D cell culture assays and impaired their ability to migrate in a wound-healing assay. We show that IFI27/ISG12 downregulation of ERα transactivation activity is mediated by its ability to facilitate the interaction between ERα and CRM1/XPO1 that mediates the nuclear export of large macromolecules to the cytoplasm. IFI27/ISG12 overexpression was shown to impair the estradiol-dependent proliferation and tamoxifen-induced apoptosis in breast cancer cells. Our results suggest that IFI27/ISG12 may be an important factor in regulating ERα activity in breast cancer cells by modifying its nuclear versus cytoplasmic protein levels. We propose that IFI27/ISG12 may be a potential target of future strategies to control the growth and proliferation of ERα-positive breast cancer tumors.
Assuntos
Neoplasias da Mama/metabolismo , Regulação para Baixo/fisiologia , Receptor alfa de Estrogênio/biossíntese , Carioferinas/biossíntese , Proteínas de Membrana/biossíntese , Receptores Citoplasmáticos e Nucleares/biossíntese , Ativação Transcricional/fisiologia , Neoplasias da Mama/genética , Bases de Dados Genéticas , Regulação para Baixo/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Carioferinas/genética , Células MCF-7 , Proteínas de Membrana/genética , Receptores Citoplasmáticos e Nucleares/genética , Tamoxifeno/farmacologia , Ativação Transcricional/efeitos dos fármacos , Proteína Exportina 1RESUMO
OBJECTIVE: The aim of this work was to evaluate a pediatric ependymoma protein expression that may be useful as a molecular biomarker candidate for prognosis, correlated with clinical features such as age, gender, histopathological grade, ependymal tumor recurrence and patient survival. PATIENTS AND METHODS: Immunohistochemistry assays were performed for GNAO1, ASAH1, IMMT, IPO7, Cyclin D1, P53 and Ki-67 proteins. Kaplan-Meier and Cox analysis were performed for age, gender, histopathological grade, relapse and survival correlation. RESULTS: We found that three proteins correlate with histopathological grade and relapse; two proteins correlate with survival; one protein does not correlate with any clinical feature. CONCLUSION: Our results suggest that, out of the proteins analyzed, five may be considered suitable prognostic biomarkers and one may be considered a predictive biomarker for response to treatment of pediatric ependymoma.
Assuntos
Ceramidase Ácida/biossíntese , Neoplasias Encefálicas/metabolismo , Ependimoma/metabolismo , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/biossíntese , Carioferinas/biossíntese , Proteínas Mitocondriais/biossíntese , Proteínas Musculares/biossíntese , Receptores Citoplasmáticos e Nucleares/biossíntese , Ceramidase Ácida/genética , Adolescente , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/genética , Criança , Pré-Escolar , Estudos de Coortes , Ependimoma/diagnóstico , Ependimoma/genética , Feminino , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Recém-Nascido , Carioferinas/genética , Masculino , Proteínas Mitocondriais/genética , Proteínas Musculares/genética , Prognóstico , Receptores Citoplasmáticos e Nucleares/genética , Fatores de TempoRESUMO
The Receptor for Activated C Kinase 1 (RACK1), a scaffold protein member of the tryptophan-aspartate (WD) repeat family, folds in a seven-bladed ß-propeller structure that permits the association of proteins to form active complexes. Mosquitoes of the genus Aedes sp., are vectors of virus producing important diseases such as: dengue, chikungunya and yellow fever. Based on the highly conserved gene sequence of AeaeRACK1 of the mosquito Aedes aegypti we characterized the mRNA and protein of the homologous AealRACK1 from the Ae. albopictus-derived cell line C6/36 HT. Two protein species differing in MW/pI values were observed at 35kDa/8.0 and 36kDa/6.5. The behavior of AealRACK1 was studied inducing stress with serum deprivation and the glucocorticoid dexamethasone. Both stressors induced increase of the expression of AealRACK1 mRNA and proteins. In serum-deprived cells AealRACK1 protein was located cortically near the plasma membrane in contrast to dexamethasone-treated cells where the protein formed a dotted pattern in the cytoplasm. In addition, 33 protein partners were identified by immunoprecipitation and mass spectrometry. Most of the identified proteins were ribosomal, involved in signaling pathways and stress responses. Our results suggest that AealRACK1 in C6/36 HT cells respond to stress increasing its synthesis and producing phosphorylated activated form. BIOLOGICAL SIGNIFICANCE: Insect cells adapt to numerous environmental stressors, including chemicals and invasion of pathogenic microorganisms among others, coordinating cellular and organismal responses. Individual cells sense the environment using receptors that trigger signaling pathways that regulate expression of specific effector proteins and/or cellular responses as movement or secretion. In the coordination of responses to stress, scaffold proteins are pivotal molecules that recruit other proteins forming active complexes. The Receptor for Activated C Kinase 1 (RACK1) is the best studied member of the conserved tryptophan-aspartate (WD) repeat family. RACK1 folds in a seven-bladed ß-propeller structure and it could be activated during stress, participating in different signaling pathways. The presence and activities of RACK1 in mosquitoes had not been documented before, in this work the molecule is demonstrated in an Aedes albopictus-derived cell line and its reaction to stress is observed under the effect of serum deprivation and the presence of glucocorticoid analog dexamethasone, a chemical used to cause stress in vitro.
Assuntos
Aedes/metabolismo , Regulação da Expressão Gênica , Proteínas de Insetos/biossíntese , Receptores Citoplasmáticos e Nucleares/biossíntese , Transdução de Sinais , Animais , Linhagem CelularRESUMO
We amplified S14R protein gene cDNA of porcine, cloned it into a prokaryotic expression plasmid, and expressed it in Escherichia coli. A pair of primers was designed based on the cDNA sequence of the porcine S14R gene in GenBank. The target gene fragment from porcine liver tissue was amplified by RT-PCR. Confirmed by auto-sequencing, the target gene fragment was subcloned into an expression vector of pET28a. The pET28a-S14R construct was subsequently transformed into E. coli BL21 (DE3). This construct was verified by restriction endonuclease digestion and sequencing. Using isopropyl ß-D-1-thiogalactopyranoside induction, a new recombinant protein with the expected relative molecular mass of 24 kDa appeared. The result was identified by SDS-PAGE electrophoresis. Porcine S14R includes 549bp (GenBank No. JN793537), with an open reading frame of 549 bp coding 182 amino acids.
Assuntos
Receptores Citoplasmáticos e Nucleares/genética , Sus scrofa/genética , Sequência de Aminoácidos , Animais , Clonagem Molecular , Escherichia coli , Expressão Gênica , Dados de Sequência Molecular , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores Citoplasmáticos e Nucleares/química , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
OBJECTIVE: The purpose of this study was to determine the expression of receptor activator of NFkappaB ligand (RANKL) and osteoprotegerin (OPG) associated with bone destruction in periapical cysts and granulomas. STUDY DESIGN: Forty human dental chronic periapical lesions were collected after periapical surgery. The lesions collected were fixed in 10% buffered formalin and histologically processed. At least 2 sections of each specimen were stained with hematoxylin and eosin for microscopic diagnosis. After that, 10 human periapical granulomas and 10 cysts were selected for immunohistochemical analysis for RANKL, OPG, and CD68+. RESULTS: Polymorphonuclear neutrophils, macrophages, endothelial cells, and lymphocytes were stained for RANKL and OPG in both lesions. Epithelial cells were also stained for RANKL and OPG in periapical cysts. Quantitative analysis was conducted and the results were expressed as a ratio of the number of immunostained cells over the total number of cells in the field (n = 100). The ratio of RANKL+/total cells was higher than OPG+/total cells in periapical granulomas (0.553 +/- 0.153 and 0.483 +/- 0.189, respectively; P < .0012; paired t test) and in cysts (0.519 +/- 0.09 and 0.339 +/- 0.117, respectively; P < .0001; paired t test). The ratios of OPG+/total cells (P < .0001; paired t test) and RANKL+/total cells (P < .0322; paired t test) were greater in granulomas than in cysts. However, the ratio RANKL+/OPG+ in granulomas (1.336 +/- 0.723) and cysts (1.404 +/- 0.385) was not significantly different. The ratio of CD68+/total cells was significantly higher in granulomas (0.381 +/- 0.040) than in cysts (0.307 +/- 0.068) (P < .0001; unpaired t test with Welch correction). CONCLUSION: Taking into account the limitations of the experimental approach employed, our findings indicate the presence of RANKL and OPG in cysts and granulomas, strongly suggesting the involvement of these gene products in the development of periapical lesions.
Assuntos
Proteínas de Transporte/biossíntese , Glicoproteínas/biossíntese , Glicoproteínas de Membrana/biossíntese , Granuloma Periapical/metabolismo , Cisto Radicular/metabolismo , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores do Fator de Necrose Tumoral/biossíntese , Adolescente , Adulto , Perda do Osso Alveolar/metabolismo , Antígenos CD/biossíntese , Antígenos de Diferenciação Mielomonocítica/biossíntese , Células Epiteliais/metabolismo , Humanos , Técnicas Imunoenzimáticas , Macrófagos/metabolismo , Pessoa de Meia-Idade , Osteoprotegerina , Ligante RANK , Receptor Ativador de Fator Nuclear kappa-BRESUMO
Several lines of evidence indicate that increases in nuclear Ca(2+) have specific biological effects that differ from those of cytosolic Ca(2+), suggesting that they occur independently. The mechanisms involved in controlling nuclear Ca(2+) signaling are both controversial and still poorly understood. Using hypotonic shock combined with mechanical disruption, we obtained and characterized a fraction of purified nuclei from cultured rat skeletal myotubes. Both immunoblot studies and radiolabeled inositol 1,4,5-trisphosphate [IP(3)] binding revealed an important concentration of IP(3) receptors in the nuclear fraction. Immunofluorescence and immunoelectron microscopy studies localized type-1 and type-3 IP(3) receptors in the nucleus with type-1 receptors preferentially localized in the inner nuclear membrane. Type-2 IP(3) receptor was confined to the sarcoplasmic reticulum. Isolated nuclei responded to IP(3) with rapid and transient Ca(2+) concentration elevations, which were inhibited by known blockers of IP(3) signals. Similar results were obtained with isolated nuclei from the 1B5 cell line, which does not express ryanodine receptors but releases nuclear Ca(2+) in an IP(3)-dependent manner. Nuclear Ca(2+) increases triggered by IP(3) evoked phosphorylation of cAMP response element binding protein with kinetics compatible with sequential activation. These results support the idea that Ca(2+) signals, mediated by nuclear IP(3) receptors in muscle cells, are part of a distinct Ca(2+) release component that originates in the nucleus and probably participates in gene regulation mediated by cAMP response element binding protein.
Assuntos
Canais de Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Músculo Esquelético/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Animais , Western Blotting , Canais de Cálcio/biossíntese , Canais de Cálcio/genética , Núcleo Celular/metabolismo , Células Cultivadas , Fluorometria , Imuno-Histoquímica , Inositol 1,4,5-Trifosfato/metabolismo , Receptores de Inositol 1,4,5-Trifosfato , Microscopia Confocal , Microscopia Imunoeletrônica , Músculo Esquelético/citologia , Músculo Esquelético/efeitos dos fármacos , Membrana Nuclear/metabolismo , Fosforilação , Ligação Proteica , Isoformas de Proteínas , Ratos , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores Citoplasmáticos e Nucleares/genéticaRESUMO
NOR-1 is an orphan member of the nuclear receptor superfamily, which includes a group of transcription factors involved in the response to steroids, fatty acids, retinoic acids, and other lipophilic molecules. The NOR-1 subfamily (NR4), composed also of Nurr1 and Nurr77, has been implicated in cell proliferation, differentiation, apoptosis, chondrosarcomas, inflammation, and atherogenesis. The NOR-1 receptor is an orphan ligand receptor which acts over gene transactivation. No ligands, if such in fact exist, are known for this receptor. Recently, the three-dimensional structure of the homolog receptor Nurr1 has been solved using protein crystallography techniques. Surprisingly, the structure does not present either a typical cavity for ligand binding or a classical co-factor binding site in the ligand binding domain (LBD). To allow for structural studies of other members of NR4 subfamily, we have subcloned, overexpressed in Escherichia coli cells, purified, and characterized the rat orphan nuclear receptor NOR-1 LBD domain. We obtained NOR-1 LBD at a high degree of purity and with an overall yield of 3 mg/L of culture media. CD spectroscopic analysis shows a high alpha-helical secondary structure content (52%), similar to that of Nurr 1 LBD three-dimensional structure. Thermal denaturation monitored by UV absorption and CD spectroscopy suggests proper folding of recombinant NOR-1 LBD.
Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/isolamento & purificação , Receptores Citoplasmáticos e Nucleares/química , Receptores Citoplasmáticos e Nucleares/isolamento & purificação , Receptores de Esteroides/química , Receptores de Esteroides/isolamento & purificação , Fatores de Transcrição/química , Fatores de Transcrição/isolamento & purificação , Sequência de Aminoácidos , Animais , Sítios de Ligação , Dicroísmo Circular , Proteínas de Ligação a DNA/biossíntese , Eletroforese em Gel de Poliacrilamida , Escherichia coli/metabolismo , Vetores Genéticos , Ligantes , Espectrometria de Massas , Dados de Sequência Molecular , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Estrutura Terciária de Proteína , Ratos , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores de Esteroides/biossíntese , Proteínas Recombinantes/química , Análise de Sequência de Proteína , Fatores de Tempo , Fatores de Transcrição/biossíntese , Raios UltravioletaRESUMO
In this study, we investigated the cellular and molecular events involved in parity-related alterations in mammary gland (MG) proliferation and differentiation. Rat MGs were removed on day 9 of either first (nulliparous), second (primiparous) or third (multiparous) pregnancy. Expression of steroid hormone receptors along with cellular biomarkers of proliferation and differentiation were quantified in all MG tissue compartments by immunohistochemistry. Wnt-4 (a Wingless-like morphogenic gene involved in MG development), ERbeta and ERbeta2 mRNA were evaluated by RT-PCR analysis. Serum levels of mammotrophic hormones were measured. In comparison to nulliparous and primiparous rats, multiparous animals exhibited decreased luminal cell proliferation and PR levels, whereas alpha-lactalbumin, ERalpha, ERbeta and ERbeta2 expression were increased. In myoepithelial cells, while parity induced a decrease in proliferative activity, subsequent pregnancies and lactations lead to an increased state of differentiation. Our results showed that at least two periods of pregnancy and lactation were necessary to modify the studied parameters. The lower proliferative activity and higher differentiation state of the multiparous MG are associated with both a decreased PR expression and increased ERalpha and ERbeta expression. Since ERbeta and/or ERbeta2 isoform expression was related to parity history, results suggest that the decreased proliferative activity and PR expression observed in the MG of multiparous animals may be associated with overexpression of ERbeta and/or the ERbeta2 isoform, thereby antagonizing the proliferative effects associated with ERalpha.
Assuntos
Glândulas Mamárias Animais/metabolismo , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores de Estrogênio/biossíntese , Animais , Diferenciação Celular , Divisão Celular , Receptor beta de Estrogênio , Estrogênios/metabolismo , Feminino , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Lactalbumina/biossíntese , Paridade , Fenótipo , Reação em Cadeia da Polimerase , Gravidez , Prenhez , Isoformas de Proteínas , Proteínas Proto-Oncogênicas/biossíntese , RNA/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Receptores de Progesterona/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Proteínas Wnt , Proteína Wnt4RESUMO
The mechanisms involved in the increase of orbital retro-ocular adipose tissue that occurs in Graves' ophthalmopathy (GO) are still unclear. In this condition, the orbital tissue shows glycosaminoglycans deposition produced by activated fibroblasts capable of undergoing adipocytic differentiation. Many genes are involved in adipogenic mechanisms including the transcription factor peroxisome proliferator-activated receptor-gamma (PPAR-gamma). We evaluated the level of expression of the PPAR-gamma gene in normal and GO orbital adipose/connective tissue specimens using a quantitative and sensitive reverse transcription (RT) competitive polymerase chain reaction (PCR) assay. Our results show that the expression of PPAR-gamma was significantly greater in adipose/connective tissue from patients in the active stage of GO than in controls (150.8 +/- 103.9 and 24.0 +/- 4.9 amol/micro g of total RNA respectively, p < 0.05), while there was no significant difference between patients with inactive GO (58.8 +/- 40.6 aM/microg total RNA) and controls. These results suggest that increased PPAR-gamma gene expression in the active stage of GO may be dependent on the inflammatory process in this disease. We speculate that the increased orbital fat tissue observed in GO may be a consequence of the anti-inflammatory PPAR-gamma action.
Assuntos
Tecido Adiposo/metabolismo , Tecido Conjuntivo/metabolismo , Doença de Graves/metabolismo , Doença de Graves/patologia , Órbita/metabolismo , Receptores Citoplasmáticos e Nucleares/biossíntese , Fatores de Transcrição/biossíntese , Tecido Adiposo/crescimento & desenvolvimento , Tecido Adiposo/patologia , Adulto , Estudos de Casos e Controles , Feminino , Expressão Gênica , Doença de Graves/genética , Doença de Graves/terapia , Humanos , Masculino , Pessoa de Meia-Idade , Órbita/crescimento & desenvolvimento , Órbita/patologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores Citoplasmáticos e Nucleares/genética , Fatores de Transcrição/genéticaRESUMO
During gestation, the perinatal neuroendocrine axis keeps clock time. In primates, the suprachiasmatic nucleus (biological clock in mammals), shows oscillatory function by midgestation. There is evidence in rodents that the mother, during pregnancy, entrains the fetal suprachiasmatic nucleus (SCN) and newborn circadian rhythms. We are investigating the role of maternal melatonin as an entraining signal for the newborn circadian time-keeping system in the Cebus apella (New World non-human primate). Twenty-four hour rhythms of temperature and cortisol are present in the 4 days old C. apella newborn. Preliminary data suggests that inhibition of maternal melatonin by exposing pregnant females to constant light alters these rhythms. We have found binding sites for melatonin and expression of mRNA for Mel 1A receptor in hypothalamus, kidney and testis. These preliminary results suggest that maternal melatonin may play a role in relating the perinatal circadian time-keeping system to environmental signals.
Assuntos
Animais Recém-Nascidos/fisiologia , Cebus/fisiologia , Ritmo Circadiano/fisiologia , Feto/fisiologia , Recém-Nascido/fisiologia , Melatonina/fisiologia , Animais , Regulação da Temperatura Corporal/fisiologia , Cebus/embriologia , Ritmo Circadiano/efeitos da radiação , Feminino , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Hidrocortisona/metabolismo , Masculino , Mamíferos/embriologia , Mamíferos/fisiologia , Troca Materno-Fetal , Neuropeptídeos/fisiologia , Gravidez , Receptores de Superfície Celular/biossíntese , Receptores de Superfície Celular/genética , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores Citoplasmáticos e Nucleares/genética , Receptores de Melatonina , Retina/fisiologia , Retina/efeitos da radiação , Núcleo Supraquiasmático/embriologia , Núcleo Supraquiasmático/fisiologiaRESUMO
Two novel cucurbitacins designated as neocucurbitacins A (1), possessing inhibitory activity of polyoma enhancer binding protein 2alphaA (PEBP2alphaA) and osteoclastogenesis-inhibitory factor (OCIF) gene expression in human osteoblast-like cells, and B (2) were isolated from the fruit of Luffa operculata. Their structures have been determined by extensive spectroscopic investigation.
Assuntos
Cucurbitaceae/química , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Glicoproteínas/biossíntese , Glicoproteínas/genética , Luffa/química , Osteoblastos/metabolismo , Proteínas Proto-Oncogênicas , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores Citoplasmáticos e Nucleares/genética , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética , Linhagem Celular , Subunidade alfa 2 de Fator de Ligação ao Core , Humanos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Osteoprotegerina , Extratos Vegetais/análise , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores do Fator de Necrose Tumoral , Espectrometria de Massas de Bombardeamento Rápido de Átomos , Espectrofotometria InfravermelhoRESUMO
We have characterized the biochemical properties of a 66-kDa poly(A)-binding protein (PABP1) in the protozoan Trypanosoma cruzi and isolated two classes of cDNAs encoding the protein. In concordance, Southern blots showed the presence of 2 gene copies. The two cDNA classes differ in the length of adenosine-rich segments in the 5' untranslated region and in point changes scattered throughout the sequence, but their 1650-bp open reading frames encode identical proteins. A single mRNA of 5.5 kb was detected, indicating that the noncoding regions are unusually long. Both the mRNA and the protein are constitutively expressed in all stages of T. cruzi life cycle. The biochemical properties and sequence comparisons show that the T. cruzi PABP1 is similar to the PABP1 of other eukaryotic organisms. These results indicate that PABP1 has been conserved throughout eukaryotic evolution.