RESUMEN
Distinct classes of neurons are generated at defined times and positions during development of the nervous system. It remains elusive how specification of neuronal identity coordinates with acquisition of pan-neuronal properties. Here we show that basic helix-loop-helix (bHLH) transcription factors Olig2 and Neurogenin2 (Ngn2) play vital roles in the coordinated induction of pan-neuronal and subtype-specific properties of motoneurons. Olig2 and Ngn2 are specifically coexpressed in motoneuron progenitors. Misexpression studies in chick demonstrate the specific, combinatorial actions of Olig2 and Ngn2 in motoneuron generation. Our results further revealed crossregulatory interactions between bHLH and homeodomain transcription factors in the specification of motoneurons. We suggest that distinct classes of transcription factors collaborate to generate motoneurons in the ventral neural tube.
Asunto(s)
Diferenciación Celular/genética , Sistema Nervioso Central/embriología , Regulación del Desarrollo de la Expresión Génica/genética , Secuencias Hélice-Asa-Hélice/genética , Neuronas Motoras/citología , Proteínas del Tejido Nervioso/genética , Células Madre/citología , Animales , Secuencia de Bases/fisiología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Tipificación del Cuerpo/genética , Sistema Nervioso Central/citología , Sistema Nervioso Central/metabolismo , Embrión de Pollo , ADN Complementario/química , ADN Complementario/aislamiento & purificación , ADN Complementario/metabolismo , Proteínas del Ojo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Inmunohistoquímica , Datos de Secuencia Molecular , Neuronas Motoras/metabolismo , Mutación Missense/fisiología , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box , ARN Mensajero/metabolismo , Ratas , Proteínas Represoras , Rombencéfalo/citología , Rombencéfalo/embriología , Rombencéfalo/metabolismo , Células Madre/metabolismoRESUMEN
Cytokines exert their activities in cell growth and differentiation by binding specific cell membrane receptors. Janus kinases (JAKs) are cytoplasmic protein tyrosine kinases that physically interact with intracellular domains of the cytokine receptors and they play crucial roles in transducing signals triggered by the cytokine-receptor interaction. We have previously shown that conditional activation of JAK through membrane-proximal dimerization confers cytokine-independence on interleukin-3 (IL-3)-dependent Ba / F3 lymphoid cells and that the cytokine-independent proliferation is completely inhibited by dominant negative Ras. In this work, we demonstrate that ectopic expression of a dominant negative form of Stat5, a major signal transducer and activator of transcription (STAT) expressed in Ba / F3 cells, also inhibits JAK-triggered mitogenesis. In contrast, overexpression of constitutively active Ras or conditional activation of Stat5 by chemical dimerization fails to confer cytokine-independence. However, concomitant activation of ectopic Ras and Stat5 molecules in Ba / F3 cells suffices for cell proliferation in the absence of IL-3. Our results indicate that Ras and STAT are essential and sufficient components of JAK-triggered mitogenesis. Our findings further indicate that the cytokine signal bifurcates into Ras and STAT pathways following JAK activation.
Asunto(s)
División Celular/fisiología , Proteínas de Unión al ADN/fisiología , Proteínas de la Leche , Proteínas Tirosina Quinasas , Proteínas/fisiología , Proteínas Proto-Oncogénicas p21(ras)/fisiología , Transducción de Señal , Transactivadores/fisiología , Animales , Línea Celular/efectos de los fármacos , Dimerización , Activación Enzimática , Vectores Genéticos , Ratones , Fosforilación , Factor de Transcripción STAT5 , TYK2 Quinasa , TransfecciónRESUMEN
A case of the Voerner type palmoplantar keratoderma was studied for abnormalities of keratinization parameters. An enzyme and materials used to build the marginal band or cellular envelope of the cornified cell were all abnormally expressed; i.e. transglutaminase I (TGK), loricrin, and involucrin were abnormally immunostained. In the normal controls, their expression was limited to the upper epidermis, mainly in the granular layer. In the lesional skin, they were detected from the suprabasal layer to the lower horny layer. Filaggrin, the protein of the keratohyalin granule, was also expressed more widely than in controls. Ultrastructural abnormalities included a significantly higher frequency of composite keratohyalin granules than controls, early formation of a marginal band in the midepidermis, and, most remarkably, the clumping of tonofilaments causing vacuolization of the cytoplasm of affected keratinocytes.
Asunto(s)
Queratinas/ultraestructura , Queratodermia Palmoplantar/patología , Piel/ultraestructura , Adulto , Femenino , Proteínas Filagrina , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Técnicas para Inmunoenzimas , Proteínas de Filamentos Intermediarios/metabolismo , Queratinas/metabolismo , Queratodermia Palmoplantar/metabolismo , Proteínas de la Membrana/metabolismo , Precursores de Proteínas/metabolismo , Piel/metabolismo , Transglutaminasas/metabolismoAsunto(s)
Melanoma Amelanótico/patología , Enfermedades de la Uña/patología , Adulto , Eritema/patología , Femenino , HumanosRESUMEN
Cytokines play crucial roles in the growth and differentiation of hematopoietic cells. They bind to specific cell membrane receptors that usually do not possess intrinsic protein-tyrosine kinase activity. Janus kinases (JAKs) are cytoplasmic protein-tyrosine kinases that physically interact with intracellular domains of the cytokine receptors and have been implicated in playing important roles in signal transduction triggered by the cytokine-cytokine receptor interaction. However, it is still uncertain whether JAK activation alone suffices to induce cell proliferation. In this work, we modified Tyk2, a member of the JAK family, by adding a membrane localization sequence and a chemical dimerizer (coumermycin)-dependent dimerization sequence. The modified Tyk2 was activated in a coumermycin-dependent manner, and the activated Tyk2 conferred cytokine independence upon interleukin-3-dependent pro-B lymphoid cells. This cytokine-independent proliferation was completely inhibited by dominant-negative Ras. These results indicate that activation of JAK through membrane-proximal dimerization is sufficient to induce cell cycle progression and that Ras is essentially involved in JAK-triggered mitogenesis.
Asunto(s)
División Celular/fisiología , Interleucina-3/fisiología , Proteínas Tirosina Quinasas/metabolismo , Proteínas/metabolismo , Proteínas Proto-Oncogénicas , Aminocumarinas , Animales , Linfocitos B , División Celular/efectos de los fármacos , Línea Celular , Membrana Celular/fisiología , Cumarinas/farmacología , Dimerización , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Interleucina-3/farmacología , Janus Quinasa 1 , Janus Quinasa 2 , Janus Quinasa 3 , Ratones , Modelos Biológicos , Regiones Promotoras Genéticas , Proteínas Recombinantes de Fusión/metabolismo , TYK2 Quinasa , TransfecciónRESUMEN
To achieve strictly on-off switching of a desired cDNA expression in a broader range of mammalian cell types, we introduced the Escherichia coli lac operator sequences into the tetracycline-repressible promoter and created a chimeric promoter (designated here as the TcIP promoter) whose activity was reciprocally controlled by tetracycline and isopropyl thiogalactopyranoside (IPTG). cDNAs were connected downstream of the TcIP promoter and stably transfected into interleukin (IL-2 or IL-3)-dependent lymphoid cells that ectopically coexpress the tetracycline-repressible transactivator and the lac repressor. Whereas the parental tetracycline-repressible promoter exhibited constitutive activities when stably introduced into the lymphoid cells, cDNA expression from the TcIP promoter was strongly inhibited by tetracycline and was potently induced by IPTG in stable transfectants. Hence, the TcIP promoter made it possible to achieve highly controlled cDNA expression in cells wherein the parental promoter did not function in an inducible manner. Potential application of this promoter was provided by expressing cyclin-dependent kinase inhibitor, p27(Kip1). Induced expression of p27(Kip1) by the TcIP promoter in the lymphoid cells strongly reduced cellular responsiveness to IL-2 or IL-3, consistent with the idea that p27(Kip1) is a critical target that must be inactivated by the interleukin-triggered mitogenic signals.
Asunto(s)
Proteínas Bacterianas/metabolismo , Proteínas de Ciclo Celular , Proteínas de Escherichia coli , Regulación de la Expresión Génica/genética , Proteínas Represoras/metabolismo , Tetraciclina/farmacología , Transactivadores/metabolismo , Proteínas Supresoras de Tumor , Animales , División Celular , Línea Celular , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , ADN Complementario , Interleucina-2/metabolismo , Interleucina-3/metabolismo , Isopropil Tiogalactósido/farmacología , Represoras Lac , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Plásmidos , Regiones Promotoras Genéticas , TransfecciónRESUMEN
BACKGROUND: Since the inception of the Charity Hospital Tumor Registry in 1948, 80 cases of malignant melanoma in blacks were treated at the Tulane University School of Medicine, Department of Surgery. Among black people, melanoma occurs on acral dermal sites. The histologic type is primarily acrallentiginous melanoma (ALM), found on acral, volar-subungual skin and junctional mucocutaneous sites. STUDY DESIGN: The registry records of 80 black patients with malignant melanoma were reviewed. The clinical data for 41 female patients were compared to those of 39 male patients. These data were analyzed according to the sex of the patient as well as the histologic type, site, and stage of disease at diagnosis. RESULTS: Among women, 44 percent of primary lesions were found on extradermal sites compared with only 10 percent among men. Only 32 percent of primary lesions among women were located on the foot, whereas 73 percent of the primary lesions in men were found on the foot. Of the seven patients with vulvar, cervical, and vaginal melanoma, none lived more than two years after diagnosis. Two female patients with anorectal melanoma succumbed to their disease within 22 months. However, 50 percent of the female patients with head and neck lesions and 75 percent of those with eye lesions lived more than five years. Forty and 26 percent of the female patients with limb lesions lived five and ten years, respectively. CONCLUSIONS: Black females have a higher rate of extracutaneous melanoma than black men or white men and women, which accounts for a distinct negative impact on survival rates among black women with melanoma. In addition, the worst prognosis of melanoma among black women is not entirely related to delays in diagnosis, as has been suggested, but to their higher rates of extracutaneous melanoma.
Asunto(s)
Población Negra , Melanoma/etnología , Neoplasias Cutáneas/etnología , Adolescente , Adulto , Anciano , Niño , Preescolar , Extremidades , Femenino , Neoplasias de los Genitales Femeninos/etnología , Neoplasias de los Genitales Femeninos/mortalidad , Neoplasias de los Genitales Femeninos/patología , Neoplasias de Cabeza y Cuello/etnología , Neoplasias de Cabeza y Cuello/mortalidad , Neoplasias de Cabeza y Cuello/patología , Humanos , Lactante , Recién Nacido , Masculino , Melanoma/mortalidad , Melanoma/patología , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Sistema de Registros , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/patología , Tasa de SupervivenciaRESUMEN
The in-gel competitive reassociation (IGCR) procedure was successfully applied to construct a comprehensive library enriched in DNA fragments containing C5mCGG sequences from mouse liver and brain genomic DNA. For IGCR, methylation-insensitive restriction enzyme (Msp I) digests were used as target DNA and methylation-sensitive restriction enzyme (Hpa II) digests as competitor DNA. Southern blot analysis indicated that 60 to 70% of the clones in the library were derived from the methylated sites and overall enrichment was 200- to 1000-fold. IGCR was further applied to construct a library for the sites differentially methylated between brain and liver DNA. In the library, approximately 20% of the Hpa II sites exhibited different degrees of methylation between these tissues.
Asunto(s)
Metilación de ADN , ADN/genética , Biblioteca de Genes , Animales , Sitios de Unión , Encéfalo/metabolismo , Clonación Molecular/métodos , Fragmentación del ADN , Genoma , Hígado/metabolismo , RatonesRESUMEN
A high pigment-producing strain of cultured Euphorbia millii cells was isolated by clonal selection. The pigment obtained was red and consisted mainly of anthocyanin. The amount of this pigment obtained after 24 selections was seven times that found in the original cells. Statistical and cell-pedigree analyses proved that this cell strain has stable productivity for this red pigment.
RESUMEN
We induced calluses from two Euphorbia species and analyzed the lipids and pigments of their cells. Growth was promoted when malt extract was added to the medium for callus induction. The lipid constituents of both E. tirucalli and E. millii calluses were the same; sitosterol, stigmasterol, campesterol, palmitic acid and linoleic acid. In addition, an anthocyanin, cyanidin glycoside, was isolated from callus that had been induced from E. millii leaves cultured on medium containing 0.1 ppm 2,4-D.