RESUMEN
Somatic inactivating mutations in epigenetic regulators are frequently found in combination in myelodysplastic syndrome (MDS). However, the mechanisms by which combinatory mutations in epigenetic regulators promote the development of MDS remain unknown. Here we performed epigenomic profiling of hematopoietic progenitors in MDS mice hypomorphic for Tet2 following the loss of the polycomb-group gene Ezh2 (Tet2KD/KDEzh2Δ/Δ). Aberrant DNA methylation propagated in a sequential manner from a Tet2-insufficient state to advanced MDS with deletion of Ezh2. Hyper-differentially methylated regions (hyper-DMRs) in Tet2KD/KDEzh2Δ/Δ MDS hematopoietic stem/progenitor cells were largely distinct from those in each single mutant and correlated with transcriptional repression. Although Tet2 hypomorph was responsible for enhancer hypermethylation, the loss of Ezh2 induced hyper-DMRs that were enriched for CpG islands of polycomb targets. Notably, Ezh2 targets largely lost the H3K27me3 mark while acquiring a significantly higher level of DNA methylation than Ezh1 targets that retained the mark. These findings indicate that Ezh2 targets are the major targets of the epigenetic switch in MDS with Ezh2 insufficiency. Our results provide a detailed trail for the epigenetic drift in a well-defined MDS model and demonstrate that the combined dysfunction of epigenetic regulators cooperatively remodels the epigenome in the pathogenesis of MDS.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Epigénesis Genética , Regulación de la Expresión Génica , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Animales , Secuencia de Bases , Sitios de Unión , Islas de CpG , Metilación de ADN , Proteínas de Unión al ADN/genética , Dioxigenasas , Modelos Animales de Enfermedad , Elementos de Facilitación Genéticos , Proteína Potenciadora del Homólogo Zeste 2/genética , Hematopoyesis/genética , Ratones , Ratones Noqueados , Ratones Transgénicos , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Motivos de Nucleótidos , Unión Proteica , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/metabolismo , Factores de Transcripción/metabolismoRESUMEN
We have determined the genomic organization of two closely related phosphoenolpyruvate carboxylase genes in soybean, GmPEPC7, which is expressed at high levels in root nodules, and the housekeeping gene GmPEPC15. Their nucleotide sequences, including most introns and 5;-flanking regions within 600 bp upstream from the transcription start sites, are well conserved, suggesting that they were duplicated quite recently. To gain insights into the process of evolution of the tissue-specifically expressed GmPEPC7gene, we produced chimeric constructs carrying either the GmPEPC7or GmPEPC15promoter fused to the beta-glucuronidase gene. The expression patterns of the reporter observed in nodules that developed on transgenic hairy roots reflected the levels of mRNA levels produced by the genes in wild-type soybean plants, indicating that the GmPEPC7promoter directs nodule-specific expression. Loss-of-function experiments showed that the segment of GmPEPC7between -466 and -400, designated as the "switch region" (SR), was necessary for expression in nodules, although proteins that bind to SR were not detectable in a gel-retardation assay. Another gel-retardation assay indicated that putative nodule nuclear proteins bind specifically to the region of GmPEPC7between -400 and -318, designated as the "amplifier region" (AR). Both SR and AR have characteristic sequences that are not found in the GmPEPC15promoter. Furthermore, experiments using hybrid promoters derived from GmPEPC15demonstrated that AR confers high-level expression in nodules only in combination with SR. When wild-type soybean plants were subjected to prolonged darkness and subsequently illuminated, the level of GmPEPC7mRNA in nodules decreased and then recovered. This study suggests that the acquisition of two interdependent cis-acting elements resulted in molecular evolution of the nodule-enhanced GmPEPC7gene.
Asunto(s)
Regulación Enzimológica de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Glycine max/enzimología , Fosfoenolpiruvato Carboxilasa/biosíntesis , Fosfoenolpiruvato Carboxilasa/genética , Secuencia de Bases , Northern Blotting , Núcleo Celular/metabolismo , Glucuronidasa/genética , Modelos Genéticos , Datos de Secuencia Molecular , Raíces de Plantas , Regiones Promotoras Genéticas , ARN Mensajero/metabolismo , Proteínas Recombinantes de Fusión/metabolismo , Homología de Secuencia de Ácido NucleicoRESUMEN
Recent advances in human islet transplantation have highlighted the need for expanding the pool of beta-cells available for transplantation. We have developed three transgenic models in which growth factors (hepatocyte growth factor [HGF], placental lactogen, or parathyroid hormone-related protein) have been targeted to the beta-cell using rat insulin promoter (RIP). Each displays an increase in islet size and islet number, and each displays insulin-mediated hypoglycemia. Of these three models, the RIP-HGF mouse displays the least impressive phenotype under basal conditions. In this study, we show that this mild basal phenotype is misleading and that RIP-HGF mice have a unique and salutary phenotype. Compared with normal islets, RIP-HGF islets contain more insulin per beta-cell (50 +/- 5 vs. 78 +/- 9 ng/islet equivalent [IE] in normal vs. RIP-HGF islets, P < 0.025), secrete more insulin in response to glucose in vivo (0.66 +/- 0.06 vs. 0.91 +/- 0.10 ng/ml in normal vs. RIP-HGF mice, P < 0.05) and in vitro (at 22.2 mmol/l glucose: 640 +/- 120.1 vs. 1,615 +/- 196.9 pg. microg protein(-1). 30 min(-1) in normal vs. RIP-HGF islets, P < 0.01), have two- to threefold higher GLUT2 and glucokinase steady-state mRNA levels, take up and metabolize glucose more effectively, and most importantly, function at least twice as effectively after transplantation. These findings indicate that HGF has surprisingly positive effects on beta-cell mitogenesis, glucose sensing, beta-cell markers of differentiation, and transplant survival. It appears to have a unique and unanticipated effective profile as an islet mass- and function-enhancing agent in vivo.
Asunto(s)
Expresión Génica , Supervivencia de Injerto , Factor de Crecimiento de Hepatocito/genética , Trasplante de Islotes Pancreáticos , Islotes Pancreáticos/fisiología , Animales , Marcación de Gen , Glucoquinasa/genética , Prueba de Tolerancia a la Glucosa , Transportador de Glucosa de Tipo 2 , Factor de Crecimiento de Hepatocito/fisiología , Insulina/genética , Islotes Pancreáticos/química , Cinética , Ratones , Ratones Transgénicos , Modelos Animales , Proteínas de Transporte de Monosacáridos/genética , Proteína Relacionada con la Hormona Paratiroidea , Lactógeno Placentario/genética , Regiones Promotoras Genéticas , Proteínas/genética , ARN Mensajero/análisisRESUMEN
The cloning of the so-called 'parathyroid hormone-related protein' (PTHrP) in 1987 was the result of a long quest for the factor which, by mimicking the actions of PTH in bone and kidney, is responsible for the hypercalcemic paraneoplastic syndrome, humoral calcemia of malignancy. PTHrP is distinct from PTH in a number of ways. First, PTHrP is the product of a separate gene. Second, with the exception of a short N-terminal region, the structure of PTHrP is not closely related to that of PTH. Third, in contrast to PTH, PTHrP is a paracrine factor expressed throughout the body. Finally, most of the functions of PTHrP have nothing in common with those of PTH. PTHrP is a poly-hormone which comprises a family of distinct peptide hormones arising from post-translational endoproteolytic cleavage of the initial PTHrP translation products. Mature N-terminal, mid-region and C-terminal secretory forms of PTHrP are thus generated, each of them having their own physiologic functions and probably their own receptors. The type 1 PTHrP receptor, binding both PTH(1-34) and PTHrP(1-36), is the only cloned receptor so far. PTHrP is a PTH-like calciotropic hormone, a myorelaxant, a growth factor and a developmental regulatory molecule. The present review reports recent aspects of PTHrP pharmacology and physiology, including: (a) the identification of new peptides and receptors of the PTH/PTHrP system; (b) the recently discovered nuclear functions of PTHrP and the role of PTHrP as an intracrine regulator of cell growth and cell death; (c) the physiological and developmental actions of PTHrP in the cardiovascular and the renal glomerulo-vascular systems; (d) the role of PTHrP as a regulator of pancreatic beta cell growth and functions, and, (e) the interactions of PTHrP and calcium-sensing receptors for the control of the growth of placental trophoblasts. These new advances have contributed to a better understanding of the pathophysiological role of PTHrP, and will help to identify its therapeutic potential in a number of diseases.
Asunto(s)
Islotes Pancreáticos/metabolismo , Hormona Paratiroidea/fisiología , Proteínas/fisiología , Receptores de Hormona Paratiroidea/fisiología , Animales , Apoptosis , Proteínas de Unión al Calcio/metabolismo , Sistema Cardiovascular/metabolismo , Núcleo Celular/metabolismo , Femenino , Humanos , Riñón/metabolismo , Ratones , Señales de Localización Nuclear , Hormona Paratiroidea/genética , Proteína Relacionada con la Hormona Paratiroidea , Placenta/metabolismo , Embarazo , Proteínas/genética , Ratas , Receptores de Hormona Paratiroidea/genética , Trofoblastos/metabolismoRESUMEN
PTHrP is secreted by most cell types. In addition to a paracrine/autocrine role, PTHrP has "intracrine" actions, entering the nuclear compartment under the direction of a classic bipartite nuclear localization signal. In vascular smooth muscle cells, nuclear entry stimulates mitogenesis. In the current study, we sought to more precisely define the regions of PTHrP required for the activation of mitogenesis in vascular smooth muscle cells. PTHrP deletion mutants missing large regions [i.e. the signal peptide, N terminus (1--36), mid region (38--86), nuclear localization signal, C terminus (108--139), or combinations of the above] were expressed in A-10 vascular smooth muscle cells. The consequences on nuclear localization and proliferation were examined. Deletion of the nuclear localization signal prevented nuclear entry and slowed proliferation. Deletion of the highly conserved N terminus or mid region had no impact on nuclear localization or on proliferation. Deletion of the C terminus had no deleterious effect on nuclear localization but dramatically reduced proliferation. Thus, the nuclear localization signal is both necessary and sufficient for nuclear localization of PTHrP. In contrast, activation of proliferation in vascular smooth muscle cells requires both an intact nuclear localization signal and an intact C terminus. Whereas the nuclear localization signal is required for nuclear entry, the C terminus may serve a trans-activating function to stimulate mitogenesis once inside the nucleus of vascular smooth muscle cells.
Asunto(s)
Músculo Liso Vascular/citología , Músculo Liso Vascular/metabolismo , Señales de Localización Nuclear/fisiología , Proteínas/química , Proteínas/fisiología , Secuencia de Aminoácidos/genética , Animales , División Celular/fisiología , Línea Celular , Núcleo Celular/metabolismo , Eliminación de Gen , Datos de Secuencia Molecular , Mutación/fisiología , Proteína Relacionada con la Hormona Paratiroidea , Proteínas/genética , Ratas , Relación Estructura-Actividad , Distribución TisularRESUMEN
This is a particularly exciting time in the field of pancreatic islet growth, development, and survival. The recent publication of a study demonstrating that human pancreatic islet transplantation is both technically and immunologically feasible has highlighted the need for large supplies of pancreatic islets or pancreatic beta cells for larger-scale islet transplantation in patients with diabetes. This, together with a rapid expansion in the past several years of the understanding of mechanisms of islet growth, development, and survival, has accelerated and invigorated efforts to therapeutically harness the cellular mechanisms responsible for pancreatic beta-cell proliferation, survival, and development and to take advantage of this new knowledge to enhance the availability, survival, and function of pancreatic beta cells in human islet transplantation for diabetes mellitus. Here, we briefly review the confluence of events that have provided optimism and energy to the islet transplant field, and we focus on peptide growth factors that eventually may be deployed in the effort to augment islet mass and function in patients with diabetes.
Asunto(s)
Sustancias de Crecimiento/uso terapéutico , Trasplante de Islotes Pancreáticos , Islotes Pancreáticos/fisiología , Animales , División Celular , Factor de Crecimiento de Hepatocito/uso terapéutico , Humanos , Islotes Pancreáticos/citología , Proteína Relacionada con la Hormona Paratiroidea , Lactógeno Placentario/uso terapéutico , Proteínas/uso terapéutico , Somatomedinas/uso terapéuticoRESUMEN
Uricase (nodulin-35) cDNA, LjUr, was isolated from nodules of a model legume, Lotus japonicus. LjUr expression was most abundant in nodules, although it was detected in nonsymbiotic tissues as well, particularly in roots. Expression in nodules was detected in uninfected cells, nodule parenchyma, and, more intensely, in vascular bundles. Phylogenetic analysis of uricase sequences from various legumes indicated that uricases of amide- and ureide-transporting legumes form two distinct clades. LjUr is in the cluster of amide-transport legumes even though L. japonicus bears determinate nodules.
Asunto(s)
Fabaceae/enzimología , Fabaceae/genética , Plantas Medicinales , Urato Oxidasa/genética , Secuencia de Aminoácidos , Exones , Fabaceae/clasificación , Expresión Génica , Biblioteca de Genes , Genes de Plantas , Datos de Secuencia Molecular , Peroxisomas/enzimología , Filogenia , Raíces de Plantas/enzimología , Estructuras de las Plantas/enzimología , ARN Mensajero/análisis , ARN Mensajero/genética , ARN de Planta/análisis , ARN de Planta/genética , Urato Oxidasa/químicaRESUMEN
Two cDNA clones, MsU2 and MsU9 encoding uricase (EC 1.7.3.3, Nodulin-35) were isolated from a cDNA library prepared from nodule tissues of alfalfa, Medicago sativa, plants. Both MsU2 and MsU9 encoded 308 amino acid polypeptides with a difference of 5 amino acids, and the deduced amino acid sequences shared 98% homology. Between these two cDNA clones and uricase genes of soybean which were designated as Nod-35s, more than 80% identity was observed in nucleotides and deduced amino acid sequences, suggesting that these MsU2 and MsU9 are homologs of Nod-35. Using the reverse transcription-PCR technique, we detected the transcripts of these two genes in almost all tissues of alfalfa. The operation of uricase genes was confirmed by the presence of ureide in the xylem sap and uricase activity in the nodules. In situ hybridization analysis revealed that MsU2 and MsU9 were expressed only in uninfected cells of the infected zone of the nodule tissue. The cell specific-expression of the two uricase genes was observed in an identical manner to that of Nod-35 in soybean nodules.
Asunto(s)
Medicago sativa/genética , Proteínas de la Membrana , Proteínas de Plantas/genética , Urato Oxidasa/genética , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Southern Blotting , ADN Complementario/química , ADN Complementario/genética , ADN Complementario/aislamiento & purificación , ADN de Plantas/genética , Regulación Enzimológica de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Hibridación in Situ , Isoenzimas/genética , Medicago sativa/enzimología , Datos de Secuencia Molecular , Raíces de Plantas/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Distribución Tisular , Urea/análogos & derivados , Urea/metabolismoRESUMEN
Hepatocyte growth factor (HGF) is produced in pancreatic mesenchyme-derived cells and in islet cells. In vitro, HGF increases the insulin content and proliferation of islets. To study the role of HGF in the islet in vivo, we have developed three lines of transgenic mice overexpressing mHGF using the rat insulin II promoter (RIP). Each RIP-HGF transgenic line displays clear expression of HGF mRNA and protein in the islet. RIP-mHGF mice are relatively hypoglycemic in post-prandial and fasting states compared with their normal littermates. They display inappropriate insulin production, striking overexpression of insulin mRNA in the islet, and a 2-fold increase in the insulin content in islet extracts. Importantly, beta cell replication rates in vivo are two to three times higher in RIP-HGF mice. This increase in proliferation results in a 2-3-fold increase in islet mass. Moreover, the islet number per pancreatic area was also increased by approximately 50%. Finally, RIP-mHGF mice show a dramatically attenuated response to the diabetogenic effects of streptozotocin. We conclude that the overexpression of HGF in the islet increases beta cell proliferation, islet number, beta cell mass, and total insulin production in vivo. These combined effects result in mild hypoglycemia and resistance to the diabetogenic effects of streptozotocin.
Asunto(s)
Factor de Crecimiento de Hepatocito/genética , Hipoglucemia/fisiopatología , Insulina/genética , Islotes Pancreáticos/citología , Islotes Pancreáticos/fisiología , Regiones Promotoras Genéticas , Animales , Glucemia/metabolismo , División Celular , Ayuno , Glucagón/análisis , Factor de Crecimiento de Hepatocito/análisis , Factor de Crecimiento de Hepatocito/fisiología , Hipoglucemia/etiología , Insulina/análisis , Islotes Pancreáticos/fisiopatología , Ratones , Ratones Transgénicos , Tamaño de los Órganos , Polipéptido Pancreático/análisis , Ratas , Somatostatina/análisisRESUMEN
The early nodulin ENOD40 has been proposed as playing a pivotal role in the organogenesis of legume root nodules. We have isolated the ENOD40 gene homologues ObENOD40 and OsENOD40 from the wild and cultivated rice genotypes Oryza brachyantha and Oryza sativa, respectively. Rice ENOD40s contain a sequence at the 5' end (region I) for encoding an oligopeptide that is highly conserved in all legume ENOD40s. Furthermore, at the 3' end (region II), the nucleotide sequence of rice ENOD40s exhibited a considerable homology to the corresponding region in legume ENOD40s. Among various organs of the rice plant, expression of OsENOD40 was detected only in stems. In situ hybridization studies revealed that, within the stem, transcription of OsENOD40 is confined to parenchyma cells surrounding the protoxylem during the early stages of development of lateral vascular bundles that conjoin an emerging leaf. Expression pattern of OsENOD40 promoter-GUS fusion in nodules developed on transgenic hairy roots of soybean was also found to be restricted to peripheral cells of nodule vascular bundles, thus evidencing that rice ENOD40 promoter activity is essentially the same as that of soybean ENOD40. Taken together, these results strongly suggest that OsENOD40 and legume ENOD40s share common, if not identical, functions in differentiation and/or function of vascular bundles.
Asunto(s)
Glycine max/genética , Oryza/genética , Proteínas de Plantas/genética , ARN no Traducido/genética , Secuencia de Aminoácidos , Secuencia de Bases , ADN Complementario , Datos de Secuencia Molecular , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas , ARN Largo no Codificante , ARN Mensajero/genética , Homología de Secuencia de AminoácidoRESUMEN
In vitro and in vivo studies revealed that Malaysian medicinal plants, Piper sarmentosum, Andrographis paniculata and Tinospora crispa produced considerable antimalarial effects. Chloroform extract in vitro did show better effect than the methanol extract. The chloroform extract showed complete parasite growth inhibition as low as 0.05 mg/ml drug dose within 24 h incubation period (Andrographis paniculata) as compared to methanol extract of drug dose of 2.5 mg/ml but under incubation time of 48 h of the same plant spesies. In vivo activity of Andrographis paniculata also demonstrated higher antimalarial effect than other two plant species.
Asunto(s)
Antimaláricos/farmacología , Extractos Vegetales/farmacología , Plantas Medicinales/química , Plasmodium berghei/efectos de los fármacos , Plasmodium falciparum/efectos de los fármacos , Animales , Femenino , Técnicas In Vitro , Malasia , Ratones , SolventesRESUMEN
Nodule-specific uricase (uricase II) is a homotetramer of a 33-kDa polypeptide, nodulin 35, and plays a key role in the assimilation of nitrogen fixed by microsymbionts in most legumes that have determinate nodules. We have isolated two distinct genes, UR2 and UR9, that encode for nodulin 35 from a soybean genomic library. Their corresponding cDNAs were also isolated from a nodule cDNA library. UR2 and UR9 both encode for 309 amino acid proteins with 12 amino acid differences. The expression of these two genes in various organs of soybean was examined by reverse transcription-polymerase chain reaction with primers specific to each cDNA sequences. Expression of UR9 was almost specific in root nodules, although it was expressed in roots, primary leaves, and developing seed at very low levels. In contrast, the UR2 transcripts were present in almost all plant organs at low levels, but no enhancement of the expression was observed in nodules. Thus, UR9 behaves as a nodulin gene, whereas UR2 is a nonsymbiotic uricase II gene. The sequences of their potential promoter regions share high homology within regions up to about 400 bp upstream from the translation initiation sites. These results suggest that symbiotic and nonsymbiotic uricase II genes diverged by gene duplication and that relatively small alterations in the promoter sequence enable the nodule-specific expression.
Asunto(s)
Genes de Plantas , Glycine max/enzimología , Proteínas de la Membrana , Proteínas de Plantas/genética , Urato Oxidasa/genética , Secuencia de Aminoácidos , Secuencia de Bases , ADN Complementario/genética , Expresión Génica , Biblioteca Genómica , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Simbiosis , Distribución TisularRESUMEN
A cDNA clone (URcot-35) was isolated from a soybean cotyledonary cDNA library using a cDNA clone (URnod-35) for nodule uricase II as a probe. URcot-35 was a 1,170-bp cDNA with an open reading frame that encoded a protein of putative 309 amino acids with a molecular mass of 35,137 Da. The nucleotide sequence of URcot-35 was identical to that of URnod-35. Expression of the URcot-35 gene in cotyledons was investigated by Northern dot-blot hybridization, by the reverse transcription-polymerase chain reaction with subsequent hybridization assays with the cDNA for nodule uricase II as a probe, and by immunoblotting analysis with a monoclonal antibody that was specific to nodule uricase II. The results suggested that the transcript of URcot-35 was present in developing cotyledons and that uricase II accumulated during the pod-filling stage. This is the first report of the isolation of cDNA for uricase II from non-symbiotic tissue and the results demonstrate that uricase II in soybean cotyledons is identical to that in soybean nodules.
Asunto(s)
Genes de Plantas , Glycine max/enzimología , Glycine max/genética , Urato Oxidasa/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Cartilla de ADN/genética , ADN Complementario/genética , ADN de Plantas/genética , Expresión Génica , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , ARN Mensajero/genética , ARN Mensajero/metabolismo , Homología de Secuencia de Ácido Nucleico , Glycine max/crecimiento & desarrolloRESUMEN
Immunoblot analysis showed that uricases in non-ureide-transporting determinate nodules (Canavalia gladiata and Lotus japonicus) did not react with a monoclonal antibody against soybean nodule uricase, suggesting different immunological reactivities from those of uricases of ureide-transporting legumes.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Expresión Génica , Glycine max/enzimología , Proteínas de Plantas/inmunología , Urato Oxidasa/inmunología , Animales , Fabaceae/metabolismo , Immunoblotting , Ratones , Ratones Endogámicos BALB C , Plantas Medicinales , Células Tumorales Cultivadas , Urato Oxidasa/genéticaRESUMEN
Although androgen receptor levels vary widely during development, our previous studies have suggested that a single promoter is active in a number of human and rat tissues and cell lines. To examine the elements controlling androgen receptor expression, we performed deletion mapping and site-directed mutagenesis of the human androgen receptor promoter and assayed these promoter fusions in human cell lines that produce the androgen receptor as well as those deficient in the androgen receptor, both in the presence and absence of androgen. Our studies identify a region (-74 to +87) surrounding the site of transcription initiation that constitutes the core of the androgen receptor promoter. When assayed in T47D cells (an AR-expressing cell line), this segment was sufficient to direct the expression of reporter genes at levels similar to that observed for larger promoter fragments, and further deletions led to an appreciable loss of promoter activity. DNA sequences surrounding this region appear to modulate the activity of this minimal promoter in a cooperative fashion. Site-directed mutagenesis and mobility shift assays demonstrated the importance of regulatory regions upstream of the transcription initiation site including an SP1 binding site and two segments with similarities to consensus HLH protein binding sites (E-boxes). Androgen treatment did not cause a decrease in activity of any of the transiently transfected promoter fusions tested, suggesting that the promoter does not contain the information necessary for autoregulation or that a posttranscriptional or posttranslational mechanism is responsible for the regulation of AR mRNA by ligand. Furthermore, the androgen receptor promoter fusions displayed differing transcriptional activities when transfected into two cell lines deficient in androgen receptor expression suggesting heterogeneity in the mechanisms responsible for this deficiency.
Asunto(s)
Regiones Promotoras Genéticas , Receptores Androgénicos/genética , Clonación Molecular , Proteínas de Unión al ADN/metabolismo , Femenino , Secuencias Hélice-Asa-Hélice , Humanos , Masculino , Mutagénesis Sitio-Dirigida , Células Tumorales CultivadasRESUMEN
Seven hundred and forty-four newly diagnosed patients with acute leukemias between 1978 and 1990 were classified on the basis of immunological phenotypes. The majority of the patients were enrolled in the Tokyo Children's Cancer Study Group (TCCSG) studies. The incidence of subclassification of acute leukemias in this study was as follows: 522 patients with ALL (70%), 139 patients with ANLL (18%), 29 patients with biphenotypic leukemia, 8 patients with Ph1-positive acute leukemia (Ph1-AL), and 45 patients with infant leukemia. ALLs were classified into common ALL (cALL, 77%), T-ALL (15%), B-ALL (4%), and unclassified ALL (3%). The incidence of ALL subtypes in this study reflected those of TCCSG. Biphenotypic leukemias were categorized into 4 groups as follows; 1) cALL with positive myelomonocytic antigen(s) (N = 11), 2) unclassified ALL with positive myelomonocytic antigen(s) (N = 5), 3) ANLL with positive B-lymphoid antigen(s) (N = 4), and 4) acute leukemia with positive T-lymphoid and myeloid antigen(s). Infant leukemias were classified into ALL type (N = 27) and ANLL type (N = 18). In this present study, clinical features and immunological phenotypes of the acute leukemias with a poor prognosis, i.e. biphenotypic leukemia, Ph1-AL, and infant leukemia are analyzed and discussed.
Asunto(s)
Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Adolescente , Antígenos CD/análisis , Niño , Preescolar , Femenino , Antígenos HLA-A/análisis , Humanos , Lactante , Recién Nacido , Leucemia/clasificación , Leucemia/inmunología , Leucemia/mortalidad , Tablas de Vida , Masculino , Leucemia-Linfoma Linfoblástico de Células Precursoras/clasificación , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidad , PronósticoRESUMEN
Growth of the penis at sexual maturation is under the control of androgens, but growth ceases as the organ reaches adult size despite continued high levels of circulating androgen. Previous studies have shown that the cessation of penile growth is associated with a decrease in the quantity of androgen receptor, as detected by ligand binding and immunological methods. In the current studies we demonstrate that the decreased androgen receptor levels correlate with a decrease in androgen receptor mRNA content in the penile corpus and os, but not in the glans penis. These findings suggest that modulation of androgen receptor mRNA levels in the body of the penis may be important to the control of androgen-dependent growth in the tissue, and that the control of androgen receptor mRNA levels differs among the different cell types that comprise the penis. To explore the mechanisms controlling androgen receptor expression, we examined the transcription initiation site of the rat androgen receptor gene in ventral prostate and in three compartments of the penis: the corpus, the urethra, and the glans penis. The same promoter is employed in all preparations, suggesting that the different patterns of androgen receptor mRNA expression in these tissues with age are controlled by factors that modulate the activity of the same promoter.
Asunto(s)
Pene/metabolismo , ARN Mensajero/metabolismo , Receptores Androgénicos/metabolismo , Envejecimiento/metabolismo , Animales , Secuencia de Bases , Exones , Expresión Génica , Inmunohistoquímica , Masculino , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Pene/crecimiento & desarrollo , Próstata/crecimiento & desarrollo , Próstata/metabolismo , Ratas , Ratas Endogámicas , Receptores Androgénicos/genética , Endonucleasas Específicas del ADN y ARN con un Solo Filamento , Distribución Tisular , Transcripción GenéticaRESUMEN
The rat phallus grows during sexual maturation as serum androgen concentrations rise to adult levels, and growth ceases when sexual maturity is attained despite the continued presence of adult serum androgen levels. This cessation of growth is correlated temporally with a diminution in the levels of androgen receptor, as detected by assays of ligand binding. To determine whether the change in androgen receptor content occurs in all cells in the tissue, immunohistochemical studies of the penile androgen receptor were performed in rats of different ages. In immature animals and animals castrated prepubertally, the androgen receptor is detected in virtually all cell types, including the corpus cavernosum penis, the small lateral cavernous bodies, the corpus cavernosum urethra, skin, urethra, and os penis. By contrast, in the mature rat penis minimal androgen receptor is evident within the corporal tissue and os, although immunoreactivity remains detectable in the penile skin and urethra. It is concluded that the cessation of androgen-mediated growth correlates with a decrease in androgen receptor levels in the body of the penis.
Asunto(s)
Pene/metabolismo , Receptores Androgénicos/metabolismo , Envejecimiento/metabolismo , Animales , Dihidrotestosterona/farmacología , Inmunohistoquímica , Masculino , Orquiectomía , Pene/citología , Ratas , Ratas EndogámicasRESUMEN
To provide insight into the factors that control androgen receptor levels in rat penis, we assessed 5 alpha-[3H]-dihydrotestosterone binding in low-salt [10 mM tris(hydroxymethyl)aminomethane (Tris), 10 mM Na2M0O4] and high-salt (10 mM Tris, 10 mM Na2M0O4, 0.5 M KCl) extracts of rat penis using sucrose density gradients. Total receptor content decreased from approximately 729 +/- 114 fmol/g tissue at 3 wk of age to less than 50 fmol/g tissue at 10 wk of age. Castration of 3-wk-old rats prevented penile growth and the age-related decline in penile androgen receptor. Treatment of 3-wk-old castrated rats with 5 alpha-dihydrotestosterone caused an acceleration in the decline in receptor levels compared with intact animals. Castration of 10-wk-old rats (after androgen receptor levels had decreased) did not result in an increase in the amount of total androgen receptor by 16 wk of age. To determine the specificity of the androgen-mediated decline in receptor levels, the amounts of prostate androgen receptor were compared with those of the penis at different ages. When expressed as femtomoles per organ, the total androgen receptor level in the prostate increased fourfold from 3 to 10 wk of age, whereas the total androgen receptor in the penis declined approximately threefold. We conclude that the downregulation of the penile androgen receptor content that occurs in the rat between 3 and 10 wk of age is androgen mediated, does not occur in all androgen target tissues, and is prevented but not reversed by castration.
Asunto(s)
Regulación hacia Abajo/efectos de los fármacos , Pene/fisiología , Receptores Androgénicos/fisiología , Envejecimiento , Animales , Centrifugación por Gradiente de Densidad , Dihidrotestosterona/metabolismo , Dihidrotestosterona/farmacología , Hipofisectomía , Masculino , Orquiectomía , Pene/efectos de los fármacos , Pene/crecimiento & desarrollo , Próstata/crecimiento & desarrollo , Próstata/fisiología , Ratas , Ratas Endogámicas , Receptores Androgénicos/efectos de los fármacos , Receptores Androgénicos/aislamiento & purificación , Valores de ReferenciaRESUMEN
We report a 17-year-old female with chronic myeloid leukemia (CML) who developed monocytic crisis. She was diagnosed as chronic phase of Ph1-chromosome positive CML at 14 years old. Three years after the diagnosis of the disease, she was admitted to the hospital because of low grade fever, lethargy and marked splenomegaly. Small dose of Ara-C relieved her symptoms and splenomegaly. Six months later, however, a marked leukocytosis over 70,000/microliters were observed, and the peripheral blood smear disclosed that about 80% of the leukocytes were relatively mature monocytoid cells. Chromosomal analysis revealed additional abnormalities (double Ph1, +8, +9, +19). Lysozyme levels in serum and urine were high and NAP score was elevated. These monocytoid cells expressed receptors for IgG-Fc and C3, phagocytic activity, and monocytoid antigens which were determined by monoclonal antibodies (MY4, Mo2, OKM5). Cytochemically, almost all of monocytoid cells were positive for peroxidase and naphthol-ASD-chloroacetate esterase (CAE), but the monocytoid cells positive for non-specific esterase were limited. These data suggested that this case was monocytic crisis in CML with proliferation of CAE positive monocytoid cells. Among several types of blast crisis, monocytic crisis is extremely rare condition. The definite monocytic crisis demonstrated by this case may support the hypothesis that target cells of CML are pluripotent hematopoietic precursors.